Percentage Of Immunoreactive Cells Research Articles

Expression analysis of leptin in nephrogenesis and renal carcinogenesis

The complex phenomenon of renal development involves several signaling molecules. Any alteration in the developmental process could largely influence the organogenesis, thus predisposing to several adulthood diseases. Leptin is a pleiotropic gene secreted by adipose cells. However, the purview of its actions is much beyond merely the extent of human adipose reserve. It regulates several cellular mechanisms such as cell proliferation, inflammation, vasculogenesis, and the production of collagen fibres. Further leptins are thought to play a putative role in embryogenesis and tumorigenesis. The aim of thIS study was to investigate the localization of the protein in fetal and cancer kidney in an attempt to understand the role of the protein in fetal kidney development and in renal cancer. Leptin expression was evaluated by subjecting tissue sections from paraffin-embedded blocks of renal tissues (fetal, adult and cancer) to immunohistochemistry staining. The tissues were scored based on the staining pattern and percentage of immunoreactive cells. The tissues were subjected to haematoxylin and eosin stain before performing immunohistochemistry. The images were analyzed and photographed. Mild staining for leptin was observed in the tubules of fetal and adult kidneys. Mild to moderate staining was seen in membranes of renal cell carcinoma tissues. It appears such that leptin may not be a key factor or rather a temporary factor in the developmental process of the kidney. The low levels of leptin in normal adult renal tissues may be physiologically significant. The role of leptin in the renal cell carcinoma progression is sceptible.

Effects of Cortactin Expression on Prognosis in Patients with Breast Cancer.

Cortactin is overexpressed in several types of invasive cancers. However, the role of cortactin expression in breast cancer prognosis has not been sufficiently elucidated. Therefore, we investigated the clinicopathological significance of cortactin in breast cancer. Tissue microarrays were prepared from a cohort of 506 patients with breast cancer, and cortactin expression was evaluated using immunohistochemistry. The cortactin immunoreactivity score (IRS) was quantified as the product of the intensity score and the percentage of immunoreactive cells. Cortactin expression was classified as low or high using the IRS (IRS ≤ 4 as a cortactin-low value and IRS > 4 as a cortactin-high value). We compared cortactin expression and clinicopathological factors according to the molecular subtypes of breast cancer. Of 506 breast cancer cases, 333 and 173 showed high and low cortactin expression, respectively. Of the 333 patients with high cortactin expression, 204, 58, and 71 had luminal, HER2, and triple-negative breast cancer (TNBC), respectively. In the univariate and multivariate analyses of patients with TNBC, cortactin expression was found to be a significant prognostic factor for overall survival (OS). However, in all patients with non-TNBC, cortactin expression had no significant association with prognosis or overall survival. Survival curves revealed that among patients with TNBC, the high-cortactin group had a better prognosis in disease-free survival and OS. Cortactin expression may be a good biomarker for predicting the prognosis of patients with TNBC.

ID PROTEIN EXPRESSION PATTERN IN MEDULLOBLASTOMA

Id proteins, also known as inhibitors of DNA binding or inhibitors of differentiation, are upregulated in a variety of neoplasms, especially in conjunction with high-grade, poorly differentiated tumours, whereas differentiated tissues have little to no Id expression. The helix-loop-helix (HLH) family of transcription factors, which the four Id genes are part of, controls transcription by binding to and sequestering HLH complexes. By doing immunohistochemistry utilising a medulloblastoma tissue microarray with 44 distinct medulloblastomas and 10 normal control cerebella, and testing antibodies specific for Id1, Id2, Id3, and Id4 it was hypothesised that Id proteins are overexpressed in medulloblastomas. A semi-quantitative staining score that considered staining intensity and the percentage of immunoreactive cells was implemented. Id1 was not found in medulloblastoma or normal cerebella, but it was strongly expressed in the endothelial nuclei of tumour arteries in 77% of tumours. In medulloblastoma, Id2 expression was more prominent than in normal cerebella (median staining score: 3). Id3 expression was seen in some neurons of the growing cerebellar cortex, but it was dramatically upregulated in tumour endothelial cells and medulloblastoma (median labelling score: 11). Id4 was not expressed in either tumour cells or normal cerebella. By modifying the expression of essential cell cycle regulatory proteins in favour of cell proliferation, Id2 or Id3 overexpression induced proliferation in medulloblastoma cell lines. This research demonstrates that enhanced Id2 and Id3 expression in medulloblastoma may play a role in tumour cell proliferation and survival and that Id1 expression in endothelial cells may contribute to angiogenic processes.

​Histomorphological and Immunohistochemical Studies on Ovaries of Crossbred Dairy Cows with Ovarian Hypoplasia

Background: Anatomical abnormalities of the ovaries can be responsible for a variety of reproductive problems, ranging from subfertility to sterility. The current study was aimed to evaluate the ovaries for histological variations and for the identification of Insulin-like Growth Factor Binding Protein - 2 (IGFBP-2) by Immunohistochemistry. IGFBP-2 was produced by ovarian tissues and was subjected to dynamic changes during follicle growth, differentiation and degeneration. Methods: The study was conducted on the genitalia of 100 dairy cows / heifers collected from Meat Technology Unit, Mannuthy. Out of these 100 animals, three showed ovarian hypoplasia. Tissue samples from different regions were fixed in 10 per cent neutral buffered formalin. Standard procedures were adopted for histoarchitectural studies and samples were subjected to immunohistochemistry. Result: Hypoplastic ovaries were appeared as small, wrinkled, flattened and elongated structures. Histologically, the ovary was not demarcated into a cortex and medulla distinctly. The ovaries were mostly devoid of ovarian follicles and corpora lutea. In partial hypoplastic ovaries, higher number of atretic follicles were noticed when compared with the normal ovary. Some of these atretic follicles showed a thickened zona pellucida. The granulosa cells surrounding the oocyte were degenerated. Tissue samples from the animals with ovarian hypoplasia showed a strong immunoreaction (for IGFBP-2) in the primordial and atretic follicles. The staining intensity and percentage of immunoreactive cells was comparatively higher than that of the control group. More number of IGFBP-2 blocked the positive effect of FSH and suppress steroidogenesis. The overall percentage of positive immunoreactive cells was more than 50 per cent with strongly positive staining intensity.

Angiogenesis and proliferation of endothelial cells in hypertrophic and nodular port-wine stain

BackgroundPort-wine stain (PWS) has been classified not as the hyperplasia of cells, but rather, as an expansion of malformed vessels. However, previous studies have reported upregulated expression of proangiogenic factors in PWS. Several studies have indicated that the pathology exhibits proliferation of numerous endothelial cells in hypertrophic/nodular PWS. This study aimed to determine the expression of vascular epithelial growth factor (VEGF), matrix metalloproteinase-9 (MMP-9), angiopoietin-2 (ANG-2), and basic fibroblast growth factor (bFGF) in hypertrophic PWS. MethodsImmunohistochemistry was used to analyze skin samples from 33 patients with hypertrophic PWS. Expression levels of VEGF, MMP-9, ANG-2, and bFGF in hypertrophic PWS were determined by multiplying the intensity by the percentage of immunoreactive cells. Immunoreactivity scores were classified as follows: negative (0), low (1), moderate (2, 3, and 4), or high (6). ResultsBased on pathological characteristics, hypertrophic PWS was divided into vascular malformation and pyogenic granuloma (PG) types. VEGF, MMP-9, ANG-2, and bFGF were significantly activated in the blood vessels of PG-type PWS samples compared with their counterparts in blood vessels of vascular malformation-type PWS samples and controls. PG-type hypertrophic PWS, which exhibited proliferation of endothelial cells, showed the strongest activation. ConclusionThe exuberant proliferation of endothelial cells in PG-type hypertrophic PWS may be associated with the regulation of proangiogenic factors during development. These proangiogenic factors that function in the angiogenesis and proliferation of endothelial cells may play an important role in the pathogenesis and progression of PWS. Furthermore, these factors may be dynamic and behave differently in various types of hypertrophic PWS.

Nicotinamide N-Methyltransferase Expression in Cervical Adenocarcinoma and Its Clinical Significance

Nicotinamide N-methyltransferase (NNMT) is a metabolic enzyme increasingly implicated in tumor progression and resistance to treatment. It is overexpressed in human cancers including glioblastoma, gastric, pancreatic, lung and colorectal cancers. The role of NNMT in cervical adenocarcinoma has not been studied thus far. We aimed to evaluate expression of NNMT in cervical adenocarcinoma and investigate its prognostic significance. NNMT expression was assayed in 18 cases of cervical adenocarcinoma and 19 cases of benign cervical tissue. NNMT immunoreactivity was scored by multiplying staining intensity with percentage of immunoreactive cells. The expression of NNMT was significantly higher in cervical adenocarcinoma than in benign tissue (p = 0.024). NNMT expression was significantly higher in patients with locally advanced than early cervical cancer (p = 0.031). NNMT expression was higher in patients with tumor size greater than 4 cm and in those with poorly differentiated tumors compared to patients with tumors of 4 cm or less and well-moderate differentiation, respectively (p = 0.044, p = 0.014). Although on a preliminary level, this is the first study to detect an overexpression of NNMT in cervical adenocarcinoma and an increased expression associated with advanced stage, increased tumor size and poor differentiation. NNMT has been shown to carry prognostic and therapeutic importance in a variety of cancers, and it can be a potential treatment target in cervical adenocarcinomas.

Preliminary study on the expression of Toll-like receptors 7 and 8 in bladder urothelial tumors

The aim of the study was to demonstrate the expression of Toll-like receptor (TLR)-8 and to confirm the expression of TLR-7 on bladder transitional cell carcinomas (TCCs) analyzing the changes in expression between low-grade (LG) and high-grade (HG) and between non-muscle invasive (NMIBC) and muscle-invasive (MIBC) bladder cancers. In our study, 25 patients who underwent transurethral resection (TURB) for bladder TCC have been selected. Thirteen of those had an LG form,while the other cases were classified as HG; the lesions were NMIBC in 18 patients and MIBC in the others seven. The analysis of TLRs expression has been performed by immuno histochemistry. TRL-8 expression seems to be higher in MIBC (85%) than in NMIBC (78%), but it is associated with a reduced percentage of immuno reactive cells and with a lower intensity. The expression had a large nuclear localization in NMIBC (80%), althoughit was mainly cytoplasmic in MIBC (72%). TLR-7 was expressed in all NMIBC samples (where the localization was mainly nucleo-cytoplasmic) and in the 71% of MIBC samples (mainly in the nucleus). A higher expression of TRL-8 in HG TCC had been observed, while TRL-7 seems to be more expressed in LG forms. Our study allowed to document the immunohistochemical expression of TLR-8 in TCCs, confirm the immunohistochemical expression of TLR-7, and suggest an increased expression of TLR-7 in LG TCC and NMIBC, and a prevalent expression of TLR-8 in HG TCC and MIBC.

CD56 Antibody: Old-Fashioned or Still Trendy in Endocrine Lung Tumors

Background: In 2010, the World Health Organization published a new classification of the endocrine tumors based on the mitotic rate and index. Concerning lung endocrine tumors, the classification of 2004 remains acceptable and widely approved. We noticed in many publications that the most used antibodies in these tumors are chromogranin and synaptophysin. This finding let us wonder about the diagnostic utility of the CD56 antibody which is widely used in our department. Material and methods: Sixty-nine endocrine lung cancers were diagnosed over a 12-month period in our Department of Pathology. Immunohistochemical technique using the three antibodies: chromogranin, synaptophysin, and CD56 was performed. The sensitivity of the three antibodies was performed using the ratio: true negative cases/true negative cases + false positive cases. The specificity wasn’t performed because the antibodies were used only in endocrine tumors. The comparison of the different percentages of expression of the three antibodies was made by the SPSS software 22.0. Results: The sensitivity of the chromogranin, synpatophysin, and CD56 accounted for 69%, 77%, and 98%, respectively. The mean percentage of immunoreactive cells with CD56 was 70% towards 15% and 20% with chromogranin and synaptophysin antibodies, respectively. The comparison of the percentages of expression showed a significant statistical difference between the expression of CD56 versus synaptophysin and CD56 versus chromogranin with P<0.001. Conclusion: CD56 antibody seems to be of diagnostic value in endocrine lung tumors with the highest sensitivity. This fact highlights the necessity of using it as a first-line neuroendocrine marker in association to chromogranin which is considered as the most specific endocrine antibody.

Immunohistochemical assessment of a unique basal pattern of p53 expression in ulcerative-colitis-associated neoplasia using computer-assisted cytometry

BackgroundThe basal pattern of p53 expression, defined as its immunoreactivity confined to the basal half of the glands, is associated with early neoplastic lesions in ulcerative colitis (UC). However, their clinical utility of this finding is limited by the use of “visual estimation” (approximate immunoreactivity on the basis of scanning the stained slide, without formal counting). This study was designed to analyze the basal pattern of p53 using computer-assisted cytometry and to identify the optimal cutoff value for discriminating between UC-associated early-stage neoplasia and regenerative atypia.MethodsThe specimens were obtained from eight UC patients undergoing colectomy and were classified according to the criteria by the Research Committee of Inflammatory Bowel Disease of the Ministry of Health and Welfare in Japan. Patients with classes UC-IIa (indefinite for dysplasia, probably regenerative), UC-IIb (indefinite for dysplasia, probably dysplastic), and UC-III (definitive dysplasia) were enrolled in the study. Based on the percentage of immunoreactive cells in the basal half of the crypt with visual estimation, basal positivity of p53 was classified into three categories: grade 1 (1 - 9%), grade 2 (10 - 19%), and grade 3 (≥20%). Next, crypts classified as grade 3 by visual estimation were analyzed by computer-assisted image analysis.ResultsUsing visual estimation, grade-3 p53 basal positivity was observed in 46.0% of UC-IIa crypts (128 of 278), 61.9% of UC-IIb crypts (39 of 63), and 94.2% of UC-III crypts (81 of 86). Using image analysis, the median p53 basal positivities were 30.3% in UC-IIa, 52.3% in UC-IIb, and 65.4% in UC-III (P ≤0.002). A receiver operating characteristics curve was generated to determine the method’s diagnostic utility in differentiating UC-IIa from UC-III. In this cohort, the sensitivity was 0.78; the specificity was 0.98; the negative predictive value was 87.4%; the positive predictive value was 95.5%, and the accuracy was 90.2% with a cutoff value for p53 basal positivity of 46.1%.ConclusionsOur findings indicate that assessing p53 basal positivity by image analysis with an optimal threshold represents an alternative to visual estimation for the accurate diagnosis of UC-associated early-stage neoplasia.Virtual SlidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/3588120501252608

Functional Screen for Secreted Proteins by Monoclonal Antibody Library and Identification of Mac-2 Binding Protein (Mac-2BP) as a Potential Therapeutic Target and Biomarker for Lung Cancer

Identification of secreted proteins of lung cancer could provide new candidates of serum biomarkers for cancer diagnosis or targets for therapeutic intervention. In this study, we developed a novel strategy that combined functional monoclonal antibody library screening technique and mass spectrometry to identify functional secreted proteins. BALB/c mice were immunized with cancer cells isolated from fresh human lung cancer tissues. The monoclonal antibody library containing 1160 mAbs was established with the mouse spleen cells, whose serum had most anti-proliferative effect on lung cancer cells. Monoclonal antibodies were subjected to an immunoreactive and functional screen and monoclonal antibodies that reacted strongly with secreted proteins in condition medium and lung cancer tissues with high inhibotion of cell proliferation were selected. Antigens that recognized by antibodies were obtained by immunoprecipitation and then identified by mass spectrometry. Mac-2-binding protein (Mac-2BP), the antigen of 13H3 antibody, was identified using this approach. Functional studies demonstrated that the 13H3 antibody suppressed lung cancer cell lines ANIP-973 and A549 proliferation in vitro and inhibit ANIP973 xenograft tumors growth in vivo by inducing cell-cycle arrest at G1 phase, with up-regulation of p27 and down-regulation of cyclin D1. Moreover, the serum level of Mac-2BP was significantly higher in lung cancer patients than healthy controls. At a cutoff value of 6 μg/ml, Mac-2BP might be a diagnostic biomarker of lung cancer, especially for SCLC. Mac-2BP concentrations of 6 μg/ml or higher was associated with poor overall survival in univariate analysis, and was an independent predictor in the multivariate COX analysis. Together, these results firstly demonstrated that Mac-2BP can be used as a therapeutic target and potential biomarker for lung cancer. Our strategy is feasible, which may facilitate the identification of novel secreted biomarkers of lung cancer.

Induction of adipose-derived stem cell into motoneuron-like cells using selegiline as preinducer

Cell therapy is one of the approaches taken to treatment of spinal cord disorders. In this study, adipose-derived stem cells (ADSCs) were induced to form motoneuron-like cells (MNLCs) using selegiline as preinducer, as well as Shh and all trans-retinoic acid (RA) as inducers. Selegiline was reported to induce the embryonic stem cells and bone marrow stromal cells into neuronal phenotype. ADSCs were evaluated using CD90, CD44, CD 49d, CD106, CD31, CD45, lipogenesis and osteogenesis. Dose response and time course studies were used in selecting the optimal concentration for selegiline using the percentage of viable cells (PVC) and percentages of immunoreactive cells (PIC) to nestin and neurofilament 68. Accordingly, such studies were used in selecting the optimal dose for RA using PVC and PIC to islet-1 and oligo-2. The expression of islet-1, oligo-2 and HLXB9 was evaluated using RT-PCR and immunocytochemistry. Real-time PCR was utilized in order to quantify the expression of islet-1, oligo-2 and HLXB9. ADSCs were immunoreactive to CD90, CD44 and CD 49d with consistent differentiation osteogenic and lipogenic cells. The optimal concentrations of selegiline and RA were 10−9mM and 2×10−8M, respectively. After two days, MNLCs showed high oligo-2 expression. MNLCs innervated myotubes; also, the release rate of synaptic vesicles using FM1-43 followed exponential decay model, and this rate in the induced MNLCs was approximately three times of that in the preinduced cells.

Nestin expression is lost in ventricular fibroblasts during postnatal development of the rat heart and re‐expressed in scar myofibroblasts

Studies have reported that the intermediate filament protein nestin was expressed in various non-stem/progenitor cells during development, downregulated during postnatal growth and re-expressed following injury. The present study tested the hypothesis that an analogous paradigm was prevalent for ventricular fibroblasts. In the neonatal rat heart, nestin protein levels were significantly higher than the adult heart and the isolation of cardiac cells revealed a selective expression in ventricular fibroblasts. In adult ventricular fibroblasts, nestin protein expression was markedly lower compared to neonatal ventricular fibroblasts. Following ischemic damage to the rat heart, nestin staining was detected in a subpopulation of scar myofibroblasts (37%) and the percentage of immunoreactive cells was greater than adult ventricular fibroblasts (7%) but significantly lower than neonatal ventricular fibroblasts (86%). Moreover, dissimilar rates of (3)H-thymidine uptake were observed among the fibroblast populations and may be related in part to the disparate percentage of nestin(+) cells. To assess the role of nestin in DNA synthesis, neonatal ventricular fibroblasts were infected with a lentivirus containing a shRNAmir directed against the intermediate filament protein. The partial depletion of nestin expression in neonatal ventricular fibroblasts significantly reduced basal DNA synthesis, in the absence of an apoptotic response. Thus, postnatal development of the rat heart was associated with a selective loss of nestin expression in ventricular fibroblasts and subsequent induction in a subpopulation of myofibroblasts following ischemic injury. The re-expression of nestin in scar myofibroblasts may represent an adaptive response to enhance their proliferative rate and accelerate the healing process.

Diagnostic Value of Automated Her2 Evaluation in Breast Cancer

Goal of this study was to asses the performance of Aperio computer-assisted analysis for HER2 immunohistochemical measurement in 292 equivocally (score2+) HercepTest immunoreactive breast cancer cases, evaluated by an experienced pathologist and analyzed with fluorescent in situ hybridization (FISH). The automatic Aperio categorization and the percentage of immunoreactive cells as evaluated by the computer and by the pathologist were recorded. Computer-assisted analysis classified 7 (2.4%) cases as negative (0), 136 (46.6%) as faintly positive (1+), 134 (40.5%) as moderately positive (2+), and 15 (5.1%) as strongly positive (3+). Correlative component analysis (CCA) classification is associated with Her2 amplification (P<0.0001). Compared with the human evaluation, automated CCA classification would save 157 (58%) FISH analyses, while not identifying 15 amplified cases (6% false-negative rate). The mean computer percentage value (CPV) is 18.44% standard deviation ±19.00 (range, 0.01 to 76.10). CPV and the pathologist percentage value are significantly associated and correlated (P<0.001) and have similar sensitivity and specificity in identifying Her2 FISH-amplified cases. CPV has a very low interobserver variation. The difference in CPV in amplified and nonamplified subgroups is statistically significant (P<0.001). Receiver operating characteristic analysis indicates that CPV is good at separating FISH nonamplified from amplified cases (P<0.001). The optimal cut-off value maximizing both sensitivity and specificity is 17.6% (sensitivity=73.3%, specificity=71.6%). Using a different cut-off value (2% of positive cells) we would have missed only 3 amplified cases (1% false-negative rate) while not submitting to FISH 52 cases (18% of the whole series). This false-negative rate is well below the expected false-negative rate usually observed in score 1 cases, supporting the use of CCA with a modified cut-off value in routine diagnostics for equivocally stained HER2 cases.

Mammalian Target of Rapamycin (mTOR) Regulates Cellular Proliferation and Tumor Growth in Urothelial Carcinoma

Mammalian target of rapamycin (mTOR) signaling has been associated with aggressive tumor growth in many cancer models, although its role in urothelial carcinoma (UCC) has not been extensively explored. Expression of phosphorylated mTOR (P-mTOR) and a downstream target, ribosomal S6 protein (P-S6), was identified in 74% (90/121) and 55% (66/121) of muscle-invasive UCCs, respectively. P-mTOR intensity and %positive cells were associated with reduced disease-specific survival (P = 0.04, P = 0.08, respectively). Moreover, P-mTOR intensity corresponded to increased pathological stage (P < 0.01), and mTOR activity was associated with cell migration in vitro. In addition, mTOR inhibition via rapamycin administration reduced cell proliferation in UCC cell lines RT4, T24, J82, and UMUC3 in a dose-dependent manner to 6% of control levels and was significant at 1 nmol/L in J82, T24, and RT4 cells (P < 0.01, P < 0.01, P = 0.03, respectively) and at 10 nmol/L in UMUC3 cells (P = 0.03). Reduced proliferation corresponded with reduced P-S6 levels by Western blot, and effects were ablated by pretreatment of cells with mTOR-specific siRNA. No effects of rapamycin on apoptosis were identified by TUNEL labeling or PARP cleavage. Administration of rapamycin to T24-xenografted mice resulted in a 55% reduction in tumor volume (P = 0.03) and a 40% reduction in proliferation (P < 0.01) compared with vehicle-injected mice. These findings indicate that mTOR pathway activation frequently occurs in UCC and that mTOR inhibition may be a potential means to reduce UCC growth.

The distribution of PAX-2 immunoreactivity in the prostate gland, seminal vesicle, and ejaculatory duct: comparison with prostatic adenocarcinoma and discussion of prostatic zonal embryogenesis

PAX-2 is a homeogene strongly expressed during development of the genitourinary tract, including the kidney and both wolffian- and müllerian-derived tissues. Expression of PAX-2 by immunohistochemistry has been studied mainly in renal epithelial neoplasms with little attention to the lower male genitourinary tract. We studied PAX-2 expression in epithelium of normal seminal vesicle, normal ejaculatory duct, normal prostatic secretory epithelium, and prostatic adenocarcinoma to define its immunoreactivity pattern throughout the prostate gland and to evaluate its potential diagnostic role in the discrimination of seminal vesicle/ejaculatory duct epithelium from prostatic adenocarcinoma. In addition, given that PAX-2 is highly expressed in tissues of wolffian duct embryologic origin, we also sought to confirm the divergent embryogenesis of the central zone, seminal vesicle, and ejaculatory duct from other regions of the prostate. Prostatectomy specimens from 12 patients were reviewed to identify blocks containing seminal vesicle, ejaculatory duct, periurethral glands, benign prostatic glands, and prostatic acinar adenocarcinoma. A total of 35 blocks from the 12 patients were evaluated. In addition, 2 tissue microarrays representing 15 additional seminal vesicles and 45 prostatic adenocarcinomas, 7 whole sections from prostatic adenocarcinomas of the central zone, and 5 core needle biopsies of seminal vesicle were also evaluated with anti-PAX-2 antibody. In the 12 radical prostatectomy whole sections, nuclear reactivity for PAX-2 was identified in 12 (100%) of 12 of the seminal vesicle epithelium, 9 (90%) of 10 of the ejaculatory duct epithelium, 0 of 12 of the prostatic adenocarcinoma, and 0 of 6 of the high-grade prostatic intraepithelial neoplasia. All 20 total additional seminal vesicles were positive for PAX-2 in the tissue microarray and biopsies; and all 52 additional prostatic adenocarcinomas were negative, including 7 of central zone origin. The staining intensity and percentage of immunoreactive cells in seminal vesicle were both 3+ in all cases. Although the ejaculatory ducts also showed diffuse staining, their staining intensity was less (2+) than that in the seminal vesicles, particularly in the ejaculatory ducts in the periurethral area (1-2+intensity). The smaller glands surrounding the main seminal vesicle duct also showed less intense staining than the luminal cells of the main duct. Of the 19 total cases with evaluable central zone glands, 2 (10.5%) had focal nuclear reactivity in normal, benign prostatic secretory cells. All other benign prostatic secretory epithelia from the peripheral and transition zones were negative for PAX-2. In conclusion, nuclear PAX-2 immunoreactivity is typical in epithelium of the seminal vesicle and ejaculatory duct; but the intensity of staining is less in the ejaculatory duct. No reactivity for PAX-2 was seen in prostatic adenocarcinoma or high-grade prostatic intraepithelial neoplasia. PAX-2 has diagnostic utility as a positive immunohistochemical marker of seminal vesicle and ejaculatory duct epithelium. In addition, these data add further support to the proposed embryogenesis of the prostatic central zone, seminal vesicle, and ejaculatory ducts from the wolffian system.

Expression of matrix metalloproteinase-9 in predicting prognosis of hepatocellular carcinoma after liver transplantation

Matrix metalloproteinases (MMPs) are known to play an important role in cell migration during cancer invasion by degrading extracellular matrix proteins. This study aimed to determine the role of MMP-9 in hepatocellular carcinoma (HCC) carcinogenesis. Eighty-nine cases who underwent liver transplantation for HCC in cirrhotic liver were selected for this study. The tumor characteristics such as nodule number, maximal diameter, portal vein invasion, and the preoperative alpha-fetoprotein levels were reviewed. The intensity of immunostaining and the percentage of immunoreactive cells with MMP-9 were evaluated. All patients were evaluated for HCC recurrence and/or death, and cause of death was noted. There was a lower survival and more recurrence risk among participants with 4 or more nodules exceeding 3 cm in diameter, with poorly differentiated tumor, and with large-vessel involvement. Eleven patients developed recurrent HCC (12.4%). Twelve patients died as a result of HCC (13.5%). Among 89 HCCs, the incidences of a weak (+) and moderate (++) expression of MMP-9 in carcinoma cells were 30.3% (23/89) and 43.8% (39/89), respectively. Increased expression and intensity of MMP-9 were found to be inversely associated with poor tumor differentiation (P = 0.016, P = 0.009, respectively). A significant correlation between expression and intensity of MMP-9 and large vascular invasion (P = 0.01, and P = 0.03) was also observed. As far as prognosis is concerned, increased immunoreactivity and intensity of MMP-9 were found to exert an unfavorable impact on overall survival rates (P < 0.01, P = 0.01, respectively) and recurrences (P = 0.001, P = 0.02). Multivariate analyses revealed that MMP-9 staining percentage (P = 0.007) and portal vein invasion (P = 0.002) were independent predictors of survival, whereas the only independent predictor of recurrences was portal vein invasion (P = 0.007). In this study, our results indicate a positive association between MMP-9 expression and histopathologic parameters that indicate poor prognosis. We conclude that together, MMP-9 staining percentage and portal vein invasion in HCC may aid to predict poor outcome. Nevertheless MMP-9 staining percentage is expected to be a potential predictive marker on survival and needs to be studied more in detail.

Precursor of prostate‐specific antigen expression in prostatic intraepithelial neoplasia and adenocarcinoma: a study of 90 cases

To assess the expression of the precursor of prostate-specific antigen (pro-PSA), a distinct molecular form of serum-free PSA that includes native and truncated forms, in benign epithelium, high-grade prostatic intraepithelial neoplasia (PIN) and prostatic adenocarcinoma. We immunohistochemically evaluated 90 formalin-fixed, paraffin-embedded prostate needle biopsies using monoclonal antibodies against [-2] pro-PSA, native [-5/-7] pro-PSA, prostate-specific membrane antigen (PSMA), PSA and racemase. Staining intensity was recorded using a scale of 0-3 (0, no staining; 3, highest staining). The percentage of immunoreactive cells in benign epithelium, high-grade PIN and adenocarcinoma was estimated in increments of 10%. All cases had [-5/-7] pro-PSA immunoreactivity. There was weak focal perinuclear cytoplasmic immunoreactivity for [-5/-7] pro-PSA in 62% (range 0-90%) of benign epithelial cells, whereas there was strong diffuse cytoplasmic staining in 83% (range 10-90%) of high-grade PIN and 87% (range 40-90%) of cancer cells. Almost all (99%) cases were immunoreactive for [-2] pro-PSA. The median (range) proportion of cells expressing [-2] pro-PSA was lower in benign epithelium, at 17 (0-80)%, than in high-grade PIN, at 55 (0-90)% (P < 0.001) and adenocarcinoma, at 55 (0-100)% (P < 0.001). The intensity of immunoreactivity for both isoforms increased from benign to neoplastic (high-grade PIN and adenocarcinoma) epithelium. A total of 31% of high-grade PIN and 11% of cancer cases with negative racemase staining showed strong staining for [-5/-7] pro-PSA. The expression of [-5/-7] pro-PSA in benign and neoplastic cells might be used in combination with high molecular weight keratin, p63, and racemase to distinguish benign epithelium from high-grade PIN and adenocarcinoma, particularly when racemase staining is negative. Both isoforms are sensitive markers for prostatic epithelium, making them possible candidates for investigating carcinoma with an unknown primary, particularly in cases in which PSA staining is negative and the level of suspicion is high.

Expression of immunoreactivity and genetic mutation in eosinophilic and ciliated metaplastic changes of endometrial glandular and stromal breakdown: cytodiagnostic implications

Various metaplastic changes may be present in endometrium, in which also cellular atypias may often be observed. Particularly, eosinophilic and ciliated changes (ECCs) occur in both nonneoplastic and neoplastic endometrium. This may cause confusion in the cytodiagnosis. This study was enterprised to investigate the possible help of immunocytochemical and cytogenetic study in the diagnostic and biologic assessment of ECC cells. In immunocytochemistry for p53 protein, Ki-67, and cyclin A, the material consists of 40 cases of cytologic smears examined by direct sampling of the endometrial cavity comprising 30 cases of ECC in endometrial glandular and stromal breakdown (EGBD) and 10 cases of well-differentiated adenocarcinoma (G1). After cytodiagnosis, immunostaining for p53 protein, Ki-67, and cyclin A was performed on multiple wet-fixed slides from each single case to evaluate the immunoreactivity, intensity of nuclear staining, and nuclear labeling index (N-LI). The intensity of nuclear staining was scored as negative (0), weak (1), moderate (2), or strong (3), and the N-LI was scored as less than 10% (0), from 10% to 25% (1), from 26% to 50% (2), or more than 50% (3), and the final score was calculated by adding both partial scores. A statistical significance test was performed by using Mann-Whitney U test, and the result was judged as significant when the P value was less than .05. For genetic mutation analysis of p53, the material comprised 6 cases of EGBD in which a high score was measured with immunocytochemistry for p53 protein, and the presence of ECC was confirmed on the hematoxylin and eosin. The ECC cells on paraffin-embedded specimens were captured using laser capture microdissection technology. Mutations in p53 gene (exons 5-8) were examined using DNA sequencing analysis. In immunocytochemistry for p53 protein, Ki-67, and cyclin A, the proportions of immunoreactive cells for p53 were statistically higher in ECC compared with those of G1 (P < .05). The proportions of the immunoreactive cells for Ki-67 and cyclin A were statistically higher in G1 compared with those of ECC (P < .05). (2) In genetic mutation analysis of p53, DNA sequencing of p53 in 6 cases revealed no mutations. The percentage of immunoreactive cells for p53 protein were higher in ECC than in G1, but the mutation point was not confirmed in genetic mutation analysis. The differential expression of these biologic parameters in ECC cells could be considered of possible relevance to the cytopathologic diagnosis in the future.

An Approach to Interpreting Immunohistochemical Stains of Adenocarcinoma in Small Needle Core Biopsy Specimens

Interpreting immunohistochemical stains of metastatic adenocarcinoma in small needle core biopsy specimens is not always straightforward. We studied the effects of small specimen size on immunohistochemical stain results in 20 colorectal adenocarcinoma hepatic resection specimens stained with cytokeratin (CK)7 and CK20. We superimposed 18- and 20-gauge needle core biopsy computer images. The results in needle core biopsy specimens correlated best with resection specimen results when immunoreactivity was assessed using the 3 images with the highest percentage of immunoreactive cells. CK7- and CK20+ needle core biopsy specimens correlated best when a higher percentage cut point was used. Immunohistochemical stains in small needle core biopsy specimens should be based on the regional area with the greatest immunoreactivity. The positive result cut point should increase as the amount of stainable carcinoma available for interpretation decreases.

P21 as a prognostic factor in non-small cell lung carcinomas

Few studies have focused on the correlation between p21 expression and survival for patients with non-small cell lung carcinoma (NSCLC), and the results are not consistent. We investigated the expression of p21 in 90 cases of NSCLC to evaluate the correlation between the p21 expression level and the clinicopathologic characteristics with patient survival. p21 was expressed in the nuclei of all the NSCLCs. The percentage of immunoreactive cells varied from 1% to 70%. All the patients were subdivided into a high and a low p21 expression group on the basis of the median percentage (17.05). There was no significant correlation between the p21 expression level and age, gender, histologic type, histologic grade, or stage. Using uni- and multivariate analyses, survival was significantly associated with gender, stage, and the p21 expression level. The survival rate for the high p21 expression group was higher than that for the low p21 expression group in the entire patient group, and especially for stage II and III patients, males, adenocarcinomas, or p53-positive tumors. Our findings showed that high p21 expression was an independent prognostic factor for NSCLC. p21 may be useful for determining the candidates for adjuvant therapies and also for selecting the appropriate chemotherapeutic agents.