Journal of Histopathology and Cytopathology 2020, July

Journal of Histopathology and Cytopathology

Official Organ of Bangladesh Academy of Pathology

Vol 4, No 2, July 2020

Front Cover PDF

Index/Contents PDF

Inside Back cover PDF


Sl No Article Title Page no
1 Histopathological Findings in Fatal COVID-19 Infections
Talukder SI
  Original Contributions  
2 Intratumoral and Peritumoral Angiogenic and Lymphangiogenic Microvessel Density in Invasive Breast Carcinoma and their Correlation with Lymph Node Metastasis
Mehjabin M,Asaduzzaman, Chakravarty S,Hassan MI, Munmun UK,Islam N,Talukder AS, Kamal M
3 Immunohistochemical Analysis and Molecular Subtyping of Breast Cancer
Sharmin S,  Ambiya AS,  Hussain M, Rima FAPDF
4 Diagnosis of Hirschsprung Disease by Frozen Section Biopsy Using Routine Hematoxylin-Eosin (HE) Stain: A Year’s Study
Banu SG, Islam T
5 Histomorphology of Gastroesophageal Junction Lesions (GEJ) and their Malignant Potential in Gastroesophageal Reflux Disease (GERD): A Study of 145 Cases in a Tertiary Level Hospital in Bangladesh
Zabin SG, Dewan RK, Jinnah SA, Jeba R, Sultana T, Rahman LY, Khan ZB
6 E-Cadherin Expression in Transitional Cell Carcinoma of the Urinary Bladder and its Correlation with Histopathological Grade and Tumour Stage
Quruni MO, Saha MK, Afrin SS, Hossain MS, Shaheen N, Dewan RK
7 Extracellular Matrix Metalloproteinase Inducer (EMMPRIN/CD147) Expression and its Correlation with Progression of Oral Squamous Cell Carcinoma
Saha MK,Quruni MO, Afrin SS, Hossain MS,Mahmud  F,  Mahmud SA, Jinnah MA, Dewan MR
Review Article
8 The Many Faces of Focal Segmental Glomerulosclerosis: A Review
  Case Reports  
9 Primary Malignant Melanoma of the Breast: A Case Report
Sadaf A, Hossain MI, Sultana N, Khan AS
10 Information for Contributors


Information for Contributors

Information for Contributors


General Information

The Journal of Histopathology and Cytopathology (JHC) aims in our understanding of the pathophysiological and pathogenetic mechanisms of human disease by publishing original papers, review articles, case reports and short communications related to basic and translational fields in pathology. It serves as bridges between basic biomedical science and clinical medicine with particular emphasis on, but is not restricted to, tissue based studies only. It is published twice a year as the Journal Committee of the Bangladesh Academy of Pathology.

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Manuscripts should be prepared in MS Word format in accordance with The Uniform Requirements for Manuscripts Submitted to Biomedical Journals

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It should begin with full title of the article. Do not write authors name in the abstract page. The abstracts should not be more than 200 words. The abstract should state the purpose of the study or investigations, basic procedures, main findings and principal conclusion. Three to ten keywords may be provided below the abstract using terms from the Medical Subject Headings (Index Medicus, NLM, USA). Abbreviations and citations should be avoided.


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Journals: van Riel D, Leijten LM, Kochs G, Osterhaus AD, Kuiken T: Decrease of Virus Receptors during Highly Pathogenic H5N1 Virus Infection in Humans and Other Mammals. Am J Pathol 2013, 183:1382-1389

 Electronic Journals: Moher D, Liberati A, Tetzlaff J, Altman DG, The PRISMA Group: Preferred Reporting Items for Systematic Reviews and Meta-Analyses: The PRISMA Statement. PLoS Med 2009, 6:e1000097.

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On receipt, manuscripts are assessed by the Editor-in-Chief, to one Associate Editor. The Reviewers’ and Associate Editor’s views are used by the Editor-in-Chief (or a Senior Editor) in reaching a decision, usually within three weeks of submission.

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Research articles are divided into sections with the headings: Abstract, Introduction, Methods, Results and Discussion. Long articles may need subheadings (especially within the Results and Discussion) to clarify their content. The sections should not be numbered. Other types of articles, such as reviews and commentaries, still need a title and abstract and should adhere as closely as possible to these guidelines.

Epidermal Nevus with Epidermolytic Hyperkeratosis: A Case Report

Epidermal Nevus with Epidermolytic Hyperkeratosis: A Case Report


*Fatima K,1 Banu SG,2 Kamal M3


  1. *Dr. Kaniz Fatima, Resident, Phase-B, Department of Pathology, Bangabandhu Sheikh Mujib Medical University (BSMMU)
  2. Sultana Gulshana Banu, Associate Professor, Department of Pathology, Bangabandhu Sheikh Mujib Medical University (BSMMU).
  3. Mohammed Kamal, Professor, Department of Pathology, Bangabandhu Sheikh Mujib Medical University (BSMMU).





Epidermal nevus has various histological patterns. Epidermal nevus having features of epidermolytic hyperkeratosis is a rare condition. This lesion is clinically indistinguishable from other epidermal nevi but shows characteristic histological features. Diagnosis of this entity is important management. Epidermolytic hyperkeratosis is an autosomal dominant disease. Offspring of these patients may have generalized epidermolytic hyperkeratosis. Here we present a case of 12 years old boy with Epidermal nevus with epidermolytic hyperkeratosis, which is a rare entity.

[Journal of Histopathology and Cytopathology, 2020 Jan; 4 (1):65-69]

 Keywords: Epidermal nevus, Epidermolytic hyperkeratosis, Generalized epidermolytic hyperkeratosis, Line of Blaschko.


Epidermal nevus comprises a heterogenous group of diseases. It is a congenital non-inflammatory cutaneous hamartoma. It may occur sporadically or as a part of several syndromes. The condition affects 1 in 1000 people in the world.1 Histologically, epidermal nevi are composed of keratinocytes, apocrine glands, eccrine glands, sebaceous glands or other components of pilosebaceous unit. Epidermal nevi are traditionally asymptomatic. Small number of cases of epidermal nevi show histologic features of epidermolysis hyperkeratosis. Epidermolytic hyperkeratosis has been observed in variety of benign and malignant skin condition or hereditary disorders. Epidermal nevus with epidermolysis hyperkeratosis has a significant clinical importance. This patient carries the risk of parenting a child of gerneralizedepidermolytichyperkeratosis.2 We report a case of epidermal nevus showing epidermolytic hyperkeratosis in a 12 year old boy for the rarity of the entity.

 Case report

A 12 year old boy of a non-consanguineous parentage, presented with non-pruritic, dark coloured elevated skin eruptions since birth. The lesion was first observed over the dorsum of right foot. After that lesions gradually appeared in front of leg and thigh, lower abdomen and flexor aspect of both forearm. Lesions were not related with any seasonal variation. Patient had a normal birth history and developmental milestones. Right sided extremities were more involved than left side.Examination revealed numerous hyperpigmented warty papules distributed in both extremities and lower abdomen (Figure. 1 and 2).  Hair, nails and oral mucosa were normal. Other system examination revealed no abnormality. No laboratory investigation was done.

Clinically it was diagnosised as Linear verrucous epidermal nevus. For histopathological examination 3 mm punch biopsy was taken from right foot. Histological examination revealed hyperkeratosis, acanthosis, papillomatosis, elongated rete ridges. The dermis revealed mild perivascular infiltration of chronic inflammatory cells.Some foci also revealed perinuclear vacuolization of the keratinocytes in spinous and granular layers, and increased number of keratohyalin granules in the stratum granulosum (Figure 3, 4 and 5).  So, histologically it was diagnosed as epidermal nevus with epidermolytic hyperkeratosis.


Epidermal nevus is hamartoma of skin, occurs due to over growth of epidermis. It arises from embryonic ectoderm as a result of mosaic postzygotic mutations. Lesions are present at birth in about half of the patients or may develop early in childhood. Depending on the affected component of the epidermis epidermal nevus can be divided into two types: keratinocytic or non organoidand organoid type.3 Keratinocytic epidermal nevus is the most common type of epidermal nevus. It occurs due to overgrowth of keratinocytes. Different varients of keratinocytic epidermal nevus are seen, such as linear epidermal nevus, hard nevus of Unna, soft epidermal nevus and nevus verrucosus etc.1 On the other hand organoid type shows predominantly another component of skin.4 Epidermal nevus occurs as a result of activated genetic mutation in FGFR-3, HRAS or PIK3CA genes. FOXN1 is highly expressed in these lesions.1 Most common pattern of keratinocytic nevus is linear epidermal nevus. The lesions are verrucous, skin-coloured dirty gray or brown coloured papule, which coalesce to form serpiginious plaques. They follow the line of Blaschko. These lines are thought to be representative pathways of epidermal cell migration and proliferation during development of fetus.5

Linear epidermal nevus may be either localized or systematized. In localized type, only one linear lesion is present and lesion is confined to one side of the body. Common sites are head, trunk and extremities. In systematized type there are many parallel linear lesions are seen. They may be unilateral or bilateral.

Localized and more commonly systematized linear epidermal nevus may be associated with skeletal deformity and CNS deficiency.6 Rarely squamous cell carcinoma or basal cell carcinoma may arise in epidermal nevus.7

Epidermal nevus may occur as a part of epidermal nevus syndrome and may be associated with internal manifestation. These syndromes have characteristic cutaneous findings and at times relevantly specific internal findings.1  The six different types of epidermal nevus syndromes are nevus sebaceous, CHILD (congenital hemidysplasia with ichthyosiform erythroderma and limb defects) syndrome, nevus comedonicus, Becker’s nevus, Proteus syndrome, phacomatosispigmentokeratotica.

Histologically epidermis of epidermal nevus is hyperplastic. There is variable hyperkeratosis, papillomatosis and acanthosis with elongation of rete ridges. Upto 62% of biopsy specimens have these pattern and these are called non-epidermolytic epidermal nevus. About 16% of epidermal nevi show features of epidermolytic hyperkeratosis. Other histologic patterns are psoriatic type, acrokeratosisverruciformis like type and a Darier’s disease like type.1 Epidermolytic hyperkeratosis is more common in systematized type than localized type. This reaction pattern of skin was first described by Ackerman in 1970.7It occurs due to defective keratin genes (KRT-1 and KRT-2), which causes excessive and abnormal keratinization. The salient histologic features are- compact hyperkeratosis, perinuclear vacuolization of the cells in stratum malpighii, irregular cellular boundaries and increased numbers of large irregular keratohyaline granules. Epidermolytic hyperkeratosis is seen in other conditions, such as- bullous congenital icthyosiformerythroderma, icthyosisbullosa of Siemens, Vorner’spalmoplanterkeratoderma, melanocytic nevus, epidermolyticacanthoma, basal cell carcinoma and squamous cell carcinoma etc.

Main clinical differential diagnosis of epidermal nevus areepidermodysplasiaverruciformis, inflammatory linear verrucous epidermal nevus, linear psoriasis and lichenstriatus.Epidermodysplasiaverruciformis is a genetic disease characterized by HPV infection. This is usually associated with HPV 5 or 8, less commonly 3, 4, 5.1 Histologically this lesion is characterized by acanthosis, hyperkeratosis, large cells with blue-gray cytoplasm, often with dysplastic change and irregular granular layer with rare perinuclear halo. Inflammatory linear verrucous epidermal nevus (ILVEN) is a type of epidermal nevus. These lesions are also distributed in linear pattern, but they are erythematous and itchy. Histologically ILVEN is characterized by some specific features, which are absent in epidermal nevus. These are- areas of depressed orthokeratosis with underlyinghypergranulosis, alternating areas of slightly raised parakeratosis with underlying hypogranulosis.8 In linear psoriasis, the lesions may be pink to red papules or silvery scales. Typical histologic findings are regular elongation of rete ridges, thin suprapaillary plates, hypo or agranulosis, Munro microabcess and spongiform pustules of Kogoj, which are absent in epidermal nevus. In lichen striatus, the lesions are erythematous papules arranged in linear pattern following lines of Blaschko. But histologically it differs from epidermal nevus by presence of vacuolar alteration of basal layer and band like lymphocytic infiltrate.7 Treatment modalities of epidermal nevus are topical cream, cryotherapy, laser and dermabrasion. But management is difficult, because the lesions  recur unless treatment extend into dermis.


Epidermal nevus with epidermolytic hyperkeratosis is a rare condition. As this is a mosaic genetic disorder of suprabasal keratin, it can be transmitted to offspring and produce generalized epidermolytic hyperkeratosis. So genetic counseling is essential for these patients.


  1. James W, Berger T, Elston D, Neuhaus I. Andrew’s Diseases of the Skin. 12th ed. Philadelphia: ELSEVIER. 2016. pp 625-26
  2. Guite Z, Pamei D, Gunto H, Das K. Epidermolytic hyperkeratosis in verrucous epidermal nevus. Journal of medical society. 2014; 28(1): 47-8.
  3. Pollozhani N, Damevska K, Silvija D, Adjievska N, Gocev G. Epidermolytic hyperkeratosis: clue for diagnosis. Global Dermatology.  2017; 4(1): 1-3.
  4. Ngan V (2003). All about the skin. Retrieved from DermNet NZ website.
  5. Kouzak SS, Mendes MS, Costa IM. Cutaneous mosaicisms: concepts, patterns and classifications. An Bras Dermatol. 2013;88(4):507-517.
  6. Edler D. in Lever’s Histopathology of the Skin. 10th Philadelphia: LIPPINCOTT WILLIAMS & WILKINS; 2009. pp: 791-92
  7. Das A, Podder I, Das A, Ghosh A, Shome K. Epidermolyticblaschkoidverrucous epidermal nevus: Report of two cases. Indian J DermatopatholDiagnDermatol. 2015; 2:46-8.
  8. Meibodi NT, Nahidi Y, Jaridi Z. Epidermolytic hyperkeratosis in inflammatory linear verrucous epidermal nevus. Indian J Dermatol. 2011; 56:309-12.


Papillary Thyroid Carcinoma Arising within Mature Ovarian Teratoma: A Case Report

Papillary Thyroid Carcinoma Arising within Mature Ovarian Teratoma: A Case Report

Sadaf A, 1 Hossain MI,2  Sultana  N,3 Nasreen S,4 Rahman Z5


  1. *Dr. Anika Sadaf, MD. (Pathology) Phase B, Resident, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh, 4203.
  2. Mohammad Ismail Hossain. Lecturer, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh, 4203.
  3. Nahid Sultana. Senior Consultant, Obstetrics & Gynaecology. 250 Bedded General Hospital,Chandpur, Bangladesh, 3600.
  4. Sayeeda Nasreen. Assistant Professor, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh, 4203.
  5. Zillur Rahman. Professor, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh, 4203.

 *For correspondence


Mature cystic teratoma is the commonest ovarian germ cell tumor. Though malignant transformation is uncommon, papillary thyroid carcinoma has rarely been described as associated with ovarian teratomas. We report a case of a 34-years old multiparous woman who presented with acute abdominal pain and an ovarian mass. After salphingo-oophorectomy, the patient was diagnosed as papillary thyroid carcinoma that arose within a mature cystic ovarian teratoma. To our knowledge, this is the first reported case of papillary thyroid carcinoma arising within a mature ovarian teratoma in this tertiary health care center in Chattogram. We recommend long term follow up to see any metastatic possibility.

 [Journal of Histopathology and Cytopathology, 2020 Jan; 4 (1):60-64]

Keywords: Mature cystic teratoma, papillary thyroid carcinoma, struma ovarii.


Among the ovarian germ cell tumors Mature Cystic Teratoma (MCT) is the most common and comprises 10–20% of all ovarian tumors. However, malignant transformation of MCT is not common and the incidence is 1–3%.1 Squamous Cell Carcinoma (SCC) is the commonest type, found in 80% of cases.2 Papillary thyroid carcinoma (PTC) within teratoma is one of the rarest types with ranges varying from 0.1% and 0.2%, and usually diagnosed postoperatively.3 The synchronous development of malignant struma ovarii and primary thyroid carcinoma is extremely rare, though a handful of cases were reported.4 So, presence of thyroid tissue in teratoma, should proceed with further work up to confirm the diagnosis and to explore the possibility of a malignant lesion in the mass- either primary or metastasis.5 Here we present a case of a patient with a PTC arising within a ovarian MCT.

Case Presentation
34-year-old multiparous woman who had abdominal pain, distention and irregular menstrual bleeding for approximately for 6 months, presented to the emergency service of Chittagong Medical College Hospital in May 2019 with the complaints of a sharp pain in lower abdomen, with accompanying vomitting. There was no previous medical or surgical history.

On abdominal examination, a tender mass adjacent to the left side of the umbilicus were detected. Manual examination of the vagina revealed tenderness and mass in left adnexal region. Paps smear was done with no abnormality detected. Haemogram and biochemical test results were normal except a hemoglobin level of 9.2 gm/dl. CA-125 was within normal limit and β-HCG was normal. Ultrasonography of the lower abdomen revealed a complex solid cystic mass measuring 82x62x55 mm, with mixed echotexture, compatible with dermoid cyst.

At laparotomy, a cystic mass of approximately 9x7cm size, with a white, smooth glistening surface, originating from the left ovary was observed. Opposite ovary was apparently normal and no adhesion or intra-abdominal deposit was observed. Left sided salphingo-oophorectomy was performed preserving the uterus and right ovary and sent to Department of Pathology, Chittagong Medical College, Chattogram for histopathological evaluation. On gross pathological examination, a cystic mass of 9 cm in diameter (fig-1) with 3cm fallopian tube were noted. On cross section, hair, sebum & fatty materials were come out and some thick greenish fluid was drained and some solid structure was observed on its wall (fig-2).

Microscopic examination revealed mature teratomatous component represented by skin with associated adnexal structures, muscles, fat, benign glands lined by mucin containing columnar epithelium (fig-3,4) and thyroid tissue, within the thyroid tissue foci papillary thyroid carcinoma (fig-5,6) was found. Lining cells had oval nuclei showing nuclear overlapping, grooving and intranuclear cytoplasmic inclusions. Follicles also contained amorphous eosinophilic thick colloid. The fallopian tube was unremarkable. Immunohistochemical (IHC) examination revealed positivity for TTF-1 (fig-8,9). With these findings, diagnosis of a “mature cystic teratoma with malignant transformation to papillary thyroid carcinoma” was made. Postoperatively plasma levels of T3, T4, TSH and thyroglobulin of the patient were normal. A normal parenchymal vasculature was identified by postoperative ultrasonography of the thyroid gland.


Mature cystic teratomas are also known as dermoid cysts, because they are mostly cystic, skin & skin appendages are the most common structures. Thyroid tissue is present in 10% of the all cases.6 Teratomas containing more than 50% of thyroid tissues are called struma ovarii, often presented as monodermal teratoma.7 Malignant transformation of MCT is rare; however, several types of malignancy can develop from any one of three germ-cell layers. Squamous cell carcinoma, derived from ectoderm is the commonest type; less common malignancies include soft tissue sarcomas, adenocarcinomas, malignant melanomas, basal cell carcinomas, carcinoid tumors, and thyroid carcinomas.2 Among thyroid carcinomas the most common histological type is the papillary carcinoma (44%), other types are follicular carcinoma (30%) and follicular variant of papillary carcinoma (26%).3

 The malignant change of an initially benign cystic teratoma is detected in patients between 40 and 60 years of age, older than its benign counterpart. Although the cancer occurs at any age, most patients are postmenopausal.1 The tumor may present as pelvic discomfort, with a pelvic mass on abdominal imaging (USG, CT, MRI) or during laparotomy for any other reason. Preoperative definitive diagnosis of stroma ovarii or papillary thyroid carcinoma is not possible. The only possibility of preoperative diagnosis is by radioactive iodine scan (not done routinely).8 Various case reports have been published over the past few years regarding the histological diagnoses and treatment options. The diagnoses of thyroid carcinomas arising in teratomas should be made following the guidelines for diagnosing carcinomas in thyroid gland. Disease is treatable with good out come in most cases. Only 7% and 14% of patients with papillary carcinoma and typical follicular carcinoma, respectively died of disease. Due to rarity of disease no consensus on treatment has been made, however treatment options include oophorectomy, additional thyroidectomy, radioactive iodine and long term follow up with serum thyroglobulin measurement.9

In order to determine metastatic disease, in MCT cases undergoing malignant transformation, follow up of thyroglobulin (Tg) levels is recommended. The only source of circulating Tg is the thyroid tissue and ovarian teratomas containing thyroid tissue, which is a very rare condition. However, high Tg level in benign thyroid diseases hamper determination of it as a convenient tumour marker in MCT, who did not undergo thyroidectomy and who contain thyroid tissue with malignant transformation. On the other hand, the high levels of anti-thyroglobulin antibody (anti-Tg Ab) may cause Tg levels to be erroneously low. For this reason, the follow-up of Tg levels is favourable for patients, who underwent thyroidectomy only and for patients left with no or very little thyroid tissue. In order to evaluate Tg levels correctly, follow-up of Tg levels together with anti-Tg Ab levels is advisable as persisting high levels of anti-Tg Ab indicate a persistent disease.3 In our case, Plasma T3, T4, TSH and Tg level were normal and normal parenchymal vasculature was identified by ultrasonography of the thyroid gland. Anti-Tg Ab level can’t be performed due to patient’s refusal. Logani et al., 2001 was commented the absence of normal thyroid tissue and features of teratoma, in favour of a metastatic lesion originating from thyroid gland.10 In the presented case, histologic evidence of mature teratoma, normal thyroid tissue along foci of papillary thyroid carcinoma, and positive immunohistochemical stain for thyroid transcription factor-1(TTF-1) indicates primary thyroid carcinoma arising within MCT.


Whether further therapy with total thyroidectomy and radioiodine ablation may be beneficial is unknown. The rarity of MCT cases undergoing PTC transformation impedes the establishment of a protocol for treatment and follow-up. We recommend that a long-term follow-up of these cases is needed to know more about the prognosis and to see any local recurrence or metastasis.


  1. Rim SY, Kim SM, Choi HS. Malignant transformation of ovarian mature cystic teratoma. Int J Gynecol Cancer. 2006; 16:140–44.
  2. Pineyro MM, Pereda J, Schou P, Santos DL, Peña SDI, Caserta B,Pisabarro R. Papillary thyroid microcarcinoma arising within a mature ovarian teratoma: case report and review of the literature. Clinical Medicine Insights: Endocrinology and Diabetes. 2017; 10: 1–3.
  3. Cokmez H,Gulbahar A, Yigit S, Aydin C. Oncocytic and tall columnar type papillary thyroid carcinoma arising on a mature cystic teratoma: A case report and literature review. J Pak Med Assoc. 2019; 69:116-19.
  4. Tzelepis EG, Barengolts E, Garzon S, Shulan J, Eisenberg Y. Unusual case of malignant strumaovarii and cervical thyroid cancer preceded by ovarian teratoma: case report and review of the literature. Hindawi: Case Reports in Endocrinology. 2019 Mar 17; 1-7.
  5. Yeasmin S. A case of papillary thyroid cancer and extraovarian pelvic Teratoma.Journal of the Endocrine Society. 2019; 3(1). available at:
  6. Bedir R,Yılmaz R. Coexistence of papillary thyroid cancer and hashimoto’sthyroiditis developing within an ovarian mature cystic teratoma. Journal of Mid-life Health. 2019 April 10; 10: 45-47.
  7. ParulskaES, Pioch A, Chyrek EC, Wolinski K, Jurczyszyn DJ, Jedynska MJ, Majewski P, Zabel M,Ruchala M. The role of immunohistochemical examination in diagnosis of papillary thyroid cancer in strumaovarii. Folia Histochemica Et Cytobiologica. 2019;57(1):35–42.
  8. Naeem M, Iqbal M, Imran MB,Tabassum R. Malignant strumaovarii: a rare case report. European Journal of Medical Case Reports. 2017; 2(1): 30-32.
  9. Haider A, Hussain M, Hassan U, Loya A. Papillary thyroid carcinoma arising in ovarian teratomas: A report of three cases. Journal of Islamabad Medical & Dental College (JIMDC). 2015 Sep 27; 4(2): 88-90.
  10. Logani S, Baloch ZW, Snyder PJ, Weinstein R, LiVolsi VA. Cystic ovarian metastasis from papillary thyroid carcinoma: A case report. Mary Ann Liebert, Inc. 2001; 11(11): 1073-1075.


Histopathological Diagnosis of Rhinofacial Entomophthoramycosis in a 16-Year-Old Girl: A Case Report

Histopathological Diagnosis of Rhinofacial Entomophthoramycosis in a 16-Year-Old Girl: A Case Report

*Asaduzzaman,1 Khandkar T,2 Rahman DA3 


  1. *Dr. Asaduzzaman, Assistant Professor of Histopathology, Sheikh HasinaNational Institute of Burn and Plastic Surgery, Dhaka, Bangladesh.
  2. Tahmina Khandkar, Assistant Registrar, Paediatric Nephrology, National Institute of Kidney Diseases and Urology, Shere-E-Bangla Nagar, Dhaka
  3. DM Arifur Rahman, Assistant Professor, Department of Pathology, TMSS Medical College, Bogura

 *For correspondence


Rhinoentomophthoramycosis is an uncommon and severely disfiguring disease. It mainly involves the mucosa of the nares, nasal passages, nasal sinuses, nasopharynx, mouth and spreads to adjacent tissues causing disfigurement of face. Histopathological examinations and mycological cultures are the gold standard for confirmation of entomophthoramycosis. We report a case of a 16-year-old girl who presented with swelling and ulcer of face. Clinical presentation along with typical histopathologic findings were diagnostic in this case.

 [Journal of Histopathology and Cytopathology, 2020 Jan; 4 (1):55-59]

 Keywords: Entomophthoramycosis, zygomycosis, fungal infection of face, fungal granuloma, splendore-Hoeppli reaction


Rhinoentomophthoramycosis is not so common in Bangladesh as well as other parts of the world. It is a grossly disfiguring disease. The medically important class zygomycetes are in two orders, the Mucorales and the Entomophthorales. Rhinofacialentomophthoralesmainly affects the mucosa of the nares, nasal passages, nasal sinuses, nasopharynx, mouth, and spreads to adjacent tissues causing disfigurement of the face. It occurs predominantly in immunocompetent individuals and live as saprophytes in soil and decaying plant matter.1 Rhinofacialconidiobolomycosis affects the subcutaneous tissues of the face, especially the paranasal sinuses as well as the deeper organs.2 We report the case of a teen-aged female who presented to us with facial swelling and ulcer and was diagnosed by histopathology.

 Case Report

An immunocompetent16-year-old girl from Chittagong presented to the outpatient department of Sheikh Hasina National Institute of Burn and Plastic Surgery with a one year history of progressive nasal and maxillofacial swelling. Swelling of face started from the nasal bridge and gradually spread into the left side of the face. For the facial swelling she had received multiple treatments, including glucocorticoids and antibiotics. But for the last one month she developed multiple ulcers with purulent discharge over the swelling. The physical examination reveals an ulcer over upper part of left cheek and swelling over bridge of nose. Adjacent area revealed erythema, edema, and tenderness over the nasal dorsum and forehead, extending to the soft tissue around left eyes (Fig. 1). Initially she has undergone biochemical and radiological investigations.

Laboratory investigations included a haemoglobin of 11.6 gm/dl, total leucocyte count 12.36×109/L with a differential count within normal range, and platelet count was 400.1x 109/L. Other biochemical tests were within normal limit. Serology for hepatitis B surface antigen and human immunodeficiency virus 1 and 2 were negative. MRI of face revealed diffuse soft tissue thickening involving the paranasal sinuses, skin and subcutaneous tissue in left side of face extending into the left zygomatoco-temporal region.  Direct naso-endoscopic examination revels left sided middle meatus. Septum and lateral wall of nose were congested. Middle meatus was adherent to the lateral wall of nose. At right side, crest of nose, middle meatus was distorted, nasal septum was absent/dehiscent on posterior part. There was no growth on nasopharynx. Biopsy specimens were obtained from multiple sites, including the forehead and the nose. The gross specimen consisted of two skin covered piece of tissue; largest one measured 4x3x1.5 cm and smaller one measures 1.5×0.8×0.5 cm. Skin surface showed multiple ulcers. The cut surface was solid and tan gray. Histopathological examination showed a chronic granulomatous inflammation (Fig. 2) with broad nonseptate branching hyphae surrounding amorphous eosinophilic substance, the Splendore-Hoeppli reaction (Fig. 3). Marked lympho-plasmacytic cell infiltrate with tissue eosinophilia and foreign body type of giant cells containing fungal elements were present. Periodic acid Schiff and Gomori-Methenamine-Silverstain highlighted the fungal elements and the surrounding amorphous eosinophilic material (Fig. 4 and 5). She was diagnosed as a case of Rhinofacialentomophthoramycosis. The patient was then on systemic antifungal therapy.


Rhinofacialentomophthoramycosis is an uncommon fungal infection; it mostly occurs in the tropical and subtropical regions of different parts of the world. G. Bras reported the first case of a Jamaican native in 1965. It is predominantly a chronic mucocutaneous and subcutaneous infection. The name Entomophthorales was coined from the Greek word “Entomon” meaning insect implicating their pathogenic nature in insects. Formerly, the two orders, namely Mucorales and Entomophthorales, were classified in the phylum Zygomycota. Hibbett et al. suggested a comprehensive phylogenetic classification of the kingdom Fungi, and the phylum Zygomycota was eliminated as a result of polyphyletic characteristics.3 Therefore, the taxa belonging to Zygomycota were distributed among the phylum Glomeromycota and four subphyla of uncertain placement (incertaesedis). Entomophthorales and Mucorales as well as two other orders (Kickxellales and Zoopagales) were raised to the rank of subphyla and renamed as EntomophthoromycotinaMucoromycotinaKickxellomycotina,and Zoopagomycotina.4 Entomophthoromycotina encompasses twogenera that cause human infection, Basidiolus and Conidiolus.

Humans suffering from rhinoentomophthoromycosis get infected by the attachment of conidia of Ccoronatus to nasal/sinusoidal mucosa. Initially, the disease presents like sinusitis.5 A nodule at the nostrils indicates expansion into the subcutaneous fat.6 The infection spreads within the subcutaneous fatty layers of the nasal bridge, eyelids, cheek, and upper lip. Swellings are firm, indolent, and, initially, often reddened and warm, while later they are often itchy.7 Mucosal swellings rarely affect laryngeal structures or cause dyspnoea. Ulcerations of skin or mucosa may occurs, as we found in our case. Skin-adherent structures, eye motility, and vision usually remain unaffected; and bones, vessels, muscle, and lymph nodes are rarely involved. The course of the disease is usually benign.8

The diagnosis is based on a combination of mycologic and histopathological tests, and clinical presentation.Histological examinations and mycological cultures are the gold standard for confirmation of entomophthoromycosis. Biopsy of skin lesions is preferred for diagnosis than pus, as the chances of positive identification with potassium hydroxide preparation and culture are better with tissue specimens.9 Entomophthoromycosis can be easily differentiated from other fungi by their characteristic hyphal morphology. The hyphae are broad, aseptate, or sparsely septate, with right-angle branching.10 The histological inflammatory reaction shows infiltration with lymphocytes, plasma cells, epithelioid cells, multinucleate giant cells, and histiocytes with an area of central necrosis that is surrounded by eosinophilic infiltration. This phenomenon is called Splendore–Hoeppli phenomenon.10 Our patient had all these typical features. PAS stain and GomoriMethenamine-Silver (GMS) stains are useful to demonstrate the fungal hyphae. Examination under fluorescent microscopy using fluorescent dye (Blankophor) wet mount preparation increases the sensitivity of diagnosis.11 Definitive diagnosis requires culture, polymerase chain reaction testing, and immunohistochemistry.

Treatment for endomophthoromycosis ismedical and surgical. Systemic antifungal therapy and or surgical debridement is the primary choice in most cases. Several antifungal agents are used for the treatment of endomophthoromycosis such as itraconazole and amphotericin B.12


The entomophthoromycosis is a severe fungal disease that can affect both immunocompetent and immunocompromised individuals. Despite the clinical features, the disease requires biopsy for diagnosis, as histological examinations and mycological cultures are the gold standard for confirmation of entomophthoramycosis.This disease have a favorable prognosis if early treatments can be ensured.


  1. Manning RJ, Waters SD, Callaghan AA. Saprotrophy of Conidiobolus and Basidiobolus in leaf litter. Mycol Res. 2007; 111: 1437–1449.
  2. Prabhu RM, Patel R. Mucormycosis and entomophthoramycosis: A review of the clinical manifestations, diagnosis and treatment. ClinMicrobiol Infect 2004;10 Suppl 1:31-47.
  3. Hibbett DS, Binder M, Bischoff JF, Blackwell M, Cannon PF, Eriksson OE, et al. A higher-level phylogenetic classification of the fungi. Mycol Res 2007;111:509-47.
  4. Kwon-Chung KJ. Taxonomy of fungi causing mucormycosis and entomophthoramycosis (zygomycosis) and nomenclature of the disease: Molecular mycologic perspectives. Clin Infect Dis 2012;54Suppl 1:S8-15.
  5. Choon SE, Kang J, Neafie RC, Ragsdale B, Klassen-Fischer M, Carlson JA. Conidiobolomycosis in a young Malaysian woman showing chronic localized fibrosingleukocytoclasticvasculitis: a case report and meta-analysis focusing on clinicopathologic and therapeutic correlations with outcome. Am J Dermatopathol. 2012; 34: 511–522.
  6. Choon SE, Kang J, Neafie RC, Ragsdale B, Klassen-Fischer M, Carlson JA. Conidiobolomycosis in a young Malaysian woman showing chronic localized fibrosingleukocytoclasticvasculitis: a case report and meta-analysis focusing on clinicopathologic and therapeutic correlations with outcome. Am J Dermatopathol. 2012; 34: 511–522.
  7. Martinson FD. Chronic Phycomycosis of the Upper Respiratory Tract: RhinophycomycosisEntomophthorae. Am J Trop Med Hyg. 1971; 20: 449–455.
  8. Ribes JA, Vanover-Sams CL, Baker DJ. Zygomycetes in Human Disease. ClinMicrobiol Infect. 2000; 13: 236–301.
  9. Chowdhary A, Randhawa HS, Khan ZU, Ahmad S, Khanna G, Gupta R, et al.Rhinoentomophthoromycosis due to Conidioboluscoronatus. A case report and an overview of the disease in India. Med Mycol 2010;48:870-9.
  10. El-Shabrawi MH, Arnaout H, Madkour L, Kamal NM. Entomophthoromycosis: A challenging emerging disease. Mycoses 2014;57Suppl 3:132-7.
  11. Kumar Verma R, Shivaprakash MR, Shanker A, Panda NK. Subcutaneous zygomycosis of the cervicotemporal region: Due to Basidiobolusranarum. Med Mycol Case Rep 2012;1:59-62.
  12. Prabhu RM, Patel R. Mucormycosis and entomophthoramycosis: A review of the clinical manifestations, diagnosis and treatment. ClinMicrobiol Infect 2004;10Suppl 1:31-47.



C3 Glomerulopathy: Overview on a New Disease Entity

C3 Glomerulopathy: Overview on a New Disease Entity


*Rahman DA,1 Banu SG2


  1. *Dr. DM Arifur Rahman, Assistant Professor, Pathology, TMSS Medical College, Bogura.
  2. Sultana Gulshana Banu, Associate Professor, Pathology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka.

 *For correspondence


The diagnosis of membranoproliferative glomerulonephritis (MPGN) has recently undergone change from an electron microscopy-based classification scheme to one based largely on immunofluorescence findings. Recent advances in our understanding of the disease pathology of membranoproliferative glomerulonephritis has resulted in its re-classification as complement C3 mediated glomerulopathy (C3G) and immune complex-mediated glomerulonephritis (IC-GN). The new concept is based on its underlying pathogenesis, with a key pathogenetic role for the complement alternative pathway (AP), rather than on histomorphological characteristics. This overview summarizes the current state of knowledge about the C3 glomerulopathy.

 [Journal of Histopathology and Cytopathology, 2020 Jan; 4 (1):48-54]

 Keywords: C3 glomerulopathy, Dense deposit disease, C3 glomerulonephritis, Membranoproliferative glomerulonephritis


C3 glomerulopathy (C3G) is an emerging kidney disease caused by dysregulation of the alternative complement pathway.1,2,3 The characteristic pathology of this disease is glomerular depositions of dominant C3 with absent or weak immunoglobulins. Therefore, C3G is basically diagnosed by immunofluorescence (IF) and it can reveal various patterns of glomerular injuries by light microscopy(LM).4,5 Following the recent trend of pathogenesis-based reclassification of glomerular diseases, glomerulonephritis associated with alternative complement dysregulation is collectively referred to as C3G.6 Because laboratory detection of alternative complement dysregulation is still uncommon in current practice, predominant C3 deposition by IF is an initial finding that suggests C3G. However, glomerular diseases caused by mechanisms other than alternative complement dysregulation may occasionally satisfy “C3-dominant deposition with scanty immunoglobulins” as stated in the current consensus report.4 Clearly, pathogenesis based classification in glomerular diseases is an important prospect for appropriate therapies, but the entity of C3G still presents dilemmas in diagnostic practice by lack of clear definition and pathogenic basis. We review the current status of C3 glomerulopathy, histological, immunofluorescence findings and treatment

MPGN and C3 Glomerulopathy

Understanding the limitations of current MPGN classification requires a brief review of complement activation pathways. There are two main pathways of complement activation: the classic pathway, which is activated when IgG or IgM antibodies bind to antigens; and the alternative pathway, which does not require the presence of antibodies and can be auto activated by spontaneous cleavage of C3 to C3b, leading to the formation of C3 convertase. The electron microscopy-based classification can result in overlap between types I and III. Both types have been considered to be immune complex-mediated glomerulonephritis but, observations suggest that some cases of MPGN type I or MPGN type III are mediated by complement, not immune complexes.7,8,9

So, the historical classification required modification. It is not based on pathogenesis and there is significant overlapping, which is described earlier. In recent years, there have been great advances in our understanding of the pathogenesis of MPGN, particularly in the area of complement-mediated C3 glomerulopathies, including DDD and C3 glomerulonephritis. 4,10

It is proposed that MPGN be classified into two major groups: immunoglobulin (Ig)-mediated and complement-mediated (C3G). If immunoglobulins are present on IF studies, the evaluation should include a work-up for infections, autoimmune diseases, and monoclonal gammopathies, including cryoglobulins. It should be kept in mind that Ig-mediated MPGN also is associated with extensive C3 (and C4) deposition along the capillary walls via activation of the classic pathway of complement. On the other hand, if the IF studies show predominantly C3 and are negative or show no significant staining for Igs, an in-depth study of the AP is warranted. Ig-mediated MPGN is more likely to be present in adults whereas complement-mediated MPGN is more likely to be present in children and young adults. It is likely that C3G noted in children and young adults is due to genetic mutations in complement-regulating proteins, whereas it is acquired in adults as a result of development of autoantibodies to complement-regulating proteins. Initial evaluation of AP should include serum MAC levels, an alternative pathway functional assay, and hemolytic complement assays. If the initial screening is positive, it should be followed by genetic analysis for mutations and enzyme-linked immunosorbent assays for the presence of autoantibodies to complement-regulating proteins.11-14

The current approach, therefore, distinguishes those forms of MPGN with isolated C3 deposits (including DDD and C3GN) as alternative complement pathway-mediated C3G from those cases of MPGN that are mediated by the classical complement pathway with deposits of Ig and complement.

Clinical Presentation

Membranoproliferative glomerulonephritis or C3G are rare diseases with an estimated incidence of 1–2 per million per total population.15  Patients with C3G present with a variety of symptoms, ranging from a mild disease with asymptomatic microhaematuria and/or proteinuria to a severe disease with nephritic or nephrotic syndrome and renal impairment. Renal survival was worse if the GFR at diagnosis is <60 ml/min/1.73 m2.16

The kidney is the major target, possibly due to the morphological specificities of the glomerular capillaries, in particular the fenestrated endothelium, with exposure of the glomerular basement membrane to serum (complement). Although low levels of C3 are considered a hallmark feature of C3G, in one study low C3 levels were only detected in about 50 % of the patients. Therefore, a normal C3 level does not rule out C3G.17

Nasr et al., 2009 studied 32 paediatric and adult patient of dense deposit disease and Lu., et al 2012 studied 92 children and adult patient of dense deposit disease. Both of the studies reveal at presentation, almost all patients have proteinuria usually with haematuria. Nephrotic-range proteinuria is present in two thirds of the patients. Full nephrotic syndrome in 12% to 65% in different series conducted by Lu et al., 2006 and Servais et al., 2012. Persistently, low serum levels of C3 are found in most patients (approximately 80%). Servais et al have reported the clinical features in 56 patients with C3 glomerulopathy without dense deposits (C3GN) and compared them with 29 patients with DDD and 49 patients with immune complex MPGN type 1. The mean age at diagnosis for C3GN was 30, which was significantly higher than for DDD; 25% of patients were below16 years of age. Twenty-seven percent of patients with C3GN had nephrotic syndrome at presentation as compared with 38% of patients with DDD and 65% of patients with MPGN type I.18,19,20

 Light Microscopy

Light microscopic findings in C3 glomerulopathy can range from membranoproliferative lesions to mesangioproliferative or endocapillary proliferative lesions with or without presence of crescents. In rare instances, light microscopy might be normal. The electron dense osmophilic deposits as seen characteristically in DDD are found within the glomerular basement membrane, and as rounded deposits in the mesangium. In many cases, deposits are also seen in Bowman’s capsule and tubular basement membranes. C3 glomerulopathy, in which deposits do not completely fulfill criteria for dense deposits, are classified as C3GN. Electron microscopy in C3GN shows a complex pattern of mesangial increase and glomerular basement membrane thickening. Differing combinations of subendothelial, intramembranous, and subepithelial deposits are noted.21,22,23

DDD is defined by the presence of dense osmiophilic transformation of the GBM on EM, and on light microscopy, the morphology is variable. While it is clear that a membranoproliferative pattern of glomerular injury with increased lobulation, mesangial expansion, and capillary wall thickening with segmental double contours is common, a range of other patterns of glomerular involvement also occur. Walker et al., 2007 collected 69 cases of DDD from centers in North America, Europe, and Japan. They identified four distinct histologic patterns on light microscopy: membranoproliferative (25%), mesangial proliferative (45%), crescentic (18%), and acute proliferative and exudative (12%).21 In the Columbia series conducted by Nasr et al., 2009 which includes 32 cases of DDD, the frequencies were MPGN (44%), mesangial proliferative(28%), endocapillary proliferative (19%), and crescentic GN (9%). These reports emphasize that fewer than 50% of cases of DDD have MPGN morphology. Morphologically, most C3GN cases show either a mesangial proliferative or membranoproliferative pattern.18

The dense deposits are recognized on light microscopy by thickening of the GBMs by ribbon-like glassy intramembranous deposits. They stain strongly with eosin and appear somewhat refractile (hyaline). They are intensely periodic acid-Schiff (PAS) positive, and the trichrome stain shows them to be fuchsinophilic (red) although this reactivity varies among specimens.23

 Immunofluorescence findings

Immunofluorescence shows characteristic C3 fragment deposition in C3GN.16  But the deposition of C3 is not always isolated. According to the current consensus report, the term ‘‘isolated’’ was replaced by “dominant staining of C3 defined as at least two orders of C3 intensity greater than that of any other immune reactant.” 4

Nasr et al., 2009, Walker et al., 2007, West and McAdams, 1998 studied the immunofluorescence findings. The invariable finding in DDD and C3GN is the presence of C3 in the glomeruli. Intense staining for C3 is noted along the glomerular capillary walls and often in the glomerular mesangial regions. The C3 deposition is usually diffuse and global. The GBM staining may be continuous or discontinuous. The early components of complement, C1q and C4, are usually absent, although occasionally C1q is found.  Immunoglobulins are usually absent or show only focal and segmental staining. If they are present, they often stain much less intensely than C3 and they are usually of the IgM type with a segmental distribution; IgG and especially IgA are less common.18,21,24

Treatment and Prognosis

Modality of treatment of MPGN is difficult and its prognosis is also guarded. About 50% develop chronic renal failure within 10 years. There is a high incidence of recurrence in transplant recipients, particularly in dense-deposit disease. Treatments with steroids, immunosuppressive agents, and antiplatelet drugs have not been proved to be materially effective.25

 Eculizumab, the first available anticomplement therapy, blocks at the level of C5 and has revolutionized the treatment of complement-mediated diseases as well as C3 glomerulopathy.13 This agent is a humanized monoclonal antibody that binds with great affinity to C5 proteins, inhibiting cleaving into C5a and C5b and blocking production of the C5b-9 membrane attack complex. Reports of individual cases showed improvement after treatment, with reduced serum creatinine and proteinuria. Bomback et al 2012, reported that, after 1 year of therapy with eculizumab, there was reduction in active glomerular proliferation and neutrophil infiltration three of five patients, consistent with effective C5 blockade.26


  1. Fakhouri F, Frémeaux-Bacchi V, Noël LH, Cook HT, Pickering MC. C3 glomerulopathy: a new classification. Nature Reviews Nephrology. 2010; 6(8):494.
  2. Barbour TD, Ruseva MM, Pickering MC. Update on C3 glomerulopathy. Nephrology Dialysis Transplantation. 2016;31(5):717-25.
  3. Barbour TD, Pickering MC, Cook Dense deposit disease and C3 glomerulopathy. InSeminars in nephrology 2013 Nov 1 (Vol. 33, No. 6, pp. 493-507). WB Saunders.
  4. Pickering MC, D’agati VD, Nester CM, Smith RJ, Haas M, Appel GB, Alpers CE, Bajema IM, Bedrosian C, Braun M, Doyle M. C3 glomerulopathy: consensus report. Kidney international. 2013; 84(6):1079-89.
  5. Cook HT, Pickering MC. Histopathology of MPGN and C3 glomerulopathies. Nature Reviews Nephrology. 2015; 11(1):14.
  6. Sethi S. Etiology-based diagnostic approach to proliferative glomerulonephritis. American journal of kidney diseases. 2014; 63(4):561-6.
  7. Levy M, Gubler MC, Sich M, Beziau A, Habib R. Immunopathology of membranoproliferative glomerulonephritis with subendothelial deposits (Type I MPGN). Clinical immunology and immunopathology. 1978; 10(4):477-92.
  8. Clardy CW, Judith F, Strife CF, West CD. A properdin dependent nephritic factor slowly activating C3, C5, and C9 in membranoproliferative glomerulonephritis, types I and III. Clinical immunology and immunopathology. 1989; 50(3):333-47.
  9. Neary JJ, Conlon PJ, Croke D, Dorman A, Keogan M, Zhang FY, Vance JM, Pericak-Vance MA, Scott WK, Winn MP. Linkage of a gene causing familial membranoproliferative glomerulonephritis type III to chromosome 1. Journal of the American Society of Nephrology. 2002; 13(8):2052-7.
  10. Sethi S, Nester CM, Smith RJ. Membranoproliferative glomerulonephritis and C3 glomerulopathy: resolving the confusion. Kidney international. 2012; 81(5):434-41.
  11. Sethi S, Fervenza FC. Membranoproliferative glomerulonephritis: pathogenetic heterogeneity and proposal for a new classification. In Seminars in nephrology 2011 Jul 1 (Vol. 31, No. 4, pp. 341-348). WB Saunders.
  12. Sethi S, Fervenza FC. Membranoproliferative glomerulonephritis – a new look at an old entity. New England Journal of Medicine. 2012; 366(12):1119-31.
  13. Bomback AS, Appel GB. Pathogenesis of the C3 glomerulopathies and reclassification of MPGN. Nature Reviews Nephrology. 2012; 8(11):634.
  14. Rabasco Ruiz C, Rabasco-Ruiz C, Huerta Arroyo A, Huerta-Arroyo A, Caro Espada J, Caro-Espada J, Gutiérrez Martínez E, Gutiérrez-Martínez E, Praga Terente M, Praga-Terente M. C3 glomerulopathies. A new perspective on glomerular diseases. Nefrología (English Edition). 2013 Mar 1;33(2):164-70.
  15. Medjeral-Thomas NR, O’Shaughnessy MM, O’Regan JA, Traynor C, Flanagan M, Wong L, Teoh CW, Awan A, Waldron M, Cairns T, O’Kelly P. C3 glomerulopathy: clinicopathologic features and predictors of outcome. Clinical Journal of the American Society of Nephrology. 2014; 9(1):46-53.
  16. Master Sankar Raj V, Gordillo R, Chand DH. Overview of C3 glomerulopathy. Frontiers in pediatrics. 2016; 4:45.
  17. Servais A, Noël LH, Roumenina LT, Le Quintrec M, Ngo S, Dragon-Durey MA, Macher MA, Zuber J, Karras A, Provot F, Moulin B. Grü nfeld JP, Niaudet P, Lesavre P, Frémeaux-Bacchi V: Acquired and genetic complement abnormalities play a critical role in dense deposit disease and other C3 glomerulopathies. Kidney Int. 2012; 82:454-64.
  18. Nasr SH, Valeri AM, Appel GB, Sherwinter J, Stokes MB, Said SM, Markowitz GS, D’Agati VD. Dense deposit disease: clinicopathologic study of 32 pediatric and adult patients. Clinical Journal of the American Society of Nephrology. 2009; 4(1):22-32.
  19. Lu Y, Shen P, Li X, Xu Y, Pan X, Wang W, Chen X, Zhang W, Ren H, Chen N. Re-evaluation of the classification system for membranoproliferative glomerulonephritis. In New Insights into Glomerulonephritis 2013 (Vol. 181, pp. 175-184). Karger Publishers.
  20. Servais A, Frémeaux-Bacchi V, Lequintrec M, Salomon R, Blouin J, Knebelmann B, Grünfeld JP, Lesavre P, Noël LH, Fakhouri F. Primary glomerulonephritis with isolated C3 deposits: a new entity which shares common genetic risk factors with haemolytic uraemic syndrome. Journal of medical genetics. 2007; 44(3):193-9.
  21. Walker PD, Ferrario F, Joh K, Bonsib SM. Dense deposit disease is not a membranoproliferative glomerulonephritis. Modern pathology. 2007; 20(6):605-16.
  22. Joh K, Aizawa S, Matsuyama N, Yamaguchi Y, Kitajima T, Sakai O, Mochizuki H, Usui N, Hamaguchi KI, Mitarai T. Morphologic variations of dense deposit disease: Light and electron microscopic, immunohistochemical and clinical findings in 10 patients. Pathology International. 1993; 43(10):552-65.
  23. Habib R, Gubler MC, Loirat C, Maiz HB, Levy M. Dense deposit disease: a variant of membranoproliferative glomerulonephritis. Kidney international. 1975; 7(4):204-15.
  24. West CD, McAdams AJ. Glomerular paramesangial deposits: association with hypocomplementemia in membranoproliferative glomerulonephritis types I and III. American journal of kidney diseases. 1998; 31(3):427-34.
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  27. Röth A, Dührsen U. Treatment of paroxysmal nocturnal hemoglobinuria in the era of eculizumab. European journal of haematology. 2011; 87(6):473-9.

Comparison between Bethesda System and Conventional System for Standardization of Reporting Thyroid Cytopathology

Comparison between Bethesda System and Conventional System for Standardization of Reporting Thyroid Cytopathology

 *Saleheen S,1 Rahman DA,2 Chowdury MA,3 Haque MM,4 Habib S,5 Khan KH6


  1. * Saied Saleheen, Assistant Professor, Department of Pathology, Sheikh Hasina Medical College, Tangail.
  2. DM. Arifur Rahman, Assistant Professor, Department of Pathology, TMSS Medical College, Gokul, Bogura,
  3. Mehdi Ashik Chowdury, Assistant Professor, Department of Pathology, Jahurul Islam Medical College, Bajitpur, Kishoreganj.
  4. Mohammad Mahbubul Hoque, Lecturer, Department of Pathology, Sheikh Sayera Khatun Medical College, Gopalganj.
  5. Saequa Habib, Associate Professor, Department of Pathology, Bangabandhu Sheikh Mujib Medical University, Dhaka.
  6. Kamrul Hasan Khan, Professor, Department of Pathology, Bangabandhu Sheikh Mujib Medical University, Dhaka.

 *For correspondence


Background: Fine-needle aspiration cytology (FNAC) is considered as an effective test to screen and diagnose patients with thyroid swelling for surgical management. But FNAC still suffers because of its inherent limitations as well as variability in its diagnostic terminology.The Bethesda System of Reporting Thyroid Cytopathology (TBSRTC) may improve the state. This study was to report the experience in using this reporting system to redistribute the cytological diagnoses made by conventional system and also to evaluate the specificity, sensitivity, accuracy and predictive values as a tool to compare both the methods based on the correlation between cytopathology and histopathology.

Methods: A total of 73 patients presenting with thyroid swelling were subjected to FNAC and reporting was done according to the conventional system and TBSRTC. The cytological diagnosis was correlated with the histopathological diagnosis. The sensitivity, specificity, predictive values were calculated considering cytology as screening test.

Results: In this study, TBSRTC was found superior because of higher sensitivity (91.66%) and specificity (97.77%) as compared to conventional system (80.76% and 87.23% respectively). Positive predictive value, negative predictive value and diagnostic accuracy of Bethesda system were 95.65%, 95.65% and 90.41% respectively which were also significantly higher as compared to those of conventional system (77.78%, 89.13% and 84.93% respectively).

Conclusions: TBSRTC may improve the efficacy of thyroid FNAC as a screening test.

 [Journal of Histopathology and Cytopathology, 2020 Jan; 4 (1):38-47]

 Keywords: Comparison, Thyroid FNAC, Bethesda system (TBSRTC), Conventional system, Histopathology, Standardization


Fine Needle Aspiration Cytology (FNAC) is a quick, cost-effective and minimally invasive outpatient procedure used worldwide in the initial diagnosis of thyroid swellings. As it can distinguish between benign and malignant lesions quite effectively, it is the preoperative screening method of choice worldwide.1 Before the routine use of thyroid Fine Needle Aspiration (FNA), the percentage of surgically resected thyroid nodules that were malignant was 14%.2 With current thyroid FNA practice, the percentage of resected nodules that are malignant surpasses 50%.3

However, due to the lack of a standardized system of reporting, pathologists have been using different terminologies and diagnostic criteria, leading to confusion amongst clinicians in the interpretation of the cytopathology report and ultimately hindering a definitive clinical management.4 It has also hindered the sharing of clinically meaningful data among multiple institutions.5 To mitigate this confusion all the pathologists need to use the same diagnostic criteria and terminology.6

Various reporting formats of thyroid FNAs have been suggested in the literature since the 1970s when thyroid FNAs began to be the subject of publications, and new formats continued to emerge.7 Several classification schemes have been suggested by various authors based on personal/institutional experiences but there is general support for the utilization of a tiered classification system.5 With this background, the National Cancer Institute (NCI), Bethesda, Maryland, United States, published an atlas and guidelines using standardized nomenclature for the interpretation of thyroid FNAC known as The Bethesda System for Reporting Thyroid Cytopathology (TBSRTC).4 It categorized the thyroid FNAC diagnoses into six groups with well-defined cancer risk and clear indications for further clinical management.8 It has also increased the interobserver reproducibility of cytopathological diagnoses of thyroid lesions.9-12

TBSRTC has been well received by both pathologists and clinicians and has been widely implemented by pathology laboratories in USA and in several European countries.13 Following publication of TBSRTC, the reporting system used in the United Kingdom since 2002, was updated by a working group of The Royal College of Pathologists and is now quite similar to TBSRTC.14 But TBSRTC still has to go a long way in getting acceptability in many countries of Asia and Africa.10

A good number of studies have been carried on in several countries in Asia (India, Pakistan, Iran, Korea and KSA) regarding usefulness of TBSRTC and it has been proved useful for management of patients with thyroid swelling in these countries.9,11,15-20 But this relatively recent six category scheme still needs to be validated by more prospective studies with histopathological correlation.16

In Bangladesh, there is no known reporting scheme using tiered classification system. The utility of thyroid cytopathology reporting according to TBSRTC in context of Bangladesh is also unexplored. Standardization of the reporting system by TBSRTC may result in improvement of the quality, homogenization and also reduce interobserver variability of thyroid cytopathology reports. In this context, this study was performed to assess the predictive values, sensitivity and specificity and accuracy of the TBSRTC in comparison with that of the conventional system used in the BSMMU, Dhaka, Bangladesh with histopathological correlation. The view was to observe the usefulness of Bethesda system for standardization of thyroid cytopathology reporting in the context of this country (Bangladesh).


This is a cross sectional study carried out at the Department of Pathology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka during the period from March 2015 to February 2017. A total of 73 patients presenting with thyroid swelling were included in this study, who has undergone FNA followed by the biopsy of the thyroid lesions. Patients presenting with thyroid swelling in any lobe of thyroid detected by clinical palpation (multinodular, solitary nodule, diffuse goiter etc) were included except those unwilling to be involved into the study.

All clinical information were recorded in a pre-designed proforma including demographic features, address and telephone contacts (for histological follow up). Results of all the routine investigations with special attention to ultrasonography of neck, serum T3, T4, and TSH levels and thyroid scintigraphy were noted where available.     

FNA of all patients was performed at the Department of Pathology, BSMMU.  Smears were stained with Papaniculaou (PAP) and Hematoxylin and Eosin (H&E) stains. Cytological findings were recorded and diagnoses were made according to both conventional and TBSRTC for each cases. For the TBSRTC reporting the Atlas10was used as manual (Table I). Reports were issued according to conventional method.

Koss’ Diagnostic Cytology and its histopathologic bases21 was followed conventionally as reference book in this department. The diagnoses made by conventional system are mostly based on its reference. The diagnoses were: Non-neoplastic lesions: Nodular goiters (NG) including cystic degeneration (CD), Lymphocytic thyroiditis (LT), Hashimoto thyroiditis (HT), Nodular goiter with co-existent thyroiditis, Multinodular goiter (MNG).

 Cellular follicular lesion (CFL): Cellular smears on cytology that included goiters with adenomatous changes, follicular adenomas and carcinomas, as well as the follicular variant of papillary carcinomas. In BSMMU suspicious cytology, atypia of undetermined significance and other gray zone cytology are also included within this diagnosis.

 Neoplastic lesions: Papillary thyroid carcinoma (PTC), suggestive of papillary carcinoma, medullary thyroid carcinoma (MTC) etc.

Biopsy specimens or histopathological reports of the patients undergoing surgery following the FNA interpretation under this study were collected. The histopathological findings were correlated with cytopathological findings and were recorded accordingly.

Descriptive analytical statistics was calculated. Thyroid FNA was considered as a ‘screening test’ and histopathology as gold standard. FNA benign was considered to be negative and the remaining categories were considered to be positive because they indicated significant risk of malignancy and led to a recommendation of surgery.8 Sensitivity, specificity, accuracy, the positive predictive value (PPV) and negative predictive value (NPV) were calculated from the available data by statistical formulae. In this study, Thyroid FNA has been considered as a screening method as TBSRTC is formed by NCI as a screening method to triage the patients effectively into groups that need surgery and the other that do not.5

For calculating statistical parameters nondiagnostic (ND) and AUS/FLUS cases were excluded as non-definitive diagnoses and categories, SFM and malignant were put together in most of the studies.15,16,22-24 In this study, the inadequate/ND cases were reaspirated and have been included rationally in other categories. Those which were still inadequate, had been excluded from statistical analysis.


Out of the 73 cases 62(84.9%) were female and 11(15%) were male. The age of the patients ranged from 16 to 70 years with average age around 40 years.

According to the conventional system forty five (61.64%) cases were diagnosed nodular goiter. Five (11.11%) of these lesions were found to be neoplastic (one benign and 4 malignant) on histopathology. Although all PTC and suggestive of PTC categories were found neoplastic and also malignant, 60% of the cellular follicular lesions (9 out of 15) were neoplastic (one benign and 8 malignant) and the rest were benign on final histological diagnosis.

The lesions when redistributed according to the Bethesda system: 46 (63%), 04 (5.4%), 03 (4.1%), 08 (10.95%), 12(16.4%) of the lesions were diagnosed as Cat-II, Cat-III, Cat-IV, Cat-V and Cat-VI respectively. The redistribution of the lesions after application of Bethesda system is shown in Table-II.

Out of 73 cases, 64 (22 neoplastic and 42 non-neoplastic) cytological diagnoses were concordant on both the conventional and Bethesda systems. One lesion was non-neoplastic on conventional but diagnosed neoplastic on Bethesda system while 04 neoplastic lesions by conventional system were categorized as non-neoplastic by Bethesda system. Three non-neoplastic and one neoplastic case according to conventional system were categorized as Cat- III (AUS) by Bethesda system. The CFL on conventional system were distributed by the Bethesda system into various categories that matched well (Table-III) with final histological diagnoses.

All the neoplastic lesions irrespective of benign and malignant neoplasm were considered positive for both the histological and cytological diagnoses, as they recommend for surgical excision. The concordances of the cytological diagnoses made by two methods with the final histological diagnoses are shown in Table IV.

Granulomatous thyroiditis: GT, Follicular adenoma: FA, Follicular carcinoma: FC, FVPTC: Follicular Variant of PTC, WDTUMP: Well differentiated tumor of uncertain malignant potential.

Diagnoses made according to TBSRTC shows increased overall concordance with hisotoathological diagnosis than that of the conventional system.

FNA interpretation by conventional method yielded significant false positive (6) and false negative (5) diagnoses. While analyzing the smears by the Bethesda system had decreased both the false positive (1) and false negative (2) interpretations. The sensitivity, specificity, accuracy and predictive values calculated for both the methods by using the statistical formulas are compared to each other in the Table V.

Sensitivity and positive predictive value (PPV) of the Bethesda system were found better (91.66% and 95.65% respectively) when compared to that the conventional system (80.76% and 77.78% respectively). The specificity and negative predictive values (NPV) were also found higher for Bethesda system (97.77% and 95.65%) in comparison to that of the conventional system (87.23% and 89.13% respectively). Diagnoses according to the Bethesda categories were 90.41% accurate where accuracy for the conventional system was 84.93%.


Reporting according to the conventional system included 41 (about 56.16%) patients into benign category, while category-II (Benign) in the Bethesda system included 44 (60.27%) patients. All (12) the suspicious for malignancy and malignant diagnoses on conventional system corresponded to category V and VI of Bethesda system. It reflects that there was actually no significant change in diagnosis of unequivocally benign and malignant cases according to the two systems of reporting.

15 (20.54%) CFL diagnosed by conventional system was the major field where the Bethesda system varied markedly. So, CFL was a “gray zone” diagnosis that included some benign (26.67%) and malignant (53.33%) cases.25 These cases when reclassified by the Bethesda system yielded more specific results with good histological correlation (Table IV). This difference can be explained by- i) Subjective underscoring of some neoplastic cytologies which are of SFM category in more objective Bethesda system and  ii) Assigning the follicular lesion diagnosis for some hyper cellular smears of goiter arranged in folded sheet appearance.

The 06 false-positive FNA diagnoses in conventional system included cellular smears of histologically nodular goiters that were interpreted as CFLs. Four of these cases were interpreted benign (category-II) by Bethesda system and 01 case was designated category-IV (suspicious for Hurthle cell neoplasm) because of relative abundance of Hurthle cells showing pleomorphism. Five cases were false negative for conventional method, 04 of which were due to compromised cytology samples. The rest one case diagnosed conventionally as nodular goiter that was histologically follicular adenoma and Category-IV (SFM) in Bethesda system.

When compared with other studies using TBSRTC with histological follow up, sensitivity and NPV of the present study were consistent with most of the studies (Table VI). The specificity and PPV were higher than most of the other studies, which may be due to small sample size in this present study.


By dint of its higher sensitivity, specificity, predictive values and accuracy TBSRTC has been proved to be better than  conventional system for reporting thyroid cytopathology. So, it should be useful for standardization of thyroid cytopathology reporting. However, studies needed to explore its effect in reducing inter-observer variation of reporting thyroid cytopathology as well as its usefulness to the clinicians in decision making.


  1. Bagga PK, Mahajan NC. Fine needle aspiration cytology of thyroid swellings: How useful and accurate is it? Indian Journal of cancer, 2010 Oct 1;47(4):437.
  2. Hamberger B, Gharib H, Melton LJ, Goellner JR, Zinsmeister AR. Fine-needle aspiration biopsy of thyroid nodules: impact on thyroid practice and cost of care. The American journal of medicine. 1982 Sep 1;73(3):381-4.
  3. Yassa L, Cibas ES, Benson CB, Frates MC, Doubilet PM, Gawande AA et al. Long‐term assessment of a multidisciplinary approach to thyroid nodule diagnostic evaluation. Cancer Cytopathology: Interdisciplinary International Journal of the American Cancer Society. 2007 Dec 25;111(6):508-16.
  4. Mondal SK, Sinha S, Basak B, Roy DN, Sinha SK. The Bethesda system for reporting thyroid fine needle aspirates: a cytologic study with histologic follow-up.Journal of Cytology. 2013; 30(2):94.
  5. Cibas ES, Ali SZ. The Bethesda system for reporting thyroid cytopathology. Thyroid. 2009 Nov 1;19(11):1159-65.
  6. Schinstine M. Pathology Hawaii Hotline: A Brief Description of the Bethesda System for Reporting Thyroid Fine Needle Aspirates. Hawaii medical journal. 2010 Jul; 69(7):176.
  7. Wang HH. Reporting thyroid fine‐needle aspiration: literature review and a proposal. Diagnostic cytopathology. 2006 Jan; 34(1):67-76.
  8. Ali SZ and Cibas ESThe Bethesda system for reporting thyroid cytopathology: definitions, criteria and explanatory notes. Springer Science & Business Media.2010
  9. Ahmed S, Ahmad M, Khan MA, Kazi F, Noreen F, Nawaz S, Sohail I. The interobserver reproducibility of thyroid cytopathology using Bethesda Reporting System: Analysis of 200 cases. J Pak Med Assoc. 2013 Oct 1;63(10):1252-5.
  10. Bhasin TS, Mannan R, Manjari M, Mehra M, Sekhon AK, Chandey M, Sharma S, Kaur P. Reproducibility of ‘The Bethesda System for reporting Thyroid Cytopathology’: a multicenter study with review of the literature. Journal of clinical and diagnostic research: JCDR. 2013 Jun; 7(6):1051.
  11. Mufti ST, Molah R. The Bethesda system for reporting thyroid cytopathology: a five-year retrospective review of one center experience. International journal of health sciences. 2012 Jun; 6(2):159.
  12. Pathak P, Srivastava R, Singh N, Arora VK, Bhatia A. Implementation of the Bethesda System for Reporting Thyroid Cytopathology: interobserver concordance and reclassification of previously inconclusive aspirates. Diagnostic cytopathology. 2014 Nov; 42(11):944-9.
  13. Auger M, Nayar R, Khalbuss WE, Barkan GA, Benedict CC, Tambouret R et al. Implementation of the Bethesda System for Reporting Thyroid Cytopathology: observations from the 2011 thyroid supplemental questionnaire of the College of American Pathologists. Archives of Pathology and Laboratory Medicine. 2013 Nov; 137(11):1555-9.
  14. Crippa S, Dina R. Interobserver reproducibility of thyroid fine-needle aspiration using the UK Royal College of Pathologists’ classification system. American journal of clinical pathology. 2012 May 1;137(5):833-5.
  15. Mamatha M, Sekhar SC, Rani HS, Anil SS and Vandana G. A comparative study between conventional system and the Bethesda system applied for reporting thyroid cytopathology. International Archives of Integrated Medicine.2015; 2(3):87 – 95
  16. Mehra P, Verma AK. Thyroid cytopathology reporting by the Bethesda system: a two-year prospective study in an academic institution. Pathology research international. 2015;2015. 240505.doi: 1155/2015/240505
  17. Ahmed S, Ahmad M, Khan MA, Kazi F, Noreen F, Nawaz S, Sohail I. The interobserver reproducibility of thyroid cytopathology using Bethesda Reporting System: Analysis of 200 cases. J Pak Med Assoc. 2013 Oct 1; 63(10):1252-5.
  18. Bukhari MH, Khan AA, Niazi S, Arshad M, Akhtar ZM, Al-Sindi KA. Better thyroid cytopathology reporting system may increase the clinical management and patients outcome. J Cytol Histol. 2012;3(6):1000158.
  19. Hajmanoochehri F, Rabiee E. FNAC accuracy in diagnosis of thyroid neoplasms considering all diagnostic categories of the Bethesda reporting system: A single-institute experience. Journal of Cytology/Indian Academy of Cytologists. 2015 Oct; 32(4):238.
  20. Jo VY, Stelow EB, Dustin SM, Hanley KZ. Malignancy risk for fine-needle aspiration of thyroid lesions according to the Bethesda System for Reporting Thyroid Cytopathology. American journal of clinical pathology. 2010 Sep 1;134(3):450-6.
  21. Sanchez AM and Stahl RE.The Thyroid, Parathyroid, and Neck Masses Other Than Lymph Koss LG, Melamed MR, editors. Koss’ diagnostic cytology and its histopathologic bases. Lippincott Williams & Wilkins; 2006.pp1148-1173.
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  26. Gupta M, Gupta S, Gupta VB. Correlation of fine needle aspiration cytology with histopathology in the diagnosis of solitary thyroid nodule. Journal of thyroid research. 2010 Apr 18;2010:379051.

Histopathological Spectrum of Prostatic Lesions Evaluated in a Tertiary Hospital

Histopathological Spectrum of Prostatic Lesions Evaluated in a Tertiary Hospital

 *Sultana SS,1 Hossain S,2 Rahman A3


  1. *Dr. Sk Salowa Sultana, Assistant Professor, Department of Pathology, Ad-Din Women’s Medical College.
  2. Shahadat Hossain, Associate Professor, Department of Pathology, Ad-Din Women’s Medical College.
  3. Afiqur Rahman, Professor of Urology, Department of Urology, Ad-Din Women’s Medical College.

*For correspondence


Objective: To determine the age distribution of various prostatic lesions, to evaluate histopathological  pattern  of prostatic lesions  and  to analyze adenocarcinoma  of prostate according to Gleason  system.

Methods: This was a retrospective study comprising of 178 cases, carried out at the department of Pathology, Ad-din Women´s  Medical College Hospital during the  period of January 2017 to December 2019.

Results: Out of 178 cases, 159(89.32%) were diagnosed as benign lesion, nodular hyperplasia being the commonest one. Prostatic adenocarcinoma was found in 16(9%) cases, majority of which belonged to Gleasons score 7.

Conclusion: Histopathological examination of prostatic biopsy specimen is essential for diagnosis of benign and malignant lesions, to rule out the incidental carcinoma and HGPIN.

 [Journal of Histopathology and Cytopathology, 2020 Jan; 4 (1):33-37]

 Keywords: Nodular hyperplasia of prostate, prostatic carcinoma, HGPIN


The  prostate that weights up to 20 gm in normal adult depends for its subsequent growth and differentiation on androgenic hormones.1,2  Lesions of prostate are extremely common over the age of 50 years and important cause of morbidity and mortality in male in advance age. Pathological processes affecting this organ include inflammation, benign nodular enlargement and tumor.1 The clinical incidence of the diseases is only 8% during the 4th decade  but it reaches 50% in the 5th decade and 75% in the 8th decade.2 Benign prostatic  hyperplasia(BPH) and carcinoma are increasingly frequent with advancing age.3  BPH results from hyperplasia of both stromal and epithelial cells, gradually being enlarged, compress and narrow the urethral canal and often leads to urinary obstruction. Inflammation of prostate is known as prostatitis and divided into acute, chronic and granulomatous prostatitis. Prostatic carcinoma is one of the most common malignancies affecting men, also it is the sixth leading cause of cancer death in male.4 Among the prostatic malignancies 95% is adenocarcinoma. premalignant lesion of prostatic adenocarcinoma  is known as high grade intraepithelial neoplasia (HGPIN). The objective of this study was to determine the age distribution of various prostatic lesions, to evaluate histopathological  pattern  of prostatic lesions  and  to analyze adenocarcinoma  of prostate according to Gleason  system.


The study was conducted in the Department  of Pathology, Ad-din Women´s Medical College Hospital – a  tertiary level hospital of Dhaka, Bangladesh. The study period was from January 2017 to December 2019. The data were collected retrospectively from histopathology register. A  total of 178 TURP (Transurethral resection of prostate) were evaluated. The received specimen were fixed in 10% formalin and routine paraffin processing followed by hematoxylin and eosin staining was done and the slides were examined under light microscope. The specimens were ananlyzed as type of specimen, age of the patient, histopathological  pattern and final diagnosis. The tumour were classified according to 2016 WHO classification and histological grading was done using modified Gleason´s system. Data  were analyzed using tables, figures and percentage.


A total of 178 cases were included in our study, all of which (100%) were TURP specimen.  In the present study, most of the prostatic lesions were benign(89.32%) followed by malignant (9.6%) and HPIN (1.12%) and ratio of benign and malignant lesions is 1:9, approximately.

In the present study, benign lesions were mostly found (45%) in the age group of 61-70 years  and 34% in the age group of 51-60 years,  whereas malignant lesions were common  41% in the age group of 71-80 years and 29% in the age group of 61-70 years of age. In our study, benign lesions(159) were nodular hyperplasia of prostate 130(73.03%) and nodular hyperplasia of prostate with co-existing prostatitis 29(16.3%). Out of 29 cases of prostatitis, 24 cases were chronic non specific prostatitis, 2 cases were granulomatous prostatitis and 3 cases were acute prostatitis.  In our study, 2(1.12%) cases were diagnosed as HGPIN and 17(9.6%) cases were diagnosed as malignant. Most of the malignant cases 16(9%)  were prostatic adenocarcinoma and 1(0.56%) was metastatic adenocarcinoma. In this study, 41% cases of malignant belong to 71-80 years age group and 29% cases belong to 61-70 years age group. According to Gleason score, most commonly (81.25%) found grade was grade 7, in this study.


We enrolled a total of 178 cases in our study, all of which (100%) were TURP specimen. Bhatta S et al5 have found 88.54% specimen of TURP in their study. Screening procedure like trans-rectal ultrasound, prostate specific antigen (PSA) are still used, but biopsy remains the gold standard for final diagnosis.6 In the present study, most of the prostatic lesions were benign (89.32%) followed by malignant (9.6%) and HPIN (1.12%). These findings are similar to the studies done by Bhatta S et al,5 Bal et al,7 Jehoram  et al,8 and Bhat S et al.9 They found (89.58%), (87%), (93%) and (92.4%) cases of BPH in their study, respectively. In our study, ratio of benign and malignant lesions is 1:9, approximately.

In the present study, benign lesions were mostly found 45% in the age group of 61-70  years  and 34% in the age group of 51-60 years,  whereas malignant lesions were common  41% in the age group of 71-80 years and 29% in the age group of 61-70 years of age. These findings are in concordance with other studies.10,11,12

In our study, benign lesions(159) were nodular hyperplasia of prostate 130(73.03%) and nodular hyperplasia of prostate with co-existing prostatitis 29(16.3%). Out of 29 cases of prostatitis, 24 cases were chronic non specific prostatitis, 2 cases were granulomatous prostatitis and 3 cases were acute prostatitis.  Bhatta S et al found 24(25%) cases of prostatitis associated with nodular hyperplasia of prostate, out of which 22 cases were chronic non specific prostatis and two cases were acuteprostatis.5 Prostatic carcinoma is one of the most common malignancies affecting men. In our study, 2(1.12%) cases were diagnosed as HGPIN and 17(9.6%) cases were diagnosed as malignant. Most of the malignant cases 16(9%)  were prostatic adenocarcinoma and 1(0.56%) was metastatic adenocarcinoma. Bhatta S et. al5found 2.08% of HGPIN and 8.34% of prostatic carcinoma. Also Deshmukh BD et. al11 and Bhat S et. al9 found similar observation. In this study, 41% cases of malignant  belong to 71-80 years age group and 29% cases belong to 61-70 years age group. Bhat S et. al12  found 75% cases of malignancy diagnosed after 60 years. All the cases of prostatic adenocarcinoma were graded according to Gleason score which determine the tumour aggressiveness.In this study, most commonly 13(81.25%) found grade was grade 7. Albasri et al.13 found Gleasons score 5-7 as the commonest among 71 cases of prostatic adenocarcinoma. Bhat S et. al9 found 56.16% of adenocarcinoma with Gleason score 8-9.


Histopathological  examination of prostatic lesions is essential for diagnosis and management. Prostatic lesions are common in the age group of 60-70 years. All the specimen received were TURP. Benign prostatic hyperplasia (BPH) was predominant type among all the lesions. Majority of the malignant lesions were incidental diagnosis and most of the malignant lesions were prostatic adenocarcinoma. Emphasis should be  given to identify premalignant lesions.


  1. Epstein JI. The lower urinary tract and male genital system. 8th rev ed. Kumar V, Abbas AK, Fausto N, Aster JC, editors. Robbins and Cotran. New Delhi: Elsevier Pathologic Basis of Disease, 2010. p993-1002.
  2. Rosai J. Male reproductive system. In: Rosai J, editor. Rosai and Ackerman’s Surgical Pathology. 10thed. New Delhi: Elsevier, 2011:1287-1333.
  3. A Josephin-2014, clinicopathological study of prostatic biopsy, ©2014journal of Clinical and Diagnostic 2014 Sep; 8(9): FC04-FC06.
  4. Chandanwale S, Jadhav PS, Anwekar SC, Kumar H, Buch AC, Chaudhari US, et al. Clinico-pathological study of benign and malignant lesions of prostate. IJPBS. 2013;3:162-178.
  5. Bhatta S, Hirachan S. Prostatic lesions: Histopathological Study in a Tertiary Care Hospital. JMMIHS. 2018;4(1):12-19.
  6. Garg M, Kaur G, Malhotra V, Garg R. Histopathological spectrum of 364 prostatic specimens including immunohistochemistry with special reference to grey zone lesions. Prostate Intl. 2013;1:146-151.
  7. Bal MS, Kanwal S, Goyal  AK, Singla N, Prostatic lesions in surgical biopsyspecimen. JK Pract 2011;16:33-4.
  8. Jehoram TA, Sitara AS, Mohammed EB. Hyperplastic , premalignant and malignant lesions of the prostate gland. Hum Pathol. 2005;36:480-5.
  9. Bhat S, Chaudhri S, Bhat P, Hatwal D. Histopathological study of prostatic Diseases in Garhwal Region. Int  J Sci stud. 2015: 3(8):136-140.
  10. Shakya G, Malla S, Shakya KN. Salient and co-morbid features in benign prostatic hyperplasia: A histopathological study of the prostate. Kathmandu Univ Med J. 2003;2:104-109.
  11. Deshmukh BD, Ramteerthakar NA, Sulhyan KR. Histopathological study of lesions of prostate- A five year study. Int J Health Sci Res. 2014;4:1-9.
  12. Yadav M, Desai H, Goswami H. Study of Various Histopathological Patterns in Prostate Biopsy. IJCRR. 2017;9(21):59-63.
  13. Albasri A, EL-Sidding A, Hussainy A, Mahrous M, Alhosaini AA, Alhujaily A. Histopathologic characterization of prostatic diseases in Madinah, Saudi Arabia. Asian Pac J Cancer prev. 2014;15:4175-9.

Histopathological Patterns of Ovarian Tumours and HER2 Expression in Epithelial Ovarian Carcinoma

Histopathological Patterns of Ovarian Tumours and HER2 Expression in Epithelial Ovarian Carcinoma

*Barua M,1  Hossain MI,2  Bini UH,3 Nasreen S,4 Ahamad MU,5 Bhattecharjee P,6 RahmanM Z7



  1. * Mitasree Barua, Lecturer, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh.
  2. Mohammad Ismail Hossain, Lecturer, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh
  3. Umme Habiba Bini, Assistant Professor, Department of Pathology, Pabna Medical College, Pabna, Bangladesh.
  4. Sayeeda Nasreen, Assistant Professor, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh.
  5. M. Sahab Uddin Ahamad, Associate Professor, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh.
  6. Pradip Bhattacharjee, Associate Professor, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh.
  7. Md. Zillur Rahman, Professor & Head, Department of Pathology, Chittagong Medical College, Chattogram, Bangladesh.

 *For correspondence


Background: Ovarian tumours are a heterogenous group of neoplasm of epithelial, stromal and germ cell origin. The management of which depends on the histological type of the tumour. Neoadjuvant chemotherapy is limited by toxicity and resistance. Hence targeted therapy is now being proposed to overcome these hurdle.

Objectives: To find out the histopathological patterns of ovarian tumours and to evaluate HER2 overexpression in epithelial ovarian carcinoma.

Methods: This study was carried out in the Department of Pathology, Chittagong Medical College, Chattogram, received from Department of Gynaecology and Obstetrics, Chittagong Medical College Hospital, Chattogram during the period from January 2016 to December 2016. A total of 92 cases of ovarian tumours were selected consecutively. The age range was 12-80 years. Histopathological sections were stained with Hematoxylin and Eosin stain and epithelial ovarian carcinoma were subjected to HER2/neu immunohistochemical stain. Histopathological patterns of ovarian neoplasm, histopathological subtyping and grading of epithelial ovarian tumours and the expression of HER2/neu among epithelial ovarian carcinomas on immunohistochemistry was analyzed.

Results: Out of total 92 cases, histopathological diagnosis showed 65 (70.7%) were benign tumours and 27 (29.3%) were malignant tumours. Histologically, surface epithelial tumours were the most common (74%) followed by germ cell tumours (21.7%) and sex cord stromal tumours (4.3%). Age incidence of benign tumour was age group of 21-40 yrs and malignant 41-60 years. One in 23 epithelial ovarian carcinoma cases (4.3%) exhibits HER2 overexpression. Only one HER2 positive case was mucinous cystadenocarcinoma grade II.

Conclusion: Surface epithelial tumours were most common followed by germ cell tumours. Serous cystadenocarcinoma were most common epithelial ovarian carcinoma.


[Journal of Histopathology and Cytopathology, 2020 Jan; 4 (1):23-32]

 Keywords: Ovarian tumour, Epithelial Carcinoma, HER2/neu.



Ovarian tumours account for 3% of all cancers in females, being the second most common cancer of the female genital tract, next only to uterine cancer.1 Ovarian cancer, traditionally known as the silent killer, is a dreaded disease because of its vague, nonspecific symptoms and late presentation. Worldwide, it is the seventh most common cause of cancer and eighth most common cause of cancer-related death among women.2 It is be a disease of developed and industrialized countries where parity of women is low. As the standard of living increases and resources for a medical infrastructure improve in Bangladesh, there will be an increased need to address less common but highly lethal cancers such as ovarian cancer. Globocan predicts a change in the reported incidence of ovarian cancer from 2912 in year 2012 to 3132 in year 2015.3

Risk factors for ovarian cancer are not well defined. However, there is general agreement on two: nulliparity and family history. Women 40 to 59 years of age who have taken oral contraceptives or undergone tubal ligation have a reduced risk of developing ovarian cancer.4 Two treatment options, available for ovarian cancers in advanced stage, are either a primary surgical cytoreduction/debulking or chemotherapy in an attempt to downstage the tumor followed by surgery. Though platinum-based therapy has produced an impressive result, chemoresistance and toxicity are creating hurdles. To overcome this, monoclonal antibodies targeting HER2/neu, a transmembrane protein homologous to epidermal growth factor receptor, has been introduced recently that might bring a new era of management.5

HER2/neu protein expression is commonly measured using immunohistochemistry (IHC).6 Overexpression of HER2/neu is seen in 20–30% patients with ovarian cancer. Some of these differences are likely to be attributable to the diagnostic technique used to measure HER2 expression. HER2/neu overexpression in epithelial ovarian carcinomas has not been studied as extensively as it has been studied in case of breast carcinomas, especially in Bangladesh scenario.

The aim of my study was to determine histopathological patterns of ovarian tumor and to evaluate the frequency of the expression of HER2 among epithelial ovarian carcinomas.


It was a cross-sectional descriptive study carried out in the Department of Pathology of Chittagong Medical College, Chattogram, Bangladesh. Study period was for one year from 1st January, 2016 to 31st December, 2016. All female patients clinically diagnosed as ovarian neoplasm received at the Department of Pathology of Chittagong Medical College, Chattogram referred from Department of Gynaecology and Obstetrics, Chittagong Medical College Hospital (CMCH) during the specified time duration. Finally, 92 consecutive patients were selected according to time frame.

 Inclusion criteria

Patient with clinically diagnosed as ovarian tumours who underwent intervention like diagnostic laparoscopy or laparotomy; at least for biopsy in Chittagong Medical College Hospital, Chattogram.  Patient with histopathologically diagnosed ovarian tumour in the Department of Pathology, Chittagong Medical College.

 Exclusion criteria

Previously diagnosed case of ovarian tumour who already has got treatment. Patient unwilling to give written consent.

Protocol was ethically reviewed and approved by the Ethical review Committee of Chittagong Medical College, Chattogram. All the patients included in the study were informed and explained about the nature of study. Informed written consent was taken from all the subjects after full explanation of nature, purpose and potential risks of all the procedures.

A brief clinical history was taken from each patient with particular reference to the age, occupation, marital history, parity, menstrual history, family history, history of breast feeding, use of contraceptives etc. Clinical history, questionnaire, thorough physical examination, and relevant investigations were recorded in details in all cases.Routine histopathology with H & E stain and Immunohistochemistry for ovarian carcinoma were done.

H&E stained slides were examined under light microscope to get a definitive diagnosis of the ovarian tumour and its type. Histological classification was based on the 2003 World Health Organization classification of ovarian tumours. The Shimizu-Silverberg three-grade histological grading system was used in epithelial ovarian carcinoma (EOC).

The most representative tumor tissue was chosen from each epithelial ovarian carcinoma case and 4 μm sections were taken to poly-L-lysine coated slides for immunohistochemical staining. For immunohistochemistry, epitope retrieval was done with a polymer-based detection system (Envision; Dako) using a Herceptin kit (HercepTest; Dako) according to the manufacturer’s instructions.Antigen retrieval for HER2 using Hercep Test was performed by immersing and incubating the slides in 10 mmol/L citrate buffer in calibrated water bath (95- 99°C) for 40 minutes. After decanting the epitope-retrieval solution, the sections was rinsed in the wash buffer and later, soaked in the buffer for 5 to 20 minutes before staining. Then the slides were rinsed, placed in 200 μL peroxidase-blocking reagent for 5 minutes, rinsed, placed in 200 μL primary anti-HER2 protein (or negative control reagent) for 30 minutes, rinsed twice, and finally immersed in 200 μL substrate chromogen solution (3,3´-diaminobenzidinetetrahydrochloride, DAB) for 10 minutes. The slides were counter stained with hematoxylin and finally were cover slipped.


In this study the age range of 92 ovarian tumours was 12-80 years, were divided into seven age groups and it was observed that maximum number of benign lesions 19 (20.6%) were in age groups 21-30 years. Maximum number of malignant tumours were found 9 (9.8%) in age group 41-50 years. Among 92 ovarian lesions, 68 (74%) were surface epithelial tumour, 20 (21.7%) were germ cell tumour, 4 (4.3%) were sex cord stromal tumours .


Current study was carried out in Departmant of pathology, Chittagong Medical College to see the histopathological patterns of ovarian tumours and HER2 expression in epithelial ovarian carcinoma (EOC).Total number of the patients in this study was 92. The age range was 12-80 years. The mean age was 34.64 years. Patients were divided into seven groups consisting of each decade as a single group and maximum number patients 23 (25%) were in 21 – 30 years group, followed by 21 (22.8%) were in 41-50 years age group. Study done in Nepal by Vaidya et al.(2014) showed highest number of patients (58%) were in 21-40 years age group which is similar to present study.7 In this study in 21-40 years age group total number of patients were 42 (45.65%). Study done in India by Bhagyalakshmi et al (2014)) patients age ranged from 11-70 years, majority of benign cases and malignant cases were between 21 to 40 years and 41 to 60 years respectively.8 Agrawal et al. (2015) also showed age ranged from 12 to 80 years with most common benign and malignant lesions cases were 21 to 40 years and 41 to 60 years respectively which was consistent with this study.9

In this study, among 92 cases, 72 (78.3%) were married and 20 (21.7%) cases were unmarried women. This is probably due to more married women presented to the outpatient department than unmarried ones. Among 72 married women 08 (11.1%) were nulliparous, 03(4.2%) were primiparous and 64 (84.7%) were multiparous.Study done in Dhaka by Dhar et al (2015) showed 8% were unmarried and 92% were married, of whom 12% were nulliparous and 80% was parous.10

Regarding socio-economic status 68 (73.9%) cases were from average socioeconomic condition, 23 (25%) were from poor and only 1 (1.1%) cases from high socioeconomic family. Among the malignant cases maximum 13 (48.1%) cases were from average socioeconomic condition. This is similar to the finding of  Dhar et al (2015).10This may be due to the fact that patients attended at a government hospital and most of the people of average and poor socioeconomic conditions usually come here to get treatment.

In this study, it was seen that, 19 (20.7%) had history of hormonal contraceptive use and 49 (53.3%) had no history. Among 27 malignant cases 3 (11.1%) had history of contraceptive use. Ness et al (2011) had reported that oral contraceptives reduce the risk of developing ovarian cancer.11

In this study 75 (81.5%) patients presented with lower abdominal pain 74 (80.4%) with lower abdominal mass. Study done by Jaffar et al (2013) showed, abdominal pain was in 90% cases, abdominal mass was present in 24% of the patients with benign tumours and in 66% with malignant tumours.12

In the present study, among 65 benign lesions 20 (21.7%) cases had increased level of CA-125 and among 27 malignant lesions 3(3.3%) cases were found within normal level.Miralles et al. (2003) suggested that variety of malignancies and benign conditions courses with increased CA-125 level and it is observed that CA-125 is very important as tumour marker for malignancy.13

In the present study, out of 65 benign tumours 62(95.3%) were unilateral and 3(4.6%) were bilateral. Out of 27 malignant tumours 5(18.51%) were bilateral and 22(81.48%) were unilateral. Modepalli et al (2016) reported 93.8% of the neoplasms were unilateral and remaining 6.2% of the lesions were bilateral.14 Benign tumours were more unilateral than malignant tumours. In this present study, out of 92 cases, maximum number of lesions were 48 (52.2%) in the right side of abdomen which is similar to the study done by Modepalli et al (2016).14

Among 92 cases 43 (46.7%) were solid and cystic lesion, 41 (44.6%) were cystic lesions and 8 (8.7%) were solid. Similar study done by Agrawal et al (2017) showed 83.3% benign tumours were cystic whereas 65.3% malignant tumours were solid and cystic.9 Benign tumors were more often cystic in consistency in this study and malignant tumors were solid consistency which was comparable to the study of Kanthikar et al. (2014).15

In this study of the 92 cases of ovarian tumours, 67 (70.7%) were benign and 27 (29.3%) were malignant. Similar studies in India carried out by GG swamy et al. (2010) showed 86 (71.6%) benign tumours, 30 (25.1%) malignant and 4 (3.3%) borderline tumors.16 In Nepal R Jha et al reported 83.9% benign, 16.1% malignant and 2.8% borderilne ovarian tumour.17 In  Pakistan a similar study by Ahmed et al showed 59.18% benign and 40.81% malignant tumour. In this study there was no borderline malignancy which were similar to the study done by Yasmine et al. (2008).18

Among the histologic types, in the present study, surface epithelial tumours were most common 68 (74%), germ cell tumour was 20 (21.7%), and sex cord stromal tumour was 4 (4.3%).Similar observation made by Pilli et al.(2002) showed 70.9% surface epithelial tumours, 21.2% germ cell tumours and 6.7% sex cord stromal tumours.19

In this present study Among 92 cases the most common benign surface epithelial tumour was mucinous cystadenoma 26 (28.3%) followed by serous cyst adenoma 18 (19.6%) and benign brenner tumour 1(1.1%). The most common benign germ cell tumour was mature cystic teratoma 18 (19.6%).In this study benign sexcord stromal tumour was fibroma 2(2.2%). Studies done by Bhagyalakshmi et al (2014)8 and Mondol et al (2011)20 showed serous cystadenoma was the most common benign tumour. Study done by Ahmed et al (2000) however found the most common benign tumour to be benign cystic teratoma.21

The commonest malignant tumour was serous cyst adenocarcinoma 13 (14.1%), followed by mucinous cystadenocarcinoma 8 (8.7%), malignant brenner tumour 1 (1.1%) and clear cell carcinoma 1 (1.1%). Similar results were shown in studies by Mondol et al (2011) and Yogambal et al (2014).22

In this present study 23 cases of epithelial ovarian carcinoma (EOC) were evaluated for HER2/neu protein expression using immunohistochemistry.The positive expression rate of HER2/neu in this study was 1(4.3%).This is comparable to the rate of  HER2/neu positivity in EOC reported in the literature,which ranges from 7-50%Verri et al (2005),23 Berchuck et al (1990),24 Mayr et al (2006).25 The variation in HER2/neu protein expression rate in different studies may be early stage and 17% of patients with advanced stage disease in the study by Kacinsky et al (1992). However, multiple factor may be involved in producing a wide range of positivity, such as sample size, the detection technique and type of monoclonal antibody used, staining intensity and the tissue where analysis is performed. Studies with populations smaller than 100 patients reported a frequency of 1.8-76%,(Serrano-Olvera et al., 2006).26

In this study among 8 mucinous cystadenocarcinoma, HER2 expression was positive in 1 (12.5%) case and was in Grade-II. In the study done by Missaoui et al (2014),27 HER2 expression was described in 14.3% of mucinous carcinoma. Study in Nijeria done by Ajani et al.(2015) 28 observed that a higher proportion of serous carcinoma (as opposed to mucinous carcinoma) was HER2/neu positive. Sarkar et al (2015)2 also showed that HER2 expression was more intense for high grade serous carcinoma.

In this study HER2 expression is relatively common in mucinous epithelial carcinoma of the ovary and trastuzumab therapy would be an option for patients with mucinous carcinoma when the tumour has HER2 overexpression.

The study also has some limitations. This study was severely constrained by the limited duration that led to have small population size and this study was done in patient who underwent surgery and samples were collected from Chittagong Medical College and Hospital only. In spite of these reason the present study showed more or less acceptable findings with consideration of the observations by others.


Ovarian cancer presents a tremendous clinical challenge to gynecologists, medical oncologists and radiotherapists. It is a silent menace and is not associated with significant symptoms. Surface Epithelial Tumours were the commonest group followed by Germ Cell Tumours. Most common Germ cell tumours were mature cystic teratoma occured in much younger women. Malignant ovarian neoplasms were more common in patients above 40 years. Sex Cord Stromal Tumours were also common with Fibroma and Granulosa cell tumour. Unilateral involvement of ovarian neoplasms was more common than bilateral and involvement of right ovary was more than the left. Mucinous cystadenoma was the commonest benign ovarian neoplasm while the commonest malignant tumour was serous cystadenocarcinoma. Mucinous epithelial carcinoma have expressed HER2.


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Correlation of Lymphovascular Density with Histological Prognostic Parameters in Gastric Carcinoma

Correlation of Lymphovascular Density with Histological Prognostic Parameters in Gastric Carcinoma

*Hassan MI,1 Kabir AN,2 Poly FH,3 Ray S,4 Munmun UK,5 Mehjabin M6


  1. * Mohammed Imran Hassan, Assistant Professor (CC), Department of Pathology, National Institute of Diseases of Chest and Hospital, Mohakhali, Dhaka.
  2. AKM Nurul Kabir, Associate Professor, Department of Pathology, BangabandhuSheikh Mujib Medical University, Shahbag,Dhaka.
  3. Ferdousi Hossain Poly, Medical Officer, OSD, DGHS, Attached- Department of Pediatric Neurology, National Institute of Neurosciences and Hospital, Dhaka.
  4. Subrata Ray, Assistant Professor, Department of Pathology, Jashore Medical College.
  5. Umme Kulsum Munmun, Lecturer, Department of Pathology, Dhaka Medical College, Dhaka.
  6. Monika Mehjabin, Lecturer, Department of Pathology, Shahid Suhrawardy Medical College, Dhaka.

*For correspondence


Background: Carcinoma of the stomach is a major cause of cancer mortality worldwide. Gastric cancer is actually a curable disease if it is detected at an appropriate stage and treated adequately. It rarely disseminates widely before it has involved the lymph nodes and, therefore, there is an opportunity to cure the disease prior to dissemination. Tumor-induced lymphangiogenesis plays a crucial role in metastasis and tumor progression. However, the significance of lymph vessel density has been controversial in gastric cancer. Specific lymphatic markers are now available making possible analysis of lymphatic in cancer. The aim of this study was to investigate the relation of lymph vessel density with histological prognostic factors in gastric cancer as potential indicator.
Objectives: To estimate of the lymph vessel density using Podoplanin immunostain in gastric cancer and then to correlate lymph vessel density with histological prognostic factors.
Methods: Sections from Paraffin blocks of gastric adenocarcinoma with routine stain were assessed to detect prognostic factors and then assessed with immunostain Podoplanin to count lymph vessel density. The mean number of lymph vessels in five hot spots was calculated in both intratumoral and peritumoral areas. Finally correlation was seen between histological prognostic factors and lymph vessel density in gastric carcinoma.
Results: A positive significant correlation was found between lymph node metastasis with lymph vessel density in both intratumoral and peritumoral area. Similarly, a positive significant correlation was found between tumor stage with lymph vessel density in both intratumoral and peritumoral area.

 [Journal of Histopathology and Cytopathology, 2020 Jan; 4 (1):12-22]

 Keywords: Podoplanin, Lymphovascular density, Gastric carcinoma


Carcinoma of the stomach is a major cause of cancer mortality worldwide. The prognosis of gastric carcinoma is poor with cure rates little better than 5–10%, although better results are obtained in Japan where the disease is common. Gastric cancer is actually a curable disease provided that it is detected at an appropriate stage and treated adequately. It rarely disseminates widely before it has involved the lymph nodes and, therefore, there is an opportunity to cure the disease prior to dissemination.1

Worldwide gastric cancer ranks fourth in frequency and second in cancer mortality rate; with a 5-year survival rate not exceeding 30% in Western countries.2 Gastric cancer incidence varies markedly with geography. In Japan, Chile, Costa Rica, and Eastern Europe the incidence is up to 20-fold higher than in North America, northern Europe, Africa, and Southeast Asia.1 The incidence of gastric carcinoma in Bangladesh is 5.2 per 100,000.3


Gastric cancer is a multi-factorial disease. Epidemiological studies point to a role for Helicobacter pylori, although its importance is disputed. Helicobacter pylori seem to be principally associated with carcinoma of the body of the stomach and the distal stomach rather than the proximal stomach. As Helicobacter is associated with gastritis, gastric atrophy and intestinal metaplasia, the association with malignancy is perhaps not surprising. Patients with pernicious anemia, gastric atrophy, duodenogastric reflux and reflux gastritis, intestinal metaplasia and cigarette smoking are at increased risk, as are those with gastric polyps. Diet appears to be important factor. The ingestion of substances such as spirits may induce gastritis and, in the long term, cancer. Excessive salt intake, deficiency of antioxidants and exposure to N-nitroso compounds are also implicated.1

The most useful classification of gastric cancer is the Lauren classification.4 In this system, there are principally two forms of gastric cancer: intestinal gastric cancer and diffuse gastric cancer. In intestinal gastric cancer, the tumor resembles carcinomas found elsewhere in the tubular gastrointestinal tract and forms polypoid tumors or ulcers. It probably arises in areas of intestinal metaplasia. In contrast, diffuse gastric cancer infiltrates deeply into the stomach without forming obvious mass lesions but spreading widely in the gastric wall.1

These tumors have several prognostic and predictive factors. The depth of invasion and the nodal and distant metastasis at the time of diagnosis remain the most important prognostic indicators for gastric carcinoma. Currently tumor size, nuclear grade and patient’s age are well known prognostic factors for patients with operable gastric cancer. These prognostic factors are widely used to determine whether to apply neo-adjuvant therapy in patient with gastric cancer.  In advanced cases, gastric carcinoma may first be detected as metastases to the supraclavicular sentinel lymph nodes, also called Virchow’s node. Gastric carcinoma can also metastasize to the periumbilical region to form a subcutaneous nodule, termed as Sister Mary Joseph nodule. Local invasion into the duodenum, pancreas, and retroperitoneum is also characteristic. In such cases efforts are usually focused on chemotherapy or radiation therapy and palliative care. However, surgical resection remains the preferred treatment for gastric carcinoma.5

Lymphangiogenesis is a critical process for tumor growth, invasion and metastasis.  That is why measurement of lymph vessel density may be clinically important in gastric cancer specimens. The induction of lymphangiogenesis by a tumor is controlled process, influenced by lymphangiogenic factors, which involve a complex interaction between tumor and endothelial cells.6 The pathologic approach to assess lymphangiogenesis involves microscopic estimation of lymph vessel density by using endothelial markers in immunohistochemistry.

On previous studies detection of lymphangiogenesis was limited due to lack of specific lymphatic endothelial markers. The recent discovery of anti-Podoplanin antibody has a major impact on lymphatic studies. Podoplanin has become a valuable marker for detecting lymph vessels and for identifying lymph vessel invasion in tumor.7

Podoplanin is a specific marker of the lymphatic endothelium and is not expressed in blood vessels. It is expressed by both developing and mature lymphatic endothelial cells and seems to be a more specific marker of lymphatic endothelial cells.  By electron microscopy and immunoelectron microscopy, it was demonstrated that Podoplanin is mainly expressed on the luminal surface of lymphatic endothelial cells.  Peritumoral (PT) lymphatic vessels are larger and more irregular than the intratumoral (IT) lymphatic’s, with a significantly lower density. Intratumoral lymphatic vessels are found in a large variety of tumors and are usually small, flattened and irregular. It was suggested that Podoplanin has potential role in invasion and metastasis of tumor. This hypothesis is mainly based on the observation that high expression of Podoplanin is consistently correlated with the presence of metastases. It was reported that Podoplanin expressing cells were found at the invasion.8 The use of an anti-Podoplanin-based therapeutic strategy could be suggested in the treatment of lymphatic metastases based on three considerations: (i) Podoplanin is a well-known marker of lymphatic endothelial cells; (ii) its expression seems to be associated with bad prognosis and high risk for lymph node metastases; (iii) it is involved in tumor invasion. In conclusion, Podoplanin is a sensitive marker of lymphatic endothelial cells and is very useful in evaluating lymphatic micro vessel density. Immunohistochemical detection of Podoplanin is helpful in the diagnosis of lymphovascular invasion.8 Some studies have suggested that lymph vessel density is associated with an increased risk of lymph node metastasis;9 however, this conclusion is not supported by all of the published studies.10












The study was conducted a) to investigate morphological prognostic factors of gastric adenocarcinoma, b) to estimate lymph vessel density by using immunostain Podoplanin and c) to correlate lymph vessel density with prognostic factors.


It was a cross sectional observational study carried out from March, 2016 to July, 2018at the Department of Pathology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh. The study population was paraffin blocks of gastric adenocarcinoma from pathological laboratory in the same Institute with consecutive convenience sampling. Inclusion Criteria was paraffin blocks of gastric samples diagnosed as gastric adenocarcinoma in pathology laboratory and Exclusion criteria were with history of treated gastric carcinoma cases like neo adjuvant therapy, blocks containing autolyzed tissue, extensive necrosis, hemorrhage and desmoplasia, blocks having no tissue or damaged blocks and blocks containing other cancers like lymphoma, carcinoid tumor, malignant GIST etc. The sample size was 53. Monoclonal Anti-Human Podoplanin antibody, clone D2-40, Ready to use (code- IR072) was used for detecting lymph vessel. In this study, sections of normal vermiform appendix were taken as positive control (as recommended by Dako). Podoplanin antigen expression was detected by immunohistochemistry in the submitted blocks (paraffin blocks with maximum tumor bulk were chosen) and were performed using DAKO Cytomation at immunohistochemistry laboratory, department Pathology, BSMMU.

Histopathological categorization of tumor and grading of all the cases were done. Lymphovascular invasion was recorded. Each lymph node was histologically examined to determine metastasis. Numbers of lymph vessels were counted by Weidner’s method in peritumoral (PT) and intratumoral (IT) areas.11

 Micro vessel Quantification

Determination of lymph vessel density (LVD) were performed according to Weidner et al.11Brown staining of cytoplasmic membrane of endothelial cells of lymph vessels were considered as positive reaction. Lymph vessel density was calculated according to Weidner’s method by Olympus microscope model BH51. The immunostained sections were scanned by light microscopy at low magnification (10x) and the areas of tissue with the greatest numbers of distinctly highlighted lymph vessels (“hot spots”) were selected. So, “hot spot” is an area with the most dense lymph vessels growth. Five hot spots were selected in intratumoral (IT) and peritumoral (PT) areas. When the hot spots were defined, lymph vessel count were performed by counting the individual stained lymph vessel (at power 20x) representing a field size of 0.74mm2 (i.e. – 20x objective lens, 10x ocular lens; equivalent to 0.7386 mm2 per 200x field).11 The mean number of lymph vessels in each case were determined and considered as the lymph vessel density.7So, at first five hot spots were chosen in intratumoral (IT) and peritumoral (PT) area. In each hot spot, lymph vessel count was performed at power x20. Finally lymph vessel density was calculated as the mean of the total number of lymph vessels in these five hot spots. Lymph vessel density (LVD) assessed in this manner may be not equivalent to LVD seen in multiple areas of multiple samples of the stomach; however, it is well accepted that these hot spots, not arbitrary areas, are representative of the entire tumor. Although there is some variability in the distribution of lymph vessels within the layers of the gastric wall, the five ‘hot spot’ areas were chosen to obtain an objective assessment and to avoid observer variation.7


A total number of fifty three diagnosed cases of gastric adenocarcinoma were taken for this study. Forty six samples were partial/subtotal gastrectomy specimen and seven samples were total gastrectomy specimen. All fifty three samples contain epigastric lymph nodes ranging one to twenty five. Thirty seven cases had lymph node metastasis and the rest sixteen were free of tumor metastasis. Thirty seven cases were LVI positive in H&E routine stain and forty two cases were LVI positive with Podoplaninimmunostain.

In the group of 53 patients with gastric carcinoma, mean patients’ age was 49.87±11.38 years (range 20-70).33 patients ware male and 20 patients ware female with a male/female ratio of 1.6:1. According to Lauren’s classification, 30 cases ware diffuse type and 23 cases were intestinal type. Two third (60.4%) samples belonged to tumor size ≤ 5 cm. The mean tumor size was 5.61±1.92 cm, ranged from (3.0 – 11.0) cm. Lymph vessel density (LVD) was more frequent in cases having tumor size > 5 cm in diameter.

The histological grade was assessed according to WHO grading system. 33(62.3%) samples belonged to grade-III (poorly differentiation), followed by grade-II (moderate differentiation) (24.5%) and grade-I (well differentiation)(13.2%). Lymph vessel density (LVD) was most frequent in grade- II group.

Cases were grouped according to depth of invasion (tumor stage) as T1, T2, T3 and T4. 41.5% samples belonged to subserosal invasion (T3). T3 was followed by T2(32.1%), T4 (20.8%) and T1(5.7%). Lymph vessel density(LVD) was most frequent in T3 group.

Numbers of lymph node ranged from 1-25. Cases were grouped according to the numbers of lymph node metastasis as N0, N1, N2 and N3. 30.2% samples had no lymph node metastasis (nodal stage N0). 28.3% samples belonged to nodal stage N1, followed by nodal stage N2 (20.8%) and N3 (20.8%). Nodal stages were increasing with increase of Lymph vessel density (LVD).

Lymph vessels in intratumoral (IT) area ranged from 1-12 and peritumoral (PT) area ranged 2-11. Mean LVD in IT area was 5.93 ±2.79 cm and mean LVD in PT area was 5.50±2.38 cm.

A positive correlation was found between Lymph vessel density(LVD) and tumor stage in intratumoral (IT) area (Figure 7).

The value of Pearson’s correlation coefficient was 0.377 and it was significant (p=0.005). Therefore, there was linear association between tumor stage and lymph vessel density (LDV) in intratumoral (IT) area with Podoplanin stain.

A positive correlation was found between Lymph vessel density(LVD) and tumor stage in peritumoral (PT) area (Figure 8).  The value of Pearson’s correlation coefficient was 0.334 and it was significant (p=0.014). Therefore, there was linear association between tumor stage and lymph vessel density (LVD) in peritumoral (PT) area with Podoplanin stain.

A positive correlation was found between Lymph vessel density (LVD) and Lymph node metastases inintratumoral (IT) area (Figure 9).

The value of Pearson’s correlation coefficient was 0.855 and it was significant (p=0.000). Therefore, there was linear association between lymph node metastasis and lymph vessel density (LVD) in intratumoral (IT) area with Podoplanin stain.

A positive correlation was found between Lymph vessel density (LVD) and Lymph node metastases in peritumoral (PT) area (Figure 10).

The value of Pearson’s correlation coefficient was 0.791 and it was significant (p=0.000). Therefore, there was linear association between lymph node metastasis and lymph vessel density (LVD) in peritumoral (PT) area with Podoplanin stain.

A positive correlation was found between LVD in IT and PT.

The value of Pearson’s correlation coefficient was 0.937 and it was significant (p=0.000). Therefore, there was linear association between LVD in intratumoral (IT) and peritumoral (PT) area with Podoplanin stain.


Gastric carcinoma is the most common leading tumor of the world. Although great efforts have been made in the field of early diagnosis and adjuvant therapy, the incidence and overall mortality of gastric carcinoma continues to increase. Since gastric tumor cells commonly infiltrate into the lymphatic system, lymph node status is routinely used to identify a patient’s prognosis, tumor stage, and treatment modality. Inhibition of tumor cells to lymph node metastasis (LNM) is a promising way to prevent distant metastasis. However, the relationship between lymphangiogenesis, lymphovascular invasion and lymph node metastasis (LNM) remains ambiguous.

This cross sectional study was carried out with an aim to estimate the density of expression of Podoplanin in lymphangiogenic vessel in gastric adenocarcinoma and to see the correlation between the percentages of area covered by lymphangiogenic vessels with prognostic factors. The present study findings were discussed and compared with previously published relevant studies.

In this study, 62.3% sample had grade-III (poorly differentiation) followed by 24.5% grade-II (moderately differentiation) and 13.2 % grade-I (well differentiation). Significant relation was found between lymph vessel density-intratumoral (LVD-IT) area (p=0.013) and lymph vessel density-peritumoral (LVD-PT) area (p= 0.006) with different histological differentiation. In their study, Pak et al., 201512 found that 53.3 % sample had grade-III and significant relation (p= 0.021) between LVD-IT area with histological differentiation, which is comparable with the current study.


It was observed that 41.5 % of samples belonged to T3, followed by 32.1% T2, 20.8% T4  and 5.7% T1 and significant relation found between LVD-IT area (p=0.019) and LVD-PT area (p=0.041) with different tumor stage. Gresta, Rodrigues Jr and Cabral, 201413 found 63.5% cases were T3, followed by 19.2% T2, 13.4% T1 and 3.8% T4. Pak et al., 2015 found 39.3% samples were T3 followed by 16.6 % T2 and T4. They also found significant association between LVD-IT area (p=0.024) with different tumor stage. Raica et al., 20088 found significant correlation between LVD with tumor stage (p<0.002) and Nakamura et al., 20067 found significant positive correlation between LVD with tumor stage (p=0.0008). So, present study result is consistent with previous study.

According to number of lymph node metastasis (LNM), it was observed that 30.2% sample belonged to nodal stage 0 (N0), followed by 28.3% stage-I (N1), 20.8% stage II (N2) and stage III (N3). There was significant relation was found between LVD-IT area (p<0.001) and LVD-PT area (p<0.001) with nodal status. Yan et al., 200814 reported that LVD was obviously higher in the colorectal cancer sample with metastasis (12.08 ± 4.96) than in those without (8.26 ± 4.08) (p<0.001). Schoppmann et al., 200115 demonstrated that lymph vascular invasion (LVI), assessed by anti-Podoplanin immunostain, has been strongly associated with presence of lymph node metastasis. Pak et al. 201512 found LVD-IT area was a higher than LVD-PT area (12.29± 4.36 vs. 11.01±3.62), Nodal stage N0 cases were more frequent and significant relation between LVD-PT in area (p=0.040) with nodal status. Nakamura et al., 20067 found significant positive correlation between LVD with nodal metastasis (p= 0.0094). Raica et al., 20088 found significant correlation between LVD with nodal metastasis (p<0.031) which is comparable with current study.

In this study, there was a positive significant Pearson’s correlation (r= 0.855, p= 0.000) was found between lymph node (LN) metastasis with LVD in IT area. Similarly, there was a positive significant Pearson’s correlation (r= 0.791 p= 0.000) was found between LN metastasis with LVD in PT area. Pak et al., 201512 found in their study that LVD in PT area was significantly associated with LN metastasis (p=0.040). Nakamura et al., 20067 found significant correlation between LN metastasis with LVD (p= 0.0094). Raica et al., 20088 reported that there was significant positive correlation between both LVD in IT and PT areas with LN metastasis (P<0.031). So current study result is consistent with previous study.

In this current study, there was a positive significant Pearson’s correlation (r= 0.377, p= 0.005) was found between tumor stages with LVD in IT area. Similarly, there was a positive significant Pearson’s correlation (r= 0.334, p= 0.014) was found between tumor stage with LVD in PT area. Nakamura et al., 20067 found significant positive correlation between tumor stage with LVD (p= 0.0008). Raica et al., 20088 reported that there was significant positive correlation between both LVD-IT area and LVD-PT area with tumor stage (P<0.02).

In this current study, there was a positive significant Pearson’s correlation (r= 0.937, p= 0.000) was found between LVD-IT areas with LVD-PT areas. Pak et al., 201512 found significant correlation (p=0.028) between LVD-IT area with LVD-PT area.


Total fifty three cases of gastric adenocarcinoma were examined for lymphangiogenic vessels in peritumoral (PT) and intratumoral (IT) areas. Thirty seven cases had lymph node metastasis and the rest sixteen were free of tumor metastasis. Both peritumoral and intratumoral lymphangiogenic vessel count (density) stained by Podoplanin antibody correlated with lymph node metastasis and tumor stage. Lymphatic vessel count is more in the intratumoral area.  The specific lymphatic endothelial marker Podoplanin proved to be a valuable tool in highlighting lymph vessel density (LVD) and lymphovascular invasion, and therefore a predictor of lymph node metastasis.


The study population was selected from one institute in Dhaka city, so that the results may not be reflect the exact picture of the country.  The study period was short. Sample size was small. Podoplanin was only marker used to detect lymph vessels. Sometimes differentiation between blood vessels and lymph vessels were difficult in tissue sections. Differentiation between blood vessels and lymph vessels was also a limitation of this study.  Use of blood vessel immunostain like factor VIIIa, CD31, and CD 34 etc. could have overcome the limitation and overall no control was used.



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