Showing papers in "Applied Immunohistochemistry & Molecular Morphology in 2013"

Pathways of colorectal carcinogenesis.

Abstract: This review of the molecular and cellular changes in the different pathways of colorectal carcinogenesis sets out the classic adenoma-carcinoma sequence of the large bowel as a stepwise series of pathologic neoplastic changes associated with accumulation of genetic and epigenetic molecular alterations. The 2 major types of genomic instability found in colorectal cancers are chromosomal instability (CIN) and microsatellite instability (MSI). CIN is often associated with mutated APC. MSI is due to defective DNA mismatch repair. The associated familial cancer susceptibility syndromes are familial adenomatous polyposis coli, due to inherited APC mutations, and Lynch Syndrome or hereditary nonpolyposis colorectal cancer syndrome, due to inherited mutations in one of the mismatch repair genes (predominantly MLH1 and MSH2). In the CpG island methylator phenotype, a number of genes become transcriptionally silenced because of hypermethylation of their promoters, and this represents a key epigenetic mechanism of inactivation of tumor suppressor genes, including certain DNA repair genes. An overview of the contributions of CIN, MSI, and CpG island methylator phenotype to the different pathways of colorectal carcinogenesis allows categorization of colorectal cancers into 5 major groups on the basis of their molecular and pathologic characteristics.

SOX10 expression in malignant melanoma, carcinoma,and normal tissues

Abstract: Sry-related HMg-Box gene 10 (SOX10) is a nuclear transcription factor that plays an important role in melanocytic cell differentiation. It has been shown to be a sensitive marker of melanoma including spindle and desmoplastic subtypes. We assessed its frequency of expression in melanoma, carcinoma, benign nevi, and non-neoplastic tissues with routine immunohistochemistry for SOX10. The 109 primary melanoma included 49 epithelioid, 19 spindle cell, 22 desmoplastic, and 19 mixed spindle cell/desmoplastic melanoma. All primary, except 8 desmoplastic melanoma, and 11 metastatic melanoma were strongly and diffusely nuclear SOX10-positive. Six desmoplastic melanoma had ≤10% cells positive, and 2 were <50% positive, all of 3+ intensity. Eighteen of 149 (12%) breast carcinoma were SOX10-positive. All 24 ovarian, 23 endometrial, 26 lung, and 25 colon carcinoma were SOX10-negative. All 43 benign nevi, 18 dysplastic nevi, 68 non-neoplastic and benign skins, and all 56 non-neoplastic breast tissue were SOX10-positive. The sensitivity and specificity for SOX10 in the diagnosis of melanoma are 1.0 and 0.93, respectively; the positive and negative predictive values are 0.87 and 1.0, respectively. SOX10 is a sensitive, specific marker for melanoma. As benign nevi also express SOX10, it cannot be used to differentiate between benign and malignant pigmented skin lesions. Only a small number of breast carcinoma (12%), and breast lobules, express SOX10; no carcinoma of the ovary, endometrium, lung, or colon expressed SOX10.

Anaplastic large cell lymphoma occurring in association with breast implants: review of pathologic and immunohistochemical features in 103 cases.

Abstract: Primary lymphomas of the breast are uncommon, and mostly of B-cell type. In the late 1990s, reports began to appear,primarily in the Pathology literature, of an apparently new category of breast lymphoma of T-cell type, having a particular association with silicone breast implants. This condition came to be recognized as implant-associated anaplastic large cell lymphoma.Appearing initially as individual case reports, the pathologic features were somewhat variable and the diagnosis was difficult. This review describes the pathologic and immunohistochemical features of implant-associated anaplastic large cell lymphoma of the breast drawn from a series of 103 cases. Recommendations are given for the management of removed implants, for the approach to differential diagnosis and the choice of initial immunohistochemical panels.

Expression of squamous cell carcinoma markers and adenocarcinoma markers in primary pulmonary neuroendocrine carcinomas.

Abstract: Recent clinical trials have revealed that accurate histologic typing of non-small cell lung cancer is essential. Until now, squamous cell carcinoma (SQC) and adenocarcinoma (ADC) markers have not been thoroughly analyzed for pulmonary neuroendocrine carcinomas (NECs). We analyzed the expression of 8 markers [p63, cytokeratin (CK) 5/6, SOX2, CK7, desmocollin 3, thyroid transcription factor-1 (8G7G3/1 and SPT24), and napsin A] in 224 NECs. SOX2 (76.2%) had the greatest expression for NECs. CK5/6 (1.4%), desmocollin 3 (0.5%), and napsin A (0%) were expressed less or not at all in NECs. Although our investigated markers have been reported useful for differentiating between SQC and ADC, some of them were also present in a portion of pulmonary NECs. In our study, CK5/6 and desmocollin 3 were highly specific markers for SQC, and napsin A was highly specific for ADC. These markers are recommended for diagnosis of poorly differentiated non-small cell lung cancer.

Impact of Progesterone Receptor Semiquantitative Immunohistochemical Result on Oncotype DX Recurrence Score: A Quality Assurance Study of 1074 Cases

Abstract: Decreased or absent progesterone receptor expression in invasive breast carcinoma is a marker for an adverse prognosis. As part of an ongoing quality assurance study, this study evaluated the relationship between Oncotype DX recurrence score and progesterone receptor immunohistochemical result within each Nottingham tumor grade in 1074 cases of invasive breast carcinoma for which an Oncotype DX recurrence score was available. In addition to a statistically significant association between Nottingham grade and Oncotype DX recurrence score categories (P < 0.001), an inverse relationship was identified between progesterone receptor expression measured by modified H-score semiquantitation and Oncotype DX recurrence score that was independent of Nottingham tumor grade. The Oncotype DX recurrence score relies heavily on parameters already available from routine pathologic examination, and consideration of progesterone receptor status may aid in selection of patients most likely to benefit from ancillary testing.

Diagnostic and prognostic role of HBME-1, galectin-3, and β-catenin in poorly differentiated and anaplastic thyroid carcinomas

Abstract: Aim Thyroid cancer represents the first endocrine malignant neoplasm, accounting for 1% of human malignancy. The majority of which are well-differentiated cancer representing up to 90% of thyroid cancer and pursuing a favorable clinical course. The groups of poorly differentiated thyroid cancer (PDC) and anaplastic thyroid cancer (ATC) have a poor outcome and need a strict clinical surveillance. Materials and methods Thirty-four cases including 23 PDC/insular cancer and 9 ATC were examined for the expression of an immunohistochemical panel made up by HBME-1, galectin-3, and β-catenin and correlated either with histologic prognostic parameters or the overall surveillance. Results HBME-1 and galectin-3 were expressed in 100% of the PDC/insular cases and in none of the ATC cases. The data for β-catenin pointed out an 80% expression (12/15) in the PDCs and only a focal and nonspecific positivity in the ATCs. A β-catenin-positive expression was found in all patients with a worse outcome/death and in the presence of vascular invasion and metastatic disease. All 3 PDC patients with β-catenin negativity are alive, whereas only 41% (5/12) are alive in the β-catenin-positive group. Conclusions Our data set up the idea that PDC represents an intermediate step in the biological process of dedifferentiation of thyroid tumors toward ATC. This shift is underlined by the β-catenin expression, which seems to be related to a worse prognostic behavior. HBME-1 and galectin-3 show a similar pattern in PDC compared with well-differentiated carcinoma, whereas they are not expressed, as well as β-catenin, in anaplastic carcinomas.

Major histocompatibility complex class I and II expression in idiopathic inflammatory myopathy.

Abstract: INTRODUCTION We sought to study the intensity and pattern of major histocompatibility complex (MHC) I and II expression in muscle from patients with biopsy-proven idiopathic inflammatory myositis (IIM) including the subgroups, polymyositis (PM), dermatomyositis (DM), and inclusion body myositis (IBM). METHODS A total of 120 muscle biopsies (61 PM, 14 DM, and 45 IBM) were immunostained for MHC I and II. Staining was graded as follows. 0: no staining, 1+: ≤10% fibers, 2+: 10% to 25%, 3+: 25% to 50%, 4+: 50% to 99%, and 5+ 100%. RESULTS All IIM biopsies showed MHC I positivity; 93% showed MHC II positivity. The proportion of patients with MHC II score ≥3+ was higher in IBM than DM or PM. In DM, MHC I expression showed a perifascicular pattern. All IBM biopsies were immunopositive for MHC I and II; 30/45 were scored 5+. DISCUSSION Immunostaining for MHC I and II is a useful adjunctive test in diagnosis and subclassification of IIM.

Specificity of TLE1 expression in unclassified high-grade sarcomas for the diagnosis of synovial sarcoma.

Abstract: Expression of the transducin-like enhancer of split 1 (TLE1) by immunohistochemistry (IHC) has been widely used as a biomarker for the diagnosis of synovial sarcoma. Although TLE1 expression can be identified in more than 90% of synovial sarcomas, positive staining has been reported in up to one third of nonsynovial sarcomas, including peripheral nerve sheath tumors and neoplasms of fibrous and adipose tissues. The low specificity of this test in soft tissue tumors raises concern on its clinical application as a diagnostic biomarker. As synovial sarcoma is frequent among the differential diagnosis of unclassified high-grade sarcomas, and considering that the specificity of TLE1 antibody in this tumor group remains unclear, we evaluated TLE1 expression by IHC in 42 unclassified high-grade sarcomas. SS18 (SYT) gene break-apart analyses by fluorescence in situ hybridization were simultaneously performed as a gold standard biomarker for synovial sarcoma. Five cases that were positive for the SS18 break-apart by fluorescence in situ hybridization were also positive for TLE1 by IHC, whereas the remaining 37 cases negative for SS18 break-apart were all negative for TLE1. The results showed no evidence of nonspecific TLE1 expression in the nonsynovial high-grade sarcomas. We concluded that TLE1 is a highly specific biomarker for synovial sarcoma in the setting of differential diagnosis of unclassified high-grade sarcomas.

Significance of Notch1 Signaling Pathway in Human Pancreatic Development and Carcinogenesis

Abstract: In animal studies, Notch1-signaling pathway plays an important role in the pancreatic embryogenesis by promoting pancreatic progenitor cells self-renewal and exocrine linage development. The persistent activation of Notch pathway could arrest the organ development and keep cells at an undifferentiated stage. Studies have shown that Notch1-signaling pathway is upregulated in invasive pancreatic ductal adenocarcinoma (PDAC). Here we examined the expression pattern of Notch1 and Hes1 in human fetal pancreatic tissues to elucidate the role of Notch1 in human pancreatic embryonic development. We also compared Notch1 expression in tissues from PDAC, chronic pancreatitis and pancreatic intraepithelial neoplasm. Our data show that Notch1/Hes1-signaling pathway is activated during early pancreatic embryogenesis and reaches the highest at birth. After pancreas is fully developed, Notch1/Hes1 pathway is inactivated even though Notch1 protein cell-surface expression is upregulated. We also showed that the expression of both Notch1 and Hes1 are present in 50% (33/66) of PDACs, but not in pancreatic intraepithelial neoplasms. These findings indicate that Notch1 activation is only apparent in late stage of pancreatic carcinogenesis, suggesting that treatment with Notch-signaling inhibitors including γ-secretase should be selectively used for PDACs with confirmed Notch1-signaling activation.

Expression of SHP2 and related markers in non-small cell lung cancer: a tissue microarray study of 80 cases.

Abstract: The purpose of this study was to assess the relationships between the expression of SHP2 and VEGF, VEGFR-1, VEGFR-2, MMP-2, MMP-9, TIMP-1, TIMP-2, and microvessel density (MVD), as well as the clinicopathologic parameters of these markers in non-small cell lung cancer (NSCLC) Using a tissue microarray, the expression of these 8 markers in 80 NSCLC cases was detected by immunohistochemistry The expression of the markers was higher in cancer tissues when compared with the surrounding tissues The MVD was lower in the CD34-positive cancer tissues than in the surrounding tissues Significantly higher positive rates of expression for SHP2, MMP-9, and MMP-2 were observed in patients with lymph node metastases The later the clinical stage was, the higher the expression of MMP-9 and MMP-2 The expression of VEGF in patients with lung squamous cell carcinomas was significantly higher than in patients with lung adenocarcinomas The positive expression of SHP2 correlated significantly with that of VEGFR-2 and the MVD and a survival disadvantage was noted in the patients with SHP2-positive tumors Therefore, our data suggest that the expression of SHP2 in NSCLC has high specificity and sensitivity and is closely related to lymph node metastasis and the expression of VEGFR-2 and the MVD in patients with NSCLC SHP2 expression may promote the invasion and metastasis of NSCLC through angiogenesis and the lymphatic system

No-cost manual method for preparation of tissue microarrays having high quality comparable to semiautomated methods.

Abstract: Manual tissue microarray (TMA) construction had been introduced to avoid the high cost of automated and semiautomated techniques. The cheapest and simplest technique for constructing manual TMA was that of using mechanical pencil tips. This study was carried out to modify this method, aiming to raise its quality to reach that of expensive ones. Some modifications were introduced to Shebl's technique. Two conventional mechanical pencil tips of different diameters were used to construct the recipient blocks. A source of mild heat was used, and blocks were incubated at 38°C overnight. With our modifications, 3 high-density TMA blocks were constructed. We successfully performed immunostaining without substantial tissue loss. Our modifications increased the number of cores per block and improved the stability of the cores within the paraffin block. This new, modified technique is a good alternative for expensive machines in many laboratories.

Immunohistochemical study of the Nrf2 pathway in colorectal cancer: Nrf2 expression is closely correlated to Keap1 in the tumor and Bach1 in the normal tissue.

Abstract: Oxidative stress is a contributing factor in the carcinogenesis of colorectal cancer. The Nrf2 [nuclear factor (erythroid-derived 2)-like 2; NFE2L2] pathway is one of the major cellular defense mechanisms against oxidative stress. This study investigated the expression of the Nrf2 pathway in colorectal cancer. Formalin-fixed paraffin-embedded tissue arrays consisting of the tumor, adjacent normal, and distant normal tissues from the resected specimens of 83 colorectal cancer patients were subjected to immunohistochemical (IHC) staining with antibodies against Nrf2, kelch-like ECH-associated protein 1 (Keap1), p21, P62, Parkinson protein 7 (Park7), prohibitin, BTB and CNC homology 1 (Bach1), CD34 and 8-hydroxy-2'-deoxyguanosine (8-OHdG). The mean IHC density of each IHC staining was digitally analyzed. The results showed that molecules of the Nrf2 pathway were actively expressed, with different expression profiles among the tumor and normal tissues. The oxidative stress, represented by the mean IHC staining density of 8-OHdG, did not differ but was correlated with the expressions of different Nrf2 pathway molecules to a varied extent in tumor and normal tissues of colorectal cancer. Keap1 [estimate, 0.49; 95% confidence interval (CI), 0.19-0.79] and Bach1 (estimate, 0.24; 95% CI, 0.11-0.38) were significant predictors for the expression of 8-OHdG and had the closest proximity to Nrf2 in the cluster dendrogram of the tumor and distant normal tissues, respectively. Advanced stage (estimate, 14.9; 95% CI, 2.99-26.8) and current smoker (estimate, 15.6; 95% CI, 1.92-29.3) were significant predictors with high estimates for Bach1 in the adjacent and distant normal tissues, respectively. In colorectal cancer, the molecules of the Nrf2 pathway have different expression profiles and a difference in their importance, especially Keap1 and Bach1, related to Nrf2 and oxidative stress among tumor and normal tissues.

Salivary gland-like tumors of the breast express basal-type immunohistochemical markers.

Abstract: Background:Salivary gland–like tumors represent approximately 2% of primary breast carcinomas. These special histologic subtypes are characteristically negative for ER, PR, and HER2 (triple negative), and include adenoid cystic carcinoma, mucoepidermoid carcinoma, acinic cell carcinoma, and polymorp

Expression of TMEM166 protein in human normal and tumor tissues.

Abstract: Transmembrane protein 166 (TMEM166) is a novel human regulator involved in both autophagy and apoptosis. In this study, we generated a specific rabbit polyclonal antibody against human TMEM166 and assessed the expression of this protein in various human normal and tumor tissue samples by tissue microarray-based immunohistochemical analysis. Varying TMEM166 protein levels were expressed in a cell-type and tissue-type-specific manner in detected tissues or organs. Strong TMEM166 expression was shown in the glomerular zona of the adrenal cortex, chromophil cells of the pituitary gland, islet cells, squamous epithelium of the esophagus mucosa, the fundic gland, and hepatocytes. Moderate or weak TMEM166 staining was identified in the parathyroid gland, the testis, vaginal stratified squamous cells, lung macrophages, hematopoietic cells, renal tubular epithelial cells, macrophages in the spleen red pulp, and neuronal cells in the cerebral cortex. Some tissues failed to stain for TMEM166, such as adipose tissue, colon, cerebellum, lymph node, mammary gland, ovary, prostate, rectum, skin, small intestine, thyroid gland, tonsil, and thymus. In comparing human normal and tumor tissues, TMEM166 expression was widely downregulated in the cancer tissues. Our studies provide the basis for future investigations into cell-type-specific functions of this protein in human normal and tumor tissues.

Evaluation of SF-1 expression in testicular germ cell tumors: a tissue microarray study of 127 cases.

Abstract: Differentiating testicular germ cell tumors from sex-cord stromal tumors can be difficult in certain cases because of overlapping morphologic features and/or an absence of clinically apparent hormonal symptoms. Immunohistochemistry may be needed as an ancillary diagnostic tool in this differential diagnostic setting. Steroidogenic factor-1 (SF-1) is a nuclear transcription factor controlling steroidogenesis and is expressed in developing Sertoli and Leydig cells. Although 1 recent study has reported SF-1 nuclear immunoreactivity in testicular sex-cord stromal tumors, the specificity for this marker in germ cell tumors has not been evaluated. After encountering several problematic cases (including some on testicular biopsy), we sought to determine the diagnostic specificity of SF-1 in a large series of germ cell tumors. Nuclear immunohistochemical expression of SF-1 was evaluated in 127 germ cell tumors using tissue microarray technology with 23 non-germ cell tumor tissues as positive internal controls. No nuclear SF-1 expression was identified in any of the 127 germ cell tumors [including choriocarcinoma (3), embryonal carcinoma (25), epidermal inclusion cyst (1), intratubular germ cell neoplasia unclassified (4), seminoma (72), spermatocytic seminoma (2), teratoma (8), and yolk sac tumor (12)]. All 23 non-germ cell tumor tissues showed strong nuclear SF-1 expression in Sertoli and/or Leydig cells [including testicular atrophy (10), cryptorchidism (2), normal testis (4), hypospermatogenesis (1), immature testis (1), intratubular large cell calcifying Sertoli cell tumor (1), Leydig cell tumor (3), and Sertoli only (1)]. This study documents the absence of SF-1 expression in testicular germ cell tumors and supports its specificity for sex-cord stromal lesions in this diagnostic context.

No evidence for BRAF-V600E mutations in gastroeosophageal tumors: results from a high-throughput analysis of 534 cases using a mutation-specific antibody

Abstract: Background BRAF-V600E mutations are found in a broad spectrum of cancer types and can be successfully targeted by specific therapeutic compounds. Little data on the prevalence of BRAF-V600E mutations in tumors of the upper gastrointestinal tract are available. Materials and methods We constructed tissue microarrays of formalin-fixed and paraffin-embedded specimens of 534 gastroesophageal tumors (119 squamous cell cancers and 72 adenocarcinomas of the esophagus, 63 cancers of the gastroesophageal junction/cardia, 199 gastric cancers of the corpus or antrum, 81 gastric gastrointestinal stromal tumors) and performed anti-BRAF-V600E immunostaining using the mutation-specific antibody VE1. As control tissue we used 3 melanoma cases with confirmed BRAF-V600E mutation and distinct VE1 immunostaining. Results None of the gastroesophageal tumor cases showed a positive immunostaining signal. Conclusions BRAF-V600E mutation is not a relevant oncogenic driver in gastroesophageal tumors. Immunohistochemical analysis using mutation-specific antibodies on tissue microarrays is a feasible, time-efficient and cost-efficient approach to high-throughput screening for specific mutations in large tumor series.

Prognostic value of microvessel density and p53 expression on the locoregional metastasis and survival of the patients with head and neck squamous cell carcinoma.

Abstract: Cancer cells need to develop microvessels in order to grow and to establish metastatic foci. A role for the p53 protein in the regulation of the angiogenic process is suggested. This study aimed to investigate the relationship between immunohistochemical expression of microvessel density (MVD), meas

Clinicopathologic correlations of liver kinase B1, E-cadherin, and N-cadherin expression in non-small cell lung cancer.

Abstract: The role of liver kinase B1 (LKB1) as a tumor suppressor has emerged from the observation of increased risk of malignancy in gastrointestinal tract in Peutz-Jeghers syndrome patients harboring LKB1 gene mutations. LKB1 gene inactivation has recently been demonstrated in a subset of lung carcinoma an

HER2 in situ hybridization in gastric and gastroesophageal adenocarcinoma: comparison of automated dual ISH to FISH.

Abstract: Patients with gastric and gastroesophageal junction (GEJ) adenocarcinomas that are HER2 positive by immunohistochemistry (IHC) or in situ hybridization show a significant survival benefit with trastuzumab therapy. In situ hybridization is traditionally done by fluorescence in situ hybridization (FISH), despite some limitations. An alternative is the dual in situ hybridization (Dual ISH) technique that is fully automated and uses differentially labeled CEP17 and HER2 probes that can be read by light microscopy on 1 slide. The aim of this study was to assess the utility of Dual ISH in gastric/GEJ cancer and to compare the results with those obtained by IHC and FISH. Cases of gastric/GEJ adenocarcinoma were analyzed by IHC, FISH, and Dual ISH and the correlation between methods calculated. Results for 50 patients were available. There was a 98% (49/50) concordance rate between Dual ISH and FISH. One discrepant case was nonamplified by FISH but showed focal amplification by Dual ISH. Discrepancy was attributed to tumor heterogeneity, which was a frequent finding (78% of HER2-positive cases). There was excellent correlation between Dual ISH and FISH for assessment of HER2 amplification. Dual ISH was rapid, easy to interpret, and maintained cell morphology, which was valuable in identifying tumor heterogeneity.

Gastrointestinal stromal tumors: clinical significance of p53 expression, MDM2 amplification, and KIT mutation status.

Abstract: Background Gastrointestinal stromal tumor (GIST) is the most common mesenchymal tumor of the gastrointestinal tract. Clinical behavior is best predicted by size and mitotic count (risk index). KIT and platelet-derived growth factor receptor α (PDGFRA) mutations have therapeutic and prognostic value but few other prognostically significant molecular markers have been identified. We investigated the prognostic value of p53 protein expression and MDM2 gene amplification in a series of GISTs. Methods Thirty-five GISTs were tested for KIT and PDGFRA mutations, p53 protein expression (high >10% positive by immunohistochemistry) and MDM2 gene amplification (ratio >1.8). Mitotic index (>5/50 HPF), MDM2 amplification status, p53 protein expression, tumor size, and KIT/PDGFRA mutational status were correlated with clinical outcome. Results Only a single (3%) GIST was amplified for MDM2. p53 protein expression, mitotic index, and KIT/PDGFRA mutations did not correlate with recurrence or metastasis (P=0.20, 0.50, and 0.08, respectively) but tumor size did (P=0.04). Risk assessment (size and mitotic index) showed a weak association with clinical behavior (P=0.19). Conclusions MDM2 amplification is uncommon in GISTs. Although high p53 expression occurred in 35% of cases, it did not correlate with clinical behavior. Only GIST size predicted clinical outcome.

Smoothelin Expression in the Gastrointestinal Tract: Implication in Colonic Inertia

Abstract: Colonic inertia is a frustrating motility disorder to patients, clinicians, and pathologists. The pathogenesis is largely unknown. The aims of this study were to: (1) characterize the expression of smoothelin, a novel smooth muscle-specific contractile protein expressed only by terminally differentiated smooth muscle cells, in the normal gastrointestinal (GI) tract; and (2) determine whether smoothelin is aberrantly expressed in patients with colonic inertia. A total of 57 resections of the normal GI tract (distal esophagus to left colon) were obtained from patients without GI motor dysfunction. Sixty-one colon resections were obtained from patients with a clinical diagnosis of colonic inertia. Smoothelin immunostaining was conducted on full-thickness tissue sections. In the nondysmotile controls, strong and diffuse cytoplasmic staining for smoothelin was observed in both the inner circular and outer longitudinal layers of the muscularis propria (MP) throughout the entire GI tract. The muscularis mucosae (MM) and muscular vessel walls were either completely negative or only patchily and weakly stained. The 1 exception to this pattern was observed in the distal esophagus, in which the MM was also diffusely and strongly stained. In cases with colonic inertia, a moderate to marked reduction of smoothelin immunoreactivity was observed in 15 of 61 (24.6%) colon resections, selectively seen in the outer layer of the MP. The data demonstrate that smoothelin is differentially expressed in the MP and MM of the normal GI tract and suggest that defective smoothelin expression may play a role in the pathogenesis of colonic inertia in a subset of patients.

Detection of HPV E7 oncoviral protein in cervical lesions by a new antibody.

Abstract: The availability of a new E7 mAb-based immunohistochemistry (IHC) detection assay, Cervimax, allowed for the first time reliable testing of the E7 protein marker in formalin-fixed and paraffin-embedded tissues from cervical lesions. E7-specific IHC staining patterns were compared with those patterns of cervical cancer biomarkers, including the viral capsid protein L1 and the surrogate biomarkers p16INK4A, p53, hTERT, ubiquitin, and Ki67. The use of a tissue microarray of 138 cervical tissue cores from different pathologic stages allowed for a first profiling of the various markers in comparison with E7. Cervimax staining patterns closely overlap with those from p16INK4A and human telomerase reverse transcriptase (hTERT) in IHC staining for high-grade cervical intraepithelial neoplasia and squamous cell carcinoma. In squamous cell carcinoma, E7 immunostaining matched better to hTERT and ubiquitin profiles. On the contrary, the pattern of E7 and L1 were different in all the squamous lesions. The nuclear staining of E7 significantly discriminates between low-grade cervical intraepithelial neoplasia and high-grade cervical intraepithelial neoplasia in the basal, parabasal, and superficial layers. The results obtained in the presented pilot study suggest E7 as a valid candidate biomarker for all the stages of the malignant progression of cervical cancer; however, more extensive studies are needed to confirm the causal effect of the oncoprotein E7 in the diagnosis of human papillomavirus-induced diseases. These results also suggest that the diagnostic interpretation of cervical lesions could be increased by the combination of E7 and L1 staining in the evaluation of risk of progression, because related to different phases of viral integration.

Study of EWS/FLI-1 rearrangement in 18 cases of CK20+/CM2B4+ Merkel cell carcinoma using FISH and correlation to the differential diagnosis of Ewing sarcoma/peripheral neuroectodermal tumor.

Abstract: Merkel cell carcinoma (MCC) and primary cutaneous Ewing sarcoma/primitive neuroectodermal tumors (PCES/PNET) pose a challenging morphologic differential diagnosis. Approximately 90% of Ewing sarcoma/primitive neuroectodermal tumors (PNETs) have a specific translocation, t(11;22) (q24;q12). The EWS-friend leukemia integration-1 (FLI-1) fusion results in FLI-1 overexpression. EWS/FLI-1 rearrangement has been suggested as a useful tool in the diagnosis of PCES/PNET. In contrast, Merkel cell polyomavirus was found to be an infective agent related to the pathogenesis of MCC. Merkel cell polyomavirus can be immunohistochemically detected with the antibody CM2B4. To the best of our knowledge, there is no case of any cytokeratin (CK)20/CM2B4 PNET. The goal of our study was to investigate whether EWS/FLI-1 rearrangement was present in cases of MCC. We have studied 18 cases of MCC. To make sure that the cases investigated by fluorescent in situ hybridization were genuine MCC, we considered only CK20/CM2B4 cases. Six cases met this criterion. EWS/FLI-1 rearrangement was not evidenced in any of the 18 cases (including the 6 "genuine" cases of MCC). Although our findings were somewhat expected, we think that they fill a gap in the literature: the confirmation that MCC is devoid of the EWS/FLI-1 rearrangement. Copyright © 2012 by Lippincott Williams & Wilkins.

Identification and characterization of 2 testicular germ cell markers, Glut3 and CyclinA2.

Abstract: Testicular germ cell tumors (TGCT) are the most common type of testicular tumor and encompass different histologic types that greatly influence treatment and prognosis. Immunohistochemical studies may be required for accurate classification, particularly when these tumors present at extragonadal sites, and to aid in distinguishing histologic types. Traditional markers for identifying and distinguishing TGCT include PLAP, CD117, AFP, and CD30. More recently, the addition of OCT3/4 and SALL4 has increased sensitivity for immunohistochemical detection of germ cell tumors. We examined gene expression data from a previously published microarray study that compared normal testis mRNA expression to various TGCT. We also performed a search of the literature to identify less well-characterized markers. Glut3 and cyclinA2 showed promise as TGCT markers. Therefore, we evaluated expression of glut3 and cyclinA2 by immunohistochemistry using tissue microarrays (TMAs). Of 66 seminomas included in the TMA, 64 (97%) showed positive nuclear staining for cyclinA2 and 58 (88%) were strongly positive. Strong positive staining for cyclinA2 was also seen in the spermatocytic seminoma. All 20 of the embryonal carcinomas stained positively with cyclinA2, and 19 (95%) displayed strong nuclear staining for cyclinA2. Twenty of the 20 embryonal carcinomas stained for glut3 in a strong membranous pattern. Of 8 yolk sac tumors, 100% stained with glut3. We also evaluated glut3 and cyclinA2 staining on a general TMA containing 486 samples representing 156 different tumors. CyclinA2 stained a number of other tumor types, but the majority of these were weak or focal staining. Glut3 was rarely positive in other tumors; interestingly, most of these were of ovarian origin. We conclude that glut3 is a sensitive (96%) and specific (92%) marker for embryonal carcinomas and yolk sac tumors. Although cyclinA2 is a sensitive marker of seminomas and embryonal carcinomas (98%), its specificity is lower if focal and weak staining of nongerm cell tumors is considered positive. The sensitivity and specificity of glut3 are comparable with that seen for SALL4.

The correlations of LMX1A and osteopontin expression to the clinicopathologic stages in pancreatic adenocarcinoma.

Abstract: AIM To test the association of LMX1A and osteopontin (OPN) expression with histologic differentiation or pathologic stage in pancreatic ductal adenocarcinoma. METHODS Immunohistochemical analysis was performed to determine LMX1A and OPN expression in 100 surgical specimens obtained from Chinese patients with well-differentiated (n=15), moderately differentiated (n=65), and poorly differentiated (n=20) pancreatic ductal adenocarcinomas. RESULTS LMX1A and OPN immunoreactivities were undetectable in normal pancreatic glandular epithelia. Stronger immunostaining for LMX1A and OPN was associated with advanced nuclear grades of pancreatic ductal adenocarcinomas (70.7 and 87.1 for grade I, 109.8 and 118.3 for grade II, and 171.3 and 183.8 for grade III), and advanced TNM and American Joint Committee on Cancer stages of pancreatic ductal adenocarcinomas. CONCLUSIONS A higher expression of LMX1A and OPN is well correlated with histologic grade and pathologic stage of pancreatic ductal adenocarcinomas.

CRX/OTX3: A Useful Marker in the Differential Diagnosis of Tumors of the Pineal Region and Indicator of Photoreceptor Differentiation in Medulloblastomas and Atypical Teratoid Rhabdoid Tumors

Abstract: CRX (OTX3) is a transcription factor of the OTX homeobox family, whose expression and function is essential for the development and differentiation of retinal and pineal cells. Among human cancers, CRX seems to be selectively expressed only by retinoblastomas and pineal tumors. In our immunohistoche

Evaluation of noninvasive versus invasive techniques for the diagnosis of Helicobacter pylori infection.

Abstract: BACKGROUND Helicobacter pylori is one of the most common bacterial strains causing chronic infections, affecting over one half of the world's population. There is increasing interest in noninvasive methods for diagnosing H. pylori infection. The aim of the study was to evaluate 3 different noninvasive methods of diagnosis: the stool antigen test (HpSA), the serum antibody test, and the stool-polymerase chain reaction (PCR) test as against invasive methods based on histopathologic diagnosis. MATERIALS AND METHODS Gastric biopsies were obtained during endoscopy. Sections were stained with hematoxylin and eosin and Giemsa stain. Serum samples were tested for H. pylori antibody using an enzyme-linked immnunosorbent assay kit for the semiquantitative determination of IgG antibodies; stool samples were tested for H. pylori antigen using polyclonal enzyme-linked immnunosorbent assay kits. DNA samples from stool specimens were extracted, followed by PCR for the detection of H. pylori UreA. RESULTS The results revealed that 18/19 (94.7%) patients were positive for H. pylori infection as detected by Giemsa stain, and 84.2% were positive on the basis of hematoxylin and eosin stain, with a sensitivity and specificity of 88.9% and 100%, respectively. Diagnosis by noninvasive methods, including the serum antibody test, revealed a sensitivity and positive predictive value of 88.9% and 94.2%, respectively, whereas the stool antigen test recorded a sensitivity and positive predictive value of 72.2% and 92.9%, respectively. The stool-PCR test recorded a sensitivity of 72.2% and specificity of 100%. CONCLUSIONS Among the noninvasive methods for diagnosis of H. pylori infection, the 3 methods used in this study recorded promising results, including good sensitivity, which was the highest in the serum antibody test, whereas the stool-PCR test recorded excellent specificity.

Expression and prognostic significance of livin, caspase-3, and ki-67 in the progression of human ampullary carcinoma.

Abstract: Livin is a new member of the inhibitor of apoptosis proteins family of proteins that interacts with downstream caspases, such as caspase-3, caspase-7, and caspase-9, however, its role in human ampullary carcinoma has not been clearly defined. Immunohistochemistry was used to evaluate tissue samples from patients with ampullary carcinomas (n=71) using antibodies against livin, Ki-67 (a proliferation marker), and caspase-3. Livin was detected in 33/71 cases (in the cytoplasm of all and in the nucleus of only 2 cases). High livin expression correlated with cell differentiation, tumor-node-metastasis stage, and lymph node metastasis (P=0.001, P<0.001, and P=0.028, respectively). Caspase-3 and Ki-67 expression were significantly associated with differentiation (P<0.001, P=0.008, respectively). There was a significant negative correlation between livin and caspase-3 (r=-0.575, P<0.001), and a positive correlation between livin and Ki-67 (r=0.308, P=0.009). Survival of patients with high livin expression was shorter compared with that of patients with low livin expression (P=0.001). Expression of caspase-3 was not associated with overall survival in this cohort (P=0.335). Livin expression was an independent prognostic factor (hazard ratio 2.693, P=0.017), as was lymph node metastasis (hazard ratio 4.959; P<0.001). In this study livin expression significantly correlated with the proliferation marker Ki-67, but was negatively correlated with caspase-3 expression. These data suggest that livin may be a valuable prognostic factor for human ampullary carcinoma.

Expression of Bcl-2 and the antiapoptotic BAG family proteins in ovarian cancer.

Abstract: In epithelial ovarian cancer (EOC), literature on the prognostic value of B-cell lymphoma 2 (Bcl-2) is limited and inconsistent. Little is known about the expression patterns and the prognostic value of prosurvival proteins of the Bcl-2-associated athanogene (BAG) family proteins interacting with Bcl-2. The major aim of this study was to further define the expression pattern and the prognostic role of Bcl-2 together with BAG-1, BAG-3, and BAG-4 proteins in EOC patients receiving platinum/taxane-based chemotherapy. A tissue array was constructed comprising 63 EOC patients. The expression and the prognostic value of Bcl-2, BAG-1, BAG-3, and BAG-4 in EOC were evaluated by immunohistochemistry and multivariate analysis. A positive cytoplasmic staining for Bcl-2 was observed in 23.8% of EOC samples and in all histologic subtypes. BAG-1, BAG-3, and BAG-4 were detected in tumor cell nuclei and cytoplasm. Interestingly, all patients presenting with a positive Bcl-2 staining showed additional positive nuclear and cytoplasmic BAG-4 expression (P=0.014). Expression of Bcl-2, or the BAG family proteins, independent of nuclear or cytoplasmic localization, had no significant impact on either disease-free or overall survival, both in univariate and multivariate survival analyses with the limitation of a small cohort of cases. In this study, no association between Bcl-2 expression in EOC tumor tissue and prognosis was found. Similarly, nuclear and cytoplasmic expression of the prosurvival proteins of the BAG family had no significant impact on patients' outcome.

The influence of tissue ischemia on biomarker expression in colorectal cancer.

Abstract: The aim of the present study was to investigate the influence of fixation delay and the perioperative ischemia on hypoxia inducible factor (HIF)-1α gene expression, HIF-1α protein expression, and immunohistochemical (IHC) expression of HIF-1α, GLUT-1, Bcl-2, and Ki-67 in colorectal cancer. The study included 25 surgically removed colorectal tumors. Three sets of samples were collected readily after removal and exposed to 0, 30, and 60 minutes of delay of fixation or freezing. The perioperative ischemia time was registered. In each set of the samples, HIF-1α gene expression was analyzed by quantitative real time polymerase chain reaction, protein concentration of HIF-1α was assessed by enzyme-linked immunosorbent assay, and IHC staining of HIF-1α, GLUT-1, Bcl-2, and Ki-67 was performed. Preoperative formalin-fixed paraffin-embedded biopsies and whole sections of the entire tumor were analyzed by IHC. We found that the HIF-1α gene expression, HIF-1α protein concentration, and IHC expression of HIF-1α, GLUT-1, Ki-67, and Bcl-2 were not systematically affected by either the fixation or freezing delay of the tissue, the perioperative ischemia time, or the total ischemia time (perioperative ischemia+delay of fixation or freezing) in colorectal tumors. However, the intraindividual variation was quite large, which may question the use of individually, non-standardized-handled single biopsies or small tissue samples for analysis of often rather heterogenously expressed biomarkers.