European Journal of Cardiovascular Medicine

The prostate is a pearshaped glandular organ that weighs up to 20 grams in normal adult male and that depends for its differentiation and subsequent growth on androgenic hormones synthesized in the testis. Benign prostatic hyperplasia, prostatitis, prostatic carcinoma are the three principal conditions involving the prostate accounting for more than 95% of lesions. There are several benign proliferations of prostate which mimic malignancy and their awareness is essential to avoid diagnostic pit falls. The incidence of prostatic lesions, both benign and malignant increases with age and are rare before 40 years of age. High grade prostatic intraepithelial neoplasia and Atypical Adenomatous Hyperplasia (AAH) are now considered to be the most likely precursors of prostate cancer. Prostatic intraepithelial neoplasia lesions can only be diagnosed by histopathological examination of prostatic tissue and it is not possible to detect clinically by direct rectal examination, prostatic specific antigen assay or ultrasound. ^1,2,3

Prostate cancer is one of the leading causes of morbidity and mortality worldwide. Cancer of prostate is typically a disease of men over age 50. Africans and Americans have higher incidence than Japanese and Asians.

The incidence of prostatic carcinoma is 0.13% in Americans and 0.09% in Japanese and 0.10% in other Asian countries. In view of the increased life expectancy the absolute number of clinically significant prostate carcinomas will probably increase in the coming decades. Despite prostate cancer high morbidity its etiology remains obscure with only established risk factors being increasing age, race, hormones, dietary factors, obesity, physical inactivity, occupation, vasectomy, smoking, sexual factors and genetic susceptibility. In the hormones androgens play an important role in prostatic cancer.  ^4,5

In the view of increasing trend of the occurrence of both neoplastic and non-neoplastic lesions of the prostate in elderly, the current study was carried out at evaluating the histopathological features of various benign, premalignant and malignant lesions of prostate. Use of immunohistochemical markers in this study helps in arriving at diagnosis and to know prognosis and develop therapeutic strategies.

AIM

The prospective work “Histopathological study of prostatic lesions in correlation with immunohistochemistry is aimed to study the histopathological spectrum of various non neoplastic and neoplastic lesions of prostate among the specimens received in a period of August 2013 to September 2015 at Department of pathology, Siddhartha medical college, Vijayawada by using immunohistochemical markers p63 and P504s that are helpful in differentiating benign, pre-malignant and malignant lesions and to compare statistical data with similar other studies.

OBJECTIVES

1. To study the age wise distribution of various non neoplastic and neoplastic lesions of prostate.
2. To study association of prostatic intraepithelial neoplasia in different prostatic lesions.
3. To find out expression of immunohistochemical markers p63 and P504s in BPH, prostatic intraepithelial neoplasia and carcinoma cases.
4. To analyze and correlate the available clinical data with other studies

This is a prospective study done from August 2013 to September 2015 at Siddhartha Medical College Vijayawada. The TURP specimens were received to the Department of Pathology, Siddhartha medical college from Government general hospital, Vijayawada.

Inclusion criteria:

1. All neoplastic and non-neoplastic lesions of prostate were included in the study.
2. All TURP specimens were included in the study.

Exclusion criteria:

1. Patients below age of 40 years were excluded from the study.
2. All metastatic tumors of prostate were excluded from the study.

The clinical details of each case were recorded according to age, clinical features, per rectal examination findings, PSA levels and clinical diagnosis. The TURP specimens obtained were fixed in 10% buffered formalin for a minimum period of 12hrs and then entire bits were submitted for routine processing, fixation, dehydration, clearing and embedding in paraffin wax. Sections were prepared from the blocks for routine staining.

The nuclei and calcium were stained blue. Cytoplasm was stained pale pink. Erythrocytes, muscle, and eosinophil granules were stained bright red. The histopathological sections were reported accordingly as inflammatory, benign or malignant lesions. All malignant lesions were graded according to Gleason’s grading developed by Gleason in conjunction with the Veterans Administrative Cooperative Urological Research Group. Subsequently all sections were subjected to immunohistochemical staining.

IMMUNOHISTOCHEMICAL STAINING (67)

This is a technique of identifying cellular or tissue constituents (antigens) by means of antigen antibody interactions, the site of antibody binding being identified either by direct labeling of the antibody, or by use of secondary labeling method. The amino acid side chains of the variable domain of an antibody form a cavity which is geometrically and chemically complementary to single type of antigen epitope. The associated antigen and antibody are held together by a combination of hydrogen bonds, electrostatic interactions and Vander Waals’ forces.

Interpretation of p63 staining in prostate glands (59)

• Negative or
• Positive – complete
• Positive- partial (<25% of gland’s circumference is stained).

P504s STAINING:

The immunohistochemistry staining procedure steps were performed same as that of p63 staining except that here the antibody used was p504s polyclonal antibody and probe is not applicable for p504s staining. Cellular localization: Granular and cytoplasmic Positive control: Prostate cancer. Normal tissue: Not applicable

Interpretation of AMACR staining (66)

Positive staining pertains to dark diffuse or granular, cytoplasmic or luminal, but circumferential. The percentage positivity was graded from 0+ to 3+ as follows: -  0% cells

• - 0+, negative 1-10% cells
• - 1+, mild 11-50% cells
• - 2+, moderate
• >50% cells - 3+, strong

The clinical details of the cases like age, clinical history, clinical diagnosis was collected from the patients requisition forms. The histopathological findings were noted and the sections were submitted for immunohistochemistry and results were noted accordingly. The master chart was prepared by using all these details and statistical analysis was done. The obtained results were interpreted in form of tables, charts, bar diagrams and pie diagrams. Further the salient features of the present study were discussed and compared with the other similar recent studies. At last the salient points of the present study were given in the form of conclusion.

OBSERVATIONS AND RESULTS

The present study comprises of 82 cases of TURP specimens received in the department of pathology, Siddhartha medical college, Vijayawada during the period from August 2013 to September 2015. The clinical data was collected from patient’s requisition forms and recorded as per proforma. The microscopic and other findings were taken into proforma and master chart was prepared from these details. Overall analysis of the results was done by using tables, pie diagrams by obtaining data from master chart.

A total number of 82 cases were studied. The cases were distributed in the age group of 46-88 years. The maximum numbers of patients were in age group 60-69 years and the next common age group affected was 70-79 years. Out of 82 cases 68 were BPH, 13 were prostatic adenocarcinomas and 1 case was urothelial carcinoma. Foci of low-grade PIN was identified in 12 cases. All low-grade PIN foci were associated with BPH. High grade PIN was identified in 13 cases. Out of these 2 high grade PIN foci were seen in BPH and 11 were seen associated with adenocarcinoma. Miscellaneous features like cystic atrophy, chronic non-specific prostatitis, basal cell hyperplasia were also seen associated with these lesions.

Immunohistochemistry was done using p63 and P504S markers in cases of benign prostatic hyperplasia, prostatic intraepithelial neoplasia and carcinoma cases. All cases of BPH showed complete positivity for p63 immunostaining in the basal nuclei of the glands and all these glands of BPH were negative for P504S immunostaining. Out of 13 cases of high-grade PIN all cases of showed positivity for p63 stain in the basal cell nuclei of HGPIN glands and 12 cases showed moderate to strong positivity for P504S immunostaining. Out of 13 cases of adenocarcinoma of prostate all the cases showed negativity for p63 immunostaining and all cases showed strong cytoplasmic, granular positivity for P504S immunostaining. One case of urothelial carcinoma showed p63 positivity ( of score 5) and strong cytoplasmic positivity for P504S immunostaining.

Table 1: Distribution of cases according to Histopathological diagnosis

In the present study out of 82 cases there were 68 (82.93%) cases of BPH constituting the majority, 13 (15.86) cases were adenocarcinomas and 1 (1.21%) case of urothelial carcinoma.

Table 2: Distribution of cases according to age group

In the present study maximum number of patients i.e., 40 (48.78%) cases were in age group 60-69 yrs, 16 (19.52%) cases were in 70-79 yrs, 12(14.63%) cases were in 50-59 yrs, 12(14.63%) cases were in 80-89 yrs and 2(2.44%) were in 40-49 yrs.

Table 3: Age wise distribution of cases according to histopathological diagnosis

In the present study maximum number of carcinoma cases i.e.,8 (57.15%) were in age group of 80-89 yrs out of 14 cases and maximum number of BPH cases i.e.,34 (50%) were in age group of 60-69 yrs.

Table 4: Mean age (yrs) of cases according to histopathological diagnosis

In the present study all cases were distributed in age group of 46-88 yrs. The mean age for carcinoma is 78.42 and for BPH is 67.10.

Table 5: Distribution of Prostatic Intraepithelial Neoplasia in different cases

In the present study out of 13 cases of adenocarcinoma, 11(84.62%) cases showed HGPIN and 2(15.38%) cases were negative for PIN and out of 68 cases of BPH, 12(17.65%) cases showed LGPIN and 2(2.94%) cases showed HGPIN.

Table 6: Different microscopic patterns of HGPIN

There were 4 patterns identified in HGPIN usually with multiple patterns in each case. Tufting pattern was seen in 10(76.92%) cases, flat pattern was seen in 7(53.85%), micropapillary pattern was seen in 2(15.38%) and cribriform pattern was seen in 1(7.69%) case out of 11 cases of adenocarcinoma and 2 cases of BPH showing HGPIN. The common pattern identified was tufting type and the next common was flat type.

Table 7: Distribution of cases of Prostatic Adenocarcinoma according to Gleason’s Grading system

The Gleason score 5,7,8,9,10 constituted 1(7.69%) case, 2(15.38%) cases, 3(23.09%) cases, 5(38.46%) cases and 2(15.38%) cases respectively.

Table 8: Expression of p63 immunostaining in different cases

All the BPH and HGPIN cases showed positivity for p63 and all adenocarcinoma cases showed negativity for p63 immunostaining

Table 9: Expression of P504s immunostaining in different cases

All adenocarcinomas and 12(92.3%) cases of HGPIN showed P504S positivity and all BPH cases were negative for P504S immunostaining.

Figure:1Showing benign prostatic hyper plasia H& E (100x)

Figure: 2Benignglandsin BPH showing positivity forp63nuclearstainingIHC(400x)

Figure 3: Showing basal cellhyperpl Asia H&E (100x)

Figure4:Basalcellhyperplasiashowingpositivityforp63nuclearstainingIHC(400x)

Figure:5 Showing lowgrade PINH&E(400x)

Figure:6 High grade PIN-Tufting pattern H&E (400x)

Figure:7 High grade PIN-Flatty pe H &E (400x)

Figure:8 High grade PIN–Micro papillary pattern H&E(400x)

Figure:9 High grade PIN- Cribriform pattern H&E(400x)

Figure:10HighgradePINshowingpositivityforp63nuclearstainingIHC (400x)

Figure:11HighgradePINshowingpositivityforP504ScytoplasmicstainingIHC (400x)

Figure:12 Prostaticadeno carcinoma Gleason’ grade 2H &E (100x)

Figure: 13 Prostaticadeno carcinoma Gleason’s grade3H &E(100x)

Figure:14Prostaticadeno carcinoma Gleason’sgrade4H&E(400x)

Figure:15Prostaticadeno carcinoma Gleason’s grade5H&E(100x)

Figure:16 Adeno carcinoma showing positivity for P504Scytoplasmicstaining IHC (400x)