TRIM29 as a novel prostate basal cell marker for diagnosis of prostate cancer

Tripartite motif protein 29 (TRIM29) is one of the TRIM family proteins, some of which function as E3 ubiquitin ligases. In this study, we investigated the usefulness of TRIM29 for diagnosis of prostate cancer. Prostate tissues including carcinoma and non-carcinoma tissues obtained by needle biopsy and radical prostatectomy were used. Immunohistochemistry was performed according to standard procedures using an antibody against TRIM29. Immunohistochemical staining with an antibody against 34βE12, which recognizes cytokeratins 1, 5, 10 and 14, was performed as a control. Basal cells of normal prostatic glands were stained with anti-TRIM29 antibody in all cases, whereas prostate cancer tissues had no or little staining with anti-TRIM29 antibody. TRIM29 is selectively expressed in basal cells of the normal prostate gland, and immunohistochemical staining with anti-TRIM29 antibody showed the same expression pattern as that with 34βE12 in prostate cancer and its benign mimics. Our data indicate that TRIM29 may be useful for distinguishing prostate cancers from benign tissues.     

Cytokeratin immunohistochemistry as a diagnostic tool for distinguishing malignant from benign epithelial lesions of the prostate.

The basal cell layer (BCL) is believed to be absent in malignant but present in nonmalignant epithelial lesions of the prostate. Using the avidin-biotin-peroxidase complex (ABC) immunoperoxidase method, we examined the value of the monoclonal antibody cocktail MA-903, which stains selectively the prostatic BCL layer, in the distinction between benign and malignant epithelial lesions of the prostate. We immunostained histologic sections of 63 prostates, containing 235 morphologic appearances: normal prostate glands, 43; benign prostate hyperplasia (BPH), 59; basal cell hyperplasia (BCH), 24; adenosis, seven; prostatic intraductal neoplasia (PIN 1), 21; PIN 2, 25; PIN 3, 16; and cancer, 40. Some degree (continuous, continuous with focal disruption, and disrupted patterns) of basal cell staining was demonstrable in all normal and BPH, BCH, and PIN 1 lesions, but was absent in 39 of 40 cancers. However, not every gland in benign lesions stained positively. Further, two of 25 PIN 2 and six of 16 PIN 3 lesions failed to reveal BCL. Our results suggest that the presence or absence of BCL, predicated on cytokeratin MA-903 immunoreactivity, may be a useful indicator in the distinction between benign and malignant epithelial lesions of the prostate.     

Immunohistochemistry of a prostate membrane specific protein during development and maturation of the human prostate

An antiserum against secretory vesicles from human seminal fluid (prostasomes) was used to study the localisation and distribution of the respective antigen(s) during prenatal development and pubertal maturation of the human prostate. The crude antiserum stained both secretory and membrane proteins in the adult prostate and other glands, such as pancreas and parotid gland. An immunoaffinity purified fraction from the antiserum selectively reacted with the apical plasma membrane of prostatic epithelium adluminal cells, recognizing a 100 kDa antigen (PMS). Even in the earliest stages of embryonic prostate specimens studied, the adluminal plasma membrane of the epithelial cells from developing glandular anlagen reacted strongly. The occurrence of PMS immunoreactivity in prostatic anlagen was directly correlated with lumen formation. As the antigen is an androgen-independently synthesised membrane protein of the prostate, it may possibly be used as a marker of cell polarity in the normal and pathologically altered prostate.     

Immunohistochemical localization of FAP-1, an inhibitor of Fas-mediated apoptosis, in normal and neoplastic human tissues

Fas, a death receptor, is widely expressed in human tissue, but its expression, although a prerequisite for the induction of apoptosis, does not predict its biological function. To understand the mechanisms of Fas resistance in human tissues in vivo, we performed immunohistochemistry using an antibody against Fas-associated phosphatase-1 (FAP-1), which interacts with the cytosolic domain of Fas and inhibits Fas-mediated apoptosis. In normal human tissues, FAP-1 immunostaining was easily detected, for example, in renal tubules, skeletal muscle, myocardiocytes, pituitary gland, parathyroid gland, pancreatic islets, hepatocytes, testicular germ cells, prostatic glands, neurons, epithelium of fallopian tube, endometrial glands, trophoblasts, bronchial epithelial cells, and some types of gastrointestinal epithelial cells. In 123 (78%) of 158 cancers of various origins, including breast carcinomas, stomach carcinomas, colon carcinomas, lung carcinomas and several types of sarcomas, variable intensities of FAP-1 expression were evident. Taken together, these findings demonstrated that FAP-1 is widely expressed in normal human tissues and partly overlapped with Fas expression described in earlier reports, suggesting that FAP-1 may have an important role in the regulation of apoptosis in vivo. In addition, FAP-1 expression in cancers suggests that many cancers may be resistant to Fas-mediated apoptosis through the action of FAP-1 in vivo.     

Expression of epidermal growth factor receptor and ERBB2 (HER-2/Neu) oncoprotein in prostatic carcinomas.

Expression of epidermal growth factor receptor (EGFR) and ERBB2 oncoprotein were studied in paraffin-embedded normal (n = 2), hyperplastic (n = 17), and malignant (n = 147) prostatic tissues by immunohistochemistry. Strong immunoreactivity was detected in the epithelial cells of all normal and hyperplastic prostates with a new EGFR antibody (Mab31G7). In prostatic carcinomas, the EGFR immunoreactivity was variable with 47% showing uniform, 39% partial, and 14% no staining. Tumors with partial or uniform EGFR immunoreactivity were locally more advanced and of higher histological grade than the EGFR-negative tumors. EGFR-positive tumors also had two to three times higher S-phase fraction, suggesting that EGFR expression conferred proliferative advantage. Patients who had either partially or uniformly EGFR-positive carcinomas had a worse 10-yr progression-free (p = 0.05), overall (p = 0.03), and prostatic carcinoma-specific (p = 0.007) survival than those with EGFR-negative carcinomas. However, according to a multivariate analysis, EGFR did not have independent prognostic value. None of the normal, hyperplastic, or malignant prostate tissues showed clearly positive ERBB2 immunoreactivity with MAb1 antibody.     

Cytokeratin and vimentin expression in normal and neoplastic canine prostate

Intermediate filament expression in the canine prostate, unlike that in human prostate, is represented in the literature by only a few reports. In this study, the expression of cytokeratin (CK) and vimentin was examined in three normal canine prostates and 11 canine prostatic carcinomas. Monoclonal antibodies directed against vimentin, CK AE1/AE3, CK 18-8 (for luminal epithelial cells), CK 5, CK clone 8.12 and CK 14 (for basal cells) were employed. As in man, normal canine prostatic luminal cells were positive for CK 8-18. Basal cells were positive for CK 5 and CK clone 8.12 but, in contrast to findings in man, were negative for CK 14. Luminal cells were vimentin-negative, whereas in man they have been reported as vimentin-positive. The majority of carcinomas showed an undifferentiated histological pattern and all were positive for CK AE1/AE3 and for vimentin. Ten tumours were positive for CK 8-12, but six of them showed many cells co-expressing CK 14. Moreover, in two of these six cases a large number of neoplastic cells also reacted with CK clone 8.12 antibody, and in one of them co-expression of CK 5 was detectable. This co-expression, of luminal and basal cytokeratins, suggests a possible origin of the tumours from prostatic epithelial stem cells. Vimentin expression is an inconstant finding in human prostatic carcinomas; its almost uniform occurrence in canine carcinomas suggests a lesser degree of differentiation than in the human neoplasm.     

Human antibodies targeting cell surface antigens overexpressed by the hormone refractory metastatic prostate cancer cells: ICAM-1 is a tumor antigen that mediates prostate cancer cell invasion

Transition from hormone-sensitive to hormone-refractory metastatic tumor types poses a major challenge for prostate cancer treatment. Tumor antigens that are differentially expressed during this transition are likely to play important roles in imparting prostate cancer cells with the ability to grow in a hormone-deprived environment and to metastasize to distal sites such as the bone and thus, are likely targets for therapeutic intervention. To identify those molecules and particularly cell surface antigens that accompany this transition, we studied the changes in cell surface antigenic profiles between a hormone-sensitive prostate cancer line LNCaP and its hormone-refractory derivative C4-2B, using an antibody library-based affinity proteomic approach. We selected a naïve phage antibody display library to identify human single-chain antibodies that bind specifically to C4-2B but not LNCaP. Using mass spectrometry, we identified one of the antibody-targeted antigens as the ICAM-1/CD54/human rhinovirus receptor. Recombinant IgG1 derived from this single-chain antibody binds to a neutralizing epitope of ICAM-1 and blocks C4-2B cell invasion through extracellular matrix in vitro. ICAM-1 is thus differentially expressed during the transition of the hormone-sensitive prostate cancer cell line LNCaP to its hormone-refractory derivative C4-2B, plays an important role in imparting the C4-2B line with the ability to invade, and may therefore be a target for therapeutic intervention.     

Expression of vascular endothelial growth factor (VEGF) and VEGF receptor Flk-1 in benign, premalignant, and malignant prostate tissue.

Vascular endothelial growth factor (VEGF) is one of the most potent mitogenic, highly specific tumor angiogenic factors, which acts via binding to 2 specific tyrosine kinase receptors. There are few studies analyzing VEGF receptor expression in prostate cancer cells, and results are contradictory. In an immunohistochemical study, we analyzed VEGF and VEGF receptor fetal liver kinase (Flk)-1 expression in benign glands, high-grade prostatic intraepithelial neoplasia (HGPIN), and prostatic carcinomas of different Gleason scores, obtained from 21 radical prostatectomy specimens. In all benign glands, VEGF and Flk-1 expression was confined almost exclusively to the basal cell layer (proliferative cell compartment). In HGPIN, labeling was no longer confined to the basal cell layer, but also was seen in all neoplastic secretory cells. All carcinomas stained positive for both markers. There was a trend for increasing labeling intensity with increasing cellular dedifferentiation. We concluded that tumor growth stimulated by the VEGF-Flk-1 system is promoted not only by neoangiogenesis, but also by tumor cell autostimulation. The VEGF-Flk-1 system may have an important role in the process of malignant transformation and tumor progression.     

Monoclonal antibody (F5) to human prostate antigen

Hybridoma culture F5 has been developed which secretes monoclonal antibody (McAb) directed to an epitope of a prostatic glycoprotein of Mr 34 kD (Prostate Antigen, PA). Tissue levels of PA have been evaluated using a competitive-binding enzyme-immunoassay based upon the inhibition of McAb binding activity to purified antigen. Results indicated the specific occurrence of high antigen concentrations in extracts prepared from prostatic tissues. The antigenicity of epitope F5 is resistant to tissue fixation and embedding protocols, and has been demonstrated upon immunoperoxidase staining procedures. Immunoperoxidase data strongly indicate that McAb F5 possesses a singular specificity towards prostatic epithelial cells. Other tissues, whether normal or cancerous, fail to express this determinant. Specimens examined included epithelial and nonepithelial tissues along with a panel of carcinomas and sarcomas. The antibody was able to detect tumor cells at extra-prostatic sites and represents a powerful probe for the detection and differential diagnosis of metastatic cancer of the prostate.     

Basal epithelial cells of human prostate gland are not myoepithelial cells. A comparative immunohistochemical and ultrastructural study with the human salivary gland.

The hypothesis that basal epithelial cells of the human prostate are of myoepithelial origin was investigated using immunohistochemical and ultrastructural methodologies. The immunohistologic analyses show significant phenotypic differences between prostatic basal cells and myoepithelial cells of the salivary gland. Although both cell types stain intensely with the 312C8-1 monoclonal antibody, only true myoepithelial cells demonstrated significant amounts of muscle-specific actin as decorated by the HHF35 monoclonal antibody. Furthermore, using double-labeling experiments, the prostatic basal cells were strongly decorated with a fluorescein-tagged basal cell-specific keratin but were negative with the rhodamine-tagged phalloidin, a chemical that binds specifically to actin microfilaments. Ultrastructural studies also showed an absence of thin microfilament bundles, dense bodies, and micropinocytotic vesicles in the prostatic basal cells. The current investigations show that the prostatic acini do not have a basal myoepithelium. Although some authors have suggested a stem cell role for prostatic basal cells, the weight of experimental work argues against this hypothesis. The exact role of the basal epithelial cells of the prostate is not known, although they may serve endocrine, paracrine, or other regulatory functions and may be involved in modulating signals between prostatic stroma and epithelium.     

AMACR/34βE12/p63鸡尾酒双染对诊断前列腺癌及癌前病变的价值

目的探讨AMACR／34βE12／p63鸡尾酒双染对诊断小灶性前列腺癌及癌前病变的价值。方法从2005年6月起对3个月内临床连续送检的105例前列腺穿刺活检标本,6例前列腺癌根治标本和19例经尿道和耻骨上摘除的良性前列腺增生标本总计130个病例,1030个组织块中需要用免疫组织化学辅助诊断的262个组织块分别作AMACR／34βE12／p63鸡尾酒双染,同时作这3个抗体的单项染色,并结合HE切片和临床资料观察结果作出诊断。结果鸡尾酒双染切片中前列腺癌和高级别上皮内瘤变（HGPIN）上皮细胞呈蓝黑色,良性腺体的基底细胞呈红色。癌的蓝黑色腺上皮周围无红色基底细胞围绕,HGPIN的蓝黑色腺上皮周围有间断或连续的红色基底细胞。共在214个（82％）组织块中发现前列腺癌,包括31处小灶性癌。64个（24％）组织块中发现HGPIN,包括局灶性HGPIN和小腺泡性HGPIN。1个组织块有不典型性腺瘤样增生。未发现上皮细胞AMACR阳性同时基底细胞34βE12和p63阴性的良性腺体。结论鸡尾酒双染有助于提高小灶性前列腺癌和HGPIN检出率。     

Immunohistochemical investigation of different cytokeratins and vimentin in the prostate from the fetal period up to adulthood and in prostate carcinoma

From the fetal period up to puberty the immature epithelium of the prostate glands, the prostatic ducts, the ejaculatory ducts and the seminal vesicles as well as the urothelium of the prostatic urethra are extensively positive for different keratin antibodies (antibody against keratins from human stratum corneum, broadly reacting antibody "AE1 and AE" and antibodies against the keratins 7, 8, 18 and 19) immunohistochemically. The epithelium of the ejaculatory ducts and seminal vesicles in addition regularly exprimates vimentin which is found in the epithelium of the prostate glands focally. During puberty, the immature epithelium of the prostate glands differentiates into the two cell types basal cell and secretory epithelium which differ immunohistochemically: Keratins from human stratum corneum are exclusively demonstrable in the basal cells, the keratins 8 and 18 only in the secretory epithelium. For keratin 7, 19 and the antibody "AE1 and AE3" both cell types are positive. Keratin 7 is demonstrable only focally. The secretory epithelium partly co-exprimates keratins and vimentin. Prostatic carcinomas of different grades virtually contain no keratins from stratum corneum. All other keratins are found in variable extension in the vast majority of the tumors independent of the differentiation. Vimentin is positive mostly focally in about 50% of the tumors. Prostatic carcinoma and the secretory epithelium of the prostate glands share identical immunohistochemical features and differ from the basal cell by several markers. This indicates that prostatic carcinoma rather derives directly from the secretory epithelium than from the basal cell.     

Immunohistochemistry of a gross cystic disease fluid protein (GCDFP-15) of the breast. A marker of apocrine epithelium and breast carcinomas with apocrine features.

Gross cystic disease fluid is a pathologic secretion from breast composed of several glycoproteins, including a unique 15,000-dalton monomer protein, GCDFP-15. By the immunoperoxidase technique, GCDFP-15 was localized in the apocrine metaplastic epithelium lining breast cysts and in apocrine glands in the axilla, vulva, eyelid, and ear canal. In normal breast tissue, a few individual epithelial cells within lobules and small ducts were focally positive for GCDFP-15. Fourteen of 30 breast carcinomas stained positively for GCDFP-15. Of 16 carcinomas with apocrine features, 12 stained positively. Benign and malignant lesions from other tissues, including lung, colon, ovary, endometrium, stomach, prostate, liver, esophagus, and kidney, revealed no immunoreactivity. The only cells of "non-apocrine" tissues that contained GCDFP-15 were serous cells of the submandibular salivary gland, submucosal glands of the bronchi, and accessory lacrimal glands. Phylogenetically, these tissues have biologic features in common with apocrine glands. This report is the first to characterize GCDFP-15 as a specific tissue marker of apocrine epithelium.     

Investigations of the estrogen (ER-ICA-test) and the progesterone receptor in the prostate and prostatic carcinoma on immunohistochemical basis

Summary Estrogen (ER) and Progesterone receptors (PR) were demonstrated immunohistochemically on frozen sections from 11 prostatectomy and 7 cystoprostatectomy specimens in the nuclei of various cell types. The periglandular fibrocytes and smooth muscle cells were extensively positive, the interglandular stromal cells were only partly so. Normal basal cells stained focally positive, hyperplastic basal cells stained extensively. The glandular secretory epithelium and atrophic glands were negative. The same findings were obtained in hyperplastic nodules. Both ER and PR also occurred in the urothelium of central prostatic ducts and of the prostatic urethra. The fibrous stroma around the ejaculatory ducts and seminal vesicles was extensively positive while the epithelium was negative. The smooth musculature of the seminal vesicles was only partly positive. On large field sections, the ER as well as the PR were numerically equally distributed throughout the inner zone of the prostate and the prostate proper. 12 prostatic carcinomas (G I–G III) were ER- and PR-negative. Estrogens may contribute to nodular hyperplasia by triggering a stromal proliferation with a secondary inductive epithelial growth. Obviously they do not act directly on prostatic carcinoma but inhibit growth via the hypophyseal-testicualr axis. The biological significance of the PR in the prostate is unknown.     

Low-grade mucoepidermoid carcinoma of salivary glands: characteristic immunohistochemical profile and evidence of striated duct differentiation

The purpose of the present study is to determine the presence and distribution of epithelial and myoepithelial cells in mucoepidermoid carcinoma (MEC) of salivary glands and to compare them with normal salivary gland tissue and other primary carcinomas. This is in order to establish novel diagnostic criteria and to better understand MEC histogenesis. Formalin-fixed paraffin-embedded tissues from ten well-differentiated MECs, three adenoid cystic carcinomas (ACC), four acinic cell carcinomas (AC), and three epithelial-myoepithelial carcinomas (EMCC) of salivary glands were studied with immunohistochemistry using antibodies that recognise antigens indicative of epithelial and myoepithelial cell differentiation. An anti-mitochondrial antibody was also employed. Normal salivary tissue was present for comparative study in non-tumorous areas of the same section from 12 cases. MEC contained numerous keratin-positive cells. Anti mitochondrial antibody was diffusely positive in all ten of these tumours. Smooth muscle actin, h-caldesmon, and smooth muscle heavy chain myosin, which are indicative of myoepithelial cell differentiation, were negative. Rare cells in only one case were stained by calponin. Cytokeratin 14 (CK14) and anti mitochondrial antibody stained cells located mainly at the periphery of neoplastic nests and cystic spaces, while CK7 was mainly present in cells bordering gland lumina (zoning pattern). The immunohistochemical cell profile was similar to that seen in striated normal ducts. All others tumours studied showed a different immunohistochemical pattern, mostly consisting of a lack of mitochondrion-rich cells and the presence of myoepithelial cells in ACC and EMCC. Immunoreactivity in MEC for CK7, CK14 and mitochondrial antibodies appears as a peculiar pattern of staining, different from that of other salivary gland tumors; this seems helpful for diagnostic purposes. In addition, a differentiation of the "striated duct phenotype" is suggested.     

Morphological and immunohistochemical characteristics of the heterogeneous prostate-like glands (paraurethral gland) seen in female Brown-Norway rats.

Morphological and immunohistochemical characteristics of the prostate-like glands (paraurethral gland) seen spontaneously in female Brown-Norway (BN) rats were investigated by gross, and light and electron microscopic examination. At 9- to 10-weeks-old, the paraurethral gland was detected in 50 out of 52 female animals examined (96.2%), and it was observed as single or paired structures located ventrolaterally in the urethra just caudal to neck of the urinary bladder. Microscopically, the glandular acini consisted of flat to cuboidal secretory epithelium surrounded by the smooth muscle. The glands displayed modest secreting activity, and a few secreting materials were observed in the acinar lumens. The main peripheral ducts were located in the urethral wall, and drained into the urethra on both sides. Ultrastructurally, abundant rough endoplasmic reticulum (RER), numerous mitochondria and lysosomes, and secretory granules in the apical portion of the epithelial cells were noted, and basal cells were also observed. These gland epithelial cells showed positive reactions when stained for androgen receptor (AR), prostate specific antigen (PSA), or prostate specific acid phosphatase (PSAP). The nature of this paraurethral gland resembled that of the prostate gland in male rats. Thus, the paraurethral glands seen in the females were considered homologous to the prostate gland in males.     

Increased Id-1 expression is significantly associated with poor survival of patients with prostate cancer

The levels of Id-1 (inhibitor of DNA binding or inhibitor of cell differentiation) expression in a series of prostate cell lines and in an archival set of prostate tissues were examined. Western blot analysis showed that the level of Id-1 expressed in the androgen sensitive cell line LNCaP was 1.2 +/- 0.2 times that detected in the benign cell line PNT-2. The level of Id-1 increased further to 1.8 +/- 0.2 and 2.9 +/- 0.3 in the androgen-insensitive cell lines Du-145 and PC-3, respectively. Immunohistochemical staining with Id-1 antibody performed on 113 cases of prostate tissues showed that among the 7 normal cases, 6 (86%) stained either negative or weakly positive whereas only 1 (14%) stained moderately positive. Among the 36 benign prostatic hyperplasia (BPH) samples, 34 (94%) stained either negative or weakly positive; only 1 (3%) stained moderately and 1 (3%) stained strongly. Of the 70 carcinomas, 8 (11.5%) stained weakly, 34 (48.5%) stained moderately, and 28 (40%) stained strongly positive. The intensity of Id-1 staining in carcinomas was significantly stronger than that detected in the normal prostate and BPH (chi(2) test, P < .001) and it was significantly increased as the increasing malignancy of carcinomas measured by Gleason score (chi(2) test, P < .001). The intensity of Id-1 staining was partially associated with the levels of prostate-specific antigen, but not related to the level of androgen receptor. Kaplan-Meier survival curve analysis showed that, similar to Gleason scores, overexpression of Id-1 was significantly associated with the reduced length of patient survival (log-rank test, P = .01). These results suggest that Id-1 is a useful prognostic marker to predict the outcomes of patients with prostate cancer.     

CD109, a new marker for myoepithelial cells of mammary, salivary, and lacrimal glands and prostate basal cells

The CD109 gene encodes a glycosylphosphatidylinositol (GPI)-anchored cell surface protein. Herein it is shown that CD109 is highly expressed in myoepithelial cells of mammary, salivary, and lacrimal glands; and in prostate basal cells. The anti-CD109 antibody generated by the authors was available for formalin-fixed paraffin section, and it strongly stained myoepithelial cells and basal cells but not ductal, acinar, and secretory cells in these glands. CD109 expression was negative in examined breast ductal carcinomas and prostate adenocarcinomas. These findings indicate that CD109 is a useful marker for the diagnosis of invasive breast and prostate carcinomas.