Immunoexpression of SDHB, FH, and CK20 among eosinophilic renal tumors: A tissue microarray study.

BACKGROUND: Differential diagnosis can be a challenge for eosinophilic subtypes of renal cell tumors due to their overlapping histomorphological and immunohistochemical features. We aimed to investigate the frequency of rare variants of renal cell carcinomas (RCCs) such as succinate dehydrogenase-deficient RCC (SDDRCC), hereditary leiomyomatosis and RCC (HLRCC)-associated RCC, and eosinophilic, solid, and cystic RCC (ESCRCC) in our population. MATERIALS AND METHODS: Renal tumors which could be considered in the eosinophilic tumor category were included: 91 conventional clear cell RCCs with eosinophilic cytoplasm, 72 papillary RCCs, 74 chromophobe RCCs, 88 oncocytomas, and 37 other rare subtypes. Using the tissue microarray method, succinate dehydrogenase B (SDHB), fumarate hydratase (FH), and cytokeratin 20 (CK20) antibodies were performed by immunohistochemistry. Immunohistochemistry was repeated on whole block sections for selected cases. The utility of these antibodies in the differential diagnosis was also investigated. RESULTS: Loss of SDHB expression was detected in three tumors, two of which showed typical morphology for SDDRCC. In additional two tumors, SDHB showed weak cytoplasmic expression without a mitochondrial pattern (possible-SDHB deficient). None of the tumors showed loss of FH expression. Heterogeneous reactions were observed with SDHB and FH antibodies. Only one ESCRCC was detected with diffuse CK20 positivity. CONCLUSION: SDDRCCs, HLRCC-associated RCCs, and ESCRCCs are very rare tumors depending on the population. Possible weak staining and focal loss of SDHB and FH expression should be kept in mind and genetic testing must be included for equivocal results.

Identification of an Autoantibody Against ErbB-3-Binding Protein-1 in the Sera of Patients With Chronic Hepatitis B Virus Infection.

Background: Studies have shown that autoimmune response contributes to chronic hepatitis B (CHB) development. Aim: This study aimed to identify autoantibodies in the sera of patients with CHB and to investigate the association of autoimmune response with disease severity in CHB. Methods: Proteins from human liver carcinoma cell line HepG2 were separated by two-dimensional electrophoresis. The candidate autoantigens were recognized by serum autoantibodies from Chinese CHB patients. Immunohistochemical staining was performed to determine the hepatic expression of the autoantigen in CHB patients with different inflammatory grades. Enzyme-linked immunosorbent assay (ELISA) was conducted to measure the prevalence and the levels of serum autoantibody in CHB patients with different disease severity. Flow cytometry analysis was carried out to assess the autoreactive T cell response in the peripheral circulation of CHB patients. Results: ErbB-3-binding protein-1 (EBP-1) was identified as an autoantigen of serum autoantibodies in CBP patients. EBP-1 protein expression was upregulated in the liver of CHB patients with high-grade hepatic inflammation. The prevalence and levels of serum anti-EBP1 IgG were significantly increased in CHB patients with severe diseases compared with those with mild or moderate diseases, but none was detectable in the healthy controls. EBP-1 peptides induced proinflammatory cytokine expression in CD4+ T cells from CHB patients. Conclusion: Our results demonstrate the presence of an autoantibody against EBP-1 in the sera as well as EBP-1-reactive T cells in the peripheral blood of CHB patient. EBP-1-induced autoimmune response is positively associated with the disease severity, suggesting that EBP-1-induced autoimmune response possibly contributes to progressive liver failure.

Urothelial Carcinoma In Situ of the Bladder: Correlation of CK20 Expression With Adaptive Immune Resistance, Response to BCG Therapy, and Clinical Outcome.

Immunohistochemical stains have been suggested to aid in diagnostically challenging cases of urothelial carcinoma in-situ (CIS). Although full thickness immunostaining for CK20 is supportive of CIS, a subset of CIS cases is CK20(-), the clinical significance of which was unknown. This study included 43 patients with primary diagnosis of bladder CIS including 32 with only CIS, 5 with CIS and separate noninvasive high-grade papillary urothelial carcinoma, and 6 with CIS and separate high-grade urothelial carcinoma with lamina propria invasion. Digital morphometric image analysis showed that the average nuclear areas of enlarged nuclei were similar in CK20(+) and CK20(-) CIS (26.9 vs. 24.5 µM2; P=0.31). Average Ki67 index for CK20(+) CIS was higher than CK20(-) CIS (31.1% vs. 18.3%; P=0.03). Patients with CK20(+) CIS [28 (65%)] and patients with CK20(-) CIS [15 (35%)] had the same rates of Bacillus Calmete-Guerin (BCG) failure but patients with CK20(-) CIS had higher stage progression [3 CK20(+) (11%) vs. 6 CK20(-) (40%); P=0.02]. Given recent approval of immune checkpoint inhibitors in patients with CIS refractory to BCG, programmed death ligand-1 expression and colocalization with CD8(+) lymphocytes was investigated as signature of adaptive immune response and was seen in 8 patients regardless of CK20 status and exclusively among patients who failed BCG. Our results confirm that negative CK20 IHC does not exclude CIS and that those patients have similar clinical outcomes as patients with CK20(+) CIS. Programmed death ligand-1 and CD8 colocalization seen among patients who failed BCG therapy is an easy assay to perform to identify patients who could potentially benefit from combined BCG therapy and immune checkpoint inhibition.

Stratification of urinary bladder carcinoma based on immunohistochemical expression of CK5, CK14 and CK20.

Molecular subtyping of urothelial carcinoma (UC) is similar to that of breast cancer and is based on the developmental biology approach. The aim of the present study is to assess the prognostic impact of CK5, CK14, and CK20 expression in urinary bladder cancer (UBC) with the potential to stratify them into different subtypes. The current study examined the immunohistochemical expression of CK5, CK14, and CK20 in 90 specimens of UBC. CK5 was expressed in 81.1% of the cases and was significantly associated with old age, muscle invasion, presence of bilharziasis, and tendency for poor overall survival. CK20 was expressed in 47.8% of the cases and was associated with nonmuscle invasion and pure UC while 50% of the cases expressed CK14 that were associated with muscle invasion and perineural invasion. Most squamous cell carcinoma and those associated with bilharziasis were belonged to Ck5+/CK20- subgroup while pure UC and those lacked bilharziasis were located in the Ck5+/CK20+ subgroup. The basal group (Ck5+/CK14+/CK20-) showed high proliferative features compared to the intermediate group (Ck5+/CK14-/CK20-). Generally, presence of CK5 is associated with adverse features especially in the group lacking CK20; however, basal and intermediate subgroups share CK5 expression but they show different proliferative capacities, so their distinction by CK14 is helpful.

Diagnosis of Lymph Node Metastasis in Colorectal Cancer by a Semi-dry Dot-blot Method.

BACKGROUND/AIM: This study aimed to examine whether the semi-dry dot-blot (SDB) method can correctly identify metastasis to lymph nodes in colorectal cancer. MATERIALS AND METHODS: A total of 200 dissected lymph nodes from 83 patients with colorectal cancer who underwent surgery between November 2013 and May 2016 were examined. Each lymph node was first examined by SDB using anti-pancytokeratin antibody (AE1/AE3). Pathological Stage I/II patients with a negative reaction were further analyzed by SDB using anti-cytokeratin 20 antibody (CK-20) to detect micrometastasis or isolated tumor cells. RESULTS: The sensitivity, specificity, and accuracy of SDB using AE1/AE3 were 91.3%, 100%, and 98.0%, respectively. Five of 99 lymph nodes of pathological Stage I/II patients had a negative reaction to AE1/AE3, but were positive to CK-20, while 3 showed isolated tumor cells. CONCLUSION: The SDB is a useful diagnostic tool to detect lymph node metastases in colorectal cancer.

Evidence of proliferative activity in human Merkel cells: implications in the histogenesis of Merkel cell carcinoma.

The cellular origin of Merkel cell carcinoma (MCC) is controversial. We previously hypothesized that MCC originates from hair follicle stem cells or Merkel cell (MC) progenitors residing within the hair follicle bulge. Examination of three cases of combined MCC led to the unexpected discovery that numerous keratin 20 (CK20)-positive MCs within the squamous cell carcinoma (SCC) component of combined MCC appeared morphologically normal with dendritic and oval shapes. Moreover, one extremely rare case of combined SCC and MCC showed both intra-epidermal and dermal MCCs. These three cases represent the first documentation of MC hyperplasia in MCC, besides various benign follicular neoplasms associated with MC hyperplasia. Therefore, to elucidate the proliferating potential of MCs and their histogenetic relationship with MCCs, we further investigated these cases based on pathological observations. We identified numerous cells co-expressing CK20 and the proliferation marker Ki-67, identical to the morphological and immunohistochemical features of normal MCs. This finding indicated that MCs can no longer be considered as pure post-mitotic cells. Instead, they have proliferative potential under specific conditions in the diseased or wounded skin, or adjacent to various skin tumors, including MCC. Intimate co-existence of two malignant cell components composed of intradermal and intra-epidermal MCCs, with the proliferation of normal-appearing MCs in the same lesion, lends support to the hypothesis that MCs and MCC cells are derived from MC progenitors residing within the hair follicle bulge. Specifically, MCCs are derived from transformed MC progenitors with potential for dual-directional differentiation towards neuroendocrine and epithelial lineages.

Alcohol Feeding in Mice Promotes Colonic Hyperpermeability and Changes in Colonic Organoid Stem Cell Fate.

BACKGROUND: Alcohol increases intestinal permeability to proinflammatory microbial products that promote liver disease, even after a period of sobriety. We sought to test the hypothesis that alcohol affects intestinal stem cells using an in vivo model and ex vivo organoids generated from jejunum and colon from mice fed chronic alcohol. METHODS: Mice were fed a control or an alcohol diet. Intestinal permeability, liver steatosis-inflammation, and stool short-chain fatty acids (SCFAs) were measured. Jejunum and colonic organoids and tissue were stained for stem cell, cell lineage, and apical junction markers with assessment of mRNA by PCR and RNA-seq. ChIP-PCR analysis was carried out for Notch1 using an antibody specific for acetylated histone 3. RESULTS: Alcohol-fed mice exhibited colonic (but not small intestinal) hyperpermeability, steatohepatitis, and decreased butyrate/total SCFA ratio in stool. Stem cell, cell lineage, and apical junction marker staining in tissue or organoids from jejunum tissue were not impacted by alcohol. Only chromogranin A (Chga) was increased in jejunum organoids by qPCR. However, colonic tissue and organoid staining exhibited an alcohol-induced significant decrease in cytokeratin 20+  (Krt20+) absorptive lineage enterocytes, a decrease in occludin and E-cadherin apical junction proteins, an increase in Chga, and an increase in the Lgr5 stem cell marker. qPCR revealed an alcohol-induced decrease in colonic organoid and tissue Notch1, Hes1, and Krt20 and increased Chga, supporting an alteration in stem cell fate due to decreased Notch1 expression. Colonic tissue ChIP-PCR revealed alcohol feeding suppressed Notch1 mRNA expression (via deacetylation of histone H3) and decreased Notch1 tissue staining. CONCLUSIONS: Data support a model for alcohol-induced colonic hyperpermeability via epigenetic effects on Notch1, and thus Hes1, suppression through a mechanism involving histone H3 deacetylation at the Notch1 locus. This decreased enterocyte and increased enteroendocrine cell colonic stem cell fate and decreased apical junctional proteins leading to hyperpermeability.

CD99 expression in Merkel cell carcinoma: a case series with an unusual paranuclear dot-like staining pattern.

BACKGROUND: Merkel cell carcinoma (MCC) is a rare neuroendocrine cancer of the skin. The utility of CD99 (MIC-2) in the diagnosis of MCC has been previously studied, with reported rates of expression ranging from 13 to 55%. When specified, a membranous or cytoplasmic staining pattern was considered significant. Recent studies of CD99 have identified a paranuclear dot-like expression pattern in certain non-neuroendocrine pancreatic and colonic lesions. We recently noted paranuclear dot-like staining in several cases of MCC, including cases lacking cytokeratin 20 (CK20) expression. METHODS: Fourteen cases of MCC were stained with CK20 and CD99 antibody, and the pattern and intensity of staining were recorded. Seven cases of pulmonary small cell carcinoma (PSCC) and one case of primitive neuroectodermal tumor (PNET) were used for comparison. RESULTS: All 14 cases of MCC showed at least focal CD99 staining, with both membranous and paranuclear dot-like staining patterns identified. CK20 staining was present in 12/14 cases, with the characteristic dot-like pattern identified. Four of seven cases of PSCC showed CD99 staining, with two showing a finely granular dot-like staining pattern. CONCLUSIONS: We report an unusual pattern of paranuclear dot-like expression of CD99 in 14 cases of MCC, two of which did not express CK20. This previously unrecognized expression pattern may be of use in differentiating MCC from other cutaneous malignancies, especially when CK20 expression is limited or absent.

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.

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.

Expression and potential clinical significance of urothelial cytodifferentiation markers in the exstrophic bladder.

PURPOSE: We characterize the urothelium from patients with classic bladder exstrophy-epispadias complex for the expression of proteins associated with urothelial differentiation, and discuss a potential impact of urothelial phenotype on the structural and functional properties of the bladder template following bladder closure. MATERIALS AND METHODS: From 2005 to 2010 bladder biopsies from 32 infants with bladder exstrophy-epispadias complex obtained at primary bladder closure were collected. After histological assessment immunochemistry was used to investigate the expression of uroplakin IIIa, cytokeratin differentiation restricted antigens CK13 and CK20, and tight junction protein claudin 4. RESULTS: Overall tissue morphology showed gross alterations with inflammatory, proliferative and metaplastic changes in most specimens. Sections of intact epithelium were present in 78% of biopsies. With respect to urothelial phenotype, CK13 was expressed in all specimens, whereas UPIIIa and CK20 were absent in 76% of the tissues examined. Of the biopsies 52% revealed an irregular expression pattern of tight junction protein Cl-4. CONCLUSIONS: This is the first study to our knowledge to characterize the urothelium from infants with bladder exstrophy-epispadias complex for the expression of urothelial differentiation associated antigens. Our findings suggest urothelial differentiation changes in a majority of exstrophic bladders, at least at primary bladder closure. Although the underlying etiology remains to be established, abnormal urothelial differentiation may result in a dysfunctional urothelial barrier with implications for the structural and functional properties of the bladder template. Despite the study limitations, our preliminary findings provide a platform for further investigation of the significance of the urothelium for the exstrophic bladder.

Clinicopathological features and immunoprofile of 30 cases of Brenner ovarian tumors.

AIM: The aim of this study was to investigate the clinicopathological and immunopathological features of Brenner ovarian tumors. MATERIALS AND METHODS: Thirty cases of Brenner ovarian tumors were examined in our laboratory among 1,680 cases of ovarian tumors, representing 1.5% of all tumors examined. Blocks of paraffin-embedded tumor tissue for all cases were available for additional immunohistochemical stain by a Ventana autoimmunostainer. Moreover, antibodies for Uroplakine III (cellmarque AU-1 clone, 1:25) Chromogen (monosan clone 5H7,1:25) WT1 (novocastra, clone 3F-H2, 1:25) NSE (DAKO, clone BB5/NC/V1-H14, 1:50), CK20 (DAKO, clone Ks20.8, 1:50),CK7 (Zymed 1:25, clone V-TL12/30)were used. RESULTS: The mean age of the patients was 51.4 years ranging from 16 to 82 years. The tumor was unilateral in 28 cases (16/28 in the right ovary and 12/28 in the left ovary) and bilateral in two cases. Twenty-eight cases (93%) were benign and two (7%) were proliferating (borderline) tumors. Seventeen cases (56%) were pure Brenner tumors, measuring from 0.5 to 2.5 cm and 13 cases (44%) were mixed tumors consisting of a Brenner tumor element and a mucinous ovarian tumor (10/13 cases, 53.8%) and a germ cell tumor in 3/13 cases. The largest diameter of the mixed tumors ranged from 7 to 22 cm. The largest area consisting of Brenner elements measured 7 cm. The immunoprofile of Brenner tumor cell was cytokeratine-7 positive (30/30 cases) cytokeratine-20 negative in the Brenner cell element but positive in the mucinous component in 5/7 cases of mixed Brenner tumors, focally WT-1 positive (5/30 cases), NSE negative (0/30 cases) and focally chromogranine positive (6/30 cases), Uroplakin-III positive in 23/30 cases, with faint cytoplasmatic or luminal distribution. In conclusion, Brenner ovarian tumors are unilateral, small and benign neoplasms in their majority and present specific histopathological and immunopathological characteristics and mixed forms with other epithelial and germ cell neoplasms. This could be explained as a form of metaplasia or a diverse histogenesis from surface epithelium and/or the germ cell ovarian component.

The diagnostic value of cytokeratins expression in the renal parenchyma tumors.

Renal carcinomas are a heterogeneous group of tumors, difficult to classify and identify precisely. Since their prognosis depends very much upon their type, precise diagnosis might mean the difference between therapeutic success and patient death. Cytokeratins are particularly useful for the identification of the epithelial nature of the tumors, because their expression is maintained even in poorly differentiated tumors. Monoclonal cytokeratins such as CK7 and CK20 stain different components of the renal tubular system and are a useful duo for the identification of the origin of the different tumors that might arise in the kidney. Along with polyclonal cytokeratins such as AE1/AE3 and high molecular weight cytokeratin antibodies (34betaE12, Cam 5.2), epithelial membrane antigen (EMA) and vimentin, they are included in every diagnostic panel for renal tumors. We have selected 138 renal parenchyma tumor specimens, performed morphological diagnosis and then stained them with polyclonal cytokeratin antibody AE1/AE3, and monoclonal antibodies to CK7 and CK20. AE1/AE3 was expressed in 61.7% of the renal parenchyma tumors, with high intensity and percentage of positive cases in the papillary carcinomas (100%), and with rare and weakly positive cells in chromophobic cells carcinomas, clear cells carcinomas and sarcomatous carcinomas. CK7 was positive in 68% of the renal parenchyma tumors, with positive reaction in 100% of the cases of chromophobic cells and sarcomatous carcinomas. Clear cells carcinomas had the less percentage of positive cells, whereas papillary carcinomas were positive in seven out of eight cases. No difference in the staining pattern was noticed between type I and type II papillary carcinomas. CK20 was negative in all cases studied.

[Immunohistochemical profile of colorectal and ovarian carcinomas--examination with cytokeratin 7, cytokeratin 20, beta catenin and cDX 2].

INTRODUCTION: Metastases from colorectal adenocarcinomas can be histologically similar to serous, mucinous and endometrioid ovarian adenocarcinomas. The differentiation between primary and metastatic ovarian tumours is of great importance for the patients because of the different treatment and prognosis. AIM: The aim of the study was to determine whether the differences in the expression of Cytokeratin 7, Cytokeratin 20, Beta catenin and CDX2 can be used to distinguish the different types of carcinomas and their metastases. MATERIALS AND METHODS: The immunohistochemical expression of the listed above antibodies was examined retrospectively and prospectively in 38 colorectal adenocarcinomas (primary and metastatic) and 32 ovarian adenocarcinomas (primary and metastatic). The metastases in both types of adenocarcinomas are located in the peritoneum. RESULTS: The immunohistochemical expression was evaluated using a semi-quantitative method. The ovarian adenocarcinomas are mostly positive for Cytokeratin 7 (in 63%), while colorectal carcinomas are mostly positive for Cytokeratin 20 (in 73%). Regarding Beta catenin, in colorectal carcinomas the expression is mostly nuclear (in 65%) and in ovarian carcinomas mostly membrane (in 68%). In cases of uncertain expression of the markers mentioned above, CDX2 was used. Positive nuclear expression was observed only in intestinal tumours (in 86%). CONCLUSION: For differential diagnosis between ovarian and colorectal adenocarcinomas, the use of antibodies, determining the intestinal differentiation of the tumours like Cytokeratin 20, Beta catenin and CDX2 is recommended.

Detection of metastatic colon cancer cells in sentinel nodes by flow cytometry.

In colon cancer the presence of metastases in the regional lymph nodes is an important prognostic factor. Currently, examination is based on histological and immunohistochemical evaluations of lymph node sections. However, these methods are time-consuming, labour intensive and micro-metastases might be missed. The sentinel node technique may be used to identify the direct tumour draining lymph node. In this study the possible use of flow cytometry for detection of sentinel node metastases in patients with colon cancer has been investigated. Peripheral blood mononuclear cells (PBMC) with the addition of DLD-1 colon cancer cells were stained intracellularly for cytokeratin 20 (CK20), and for the cell surface markers epithelial cell adhesion molecule (EpCAM) and carbohydrate antigen 19-9 (CA19-9). CK20 positive colon cancer cells were reliably detected with as few as 0.037% events. However, PBMCs from both colon cancer patients and from healthy individuals contained CK20 positive cells. Staining for EpCAM resulted in an almost complete recovery of tumour cells, and as few as 0.037% added cells were detected. Low intra-assay variability was determined for EpCAM (CV 8.8) and CA19-9 (CV 9.7) stainings. The results from staining for CK20 or for EpCAM and CA19-9 concorded, but the degree to which respective antigens were expressed varied. The use of flow cytometry for detection of metastatic colon cancer cells was verified in fourteen sentinel nodes specimens from patients with colon cancer. Herein we have explored the potential of flow cytometry to become a fast, sensitive and reliable method for detection of lymphatic metastases in patients with colon cancer using direct fluorophore-conjugated antibodies against multiple surface antigens. The method seems feasible and further testing is warranted.

Immunoreactivity of p63 in monolayered and in vitro stratified human urothelial cell cultures compared with native urothelial tissue.

OBJECTIVE: To investigate the immunoreactivity of p63 in monolayered and stratified human urothelial cell cultures and in normal urothelial tissues to assess the differentiation status of in vitro stratified urothelial constructs. METHODS: p63 expression was detected immunohistochemically in native normal human bladder, ureter, and renal pelvis tissues and immunocytochemically in monolayered urothelial cell cultures and urothelial constructs stratified in vitro. Additionally, expression of pancytokeratin, cytokeratin 20 (CK20), uroplakin III, and fibroblast surface antigen was investigated. RESULTS: In native tissues, urothelial cell layers showed the most intensive p63 staining in the basal cells; the superficial umbrella cells were predominantly negative. Monolayered urothelial cell cultures revealed reduced p63 expression with ongoing culture passages. In vitro stratified urothelial constructs exhibited p63 expression similar to that of native urothelium. CK20-reactive cells were absent in the monolayered cultures but present in the stratified cell cultures and in the urothelial constructs. In native urothelium, only superficial cells stained positive for CK20. Uroplakin III was not present in either monolayered urothelial cell cultures or stratified urothelial constructs. Cultured cells were always positive for pancytokeratin and negative for fibroblast surface antigen. CONCLUSIONS: p63 is a new biomarker for differentiation and stratification of urothelium created in vitro. For proposed clinical applications of in vitro stratified urothelium in reconstructive urology, urothelial constructs should exhibit expression of significant marker proteins similar to that of native urothelium. Our results show such similarity of expression for pancytokeratin, p63, and CK20, an encouraging possibility for confirming the functionality of tissue-engineered urothelia after clinical application.

Usefulness of cytokeratin immunoreactivity pattern for distinction of Barrett's esophagus from intestinal metaplasia of the stomach.

The histological distinction between Barrett's esophagus involving the distal esophagus and intestinal metaplasia of the stomach has important clinical implications and can be difficult even with the use of histochemical stains. Cytokeratin (CK) 7 and 20 are cytoplastic structural proteins that show restricted expression in normal and malignant epithelia of the gastrointestinal tract. CK7 and 20 immunostaining were performed on a 67-year-old male with cardiac cancer with reflux esophagitis due to sliding hernia. The CK7/20 immunoreactivity pattern of cancer and reflux esophagitis in this case showed superficial CK20 staining and strong CK7 staining of both superficial and deep glands. In intestinal metaplasia of the stomach, strong CK20 immunostaining in superficial and deep glands and absent CK7 immunoreactivity were noted. Neither CK7 nor CK20 immunoreactivity was noted in squamous cell epithelium. Therefore, we concluded that in this patient intestinal metaplasia of the esophagus was BE. The CK7/20 reactivity pattern is useful for identifying the intestinal metaplasia of the esophagus from the stomach using histological materials from biopsy and surgically resected specimens.

Immunocytological detection of lymph node metastases in dogs with malignant epithelial tumours.

The aim of this study was to compare immunolabelling of cytological specimens with conventional staining in the detection of metastases in lymph nodes from dogs with carcinoma. Cytological touch imprints of 161 lymph nodes from 72 dogs, as well as 50 fine needle aspirates from 23 dogs, with malignant epithelial tumours were included in the study. Immunolabelling was performed with commercially available human antibodies. Touch imprints of all lymph nodes were labelled with broad spectrum anticytokeratins AE1/AE3 and KL1. In addition, lymph node touch imprints from dogs with primary tumours that reacted positively with the specific anticytokeratins CK7 (n=104) and CK20 (n=20) were also labelled with CK7 and CK20. Fine needle aspirates of 50 lymph nodes were examined by immunolabelling with AE1/AE3. "Reference investigations" with a combination of histological and immunohistochemical methods were performed on all lymph nodes. The immunocytological detection of lymph node metastases with the broad spectrum anti-cytokeratin AE1/AE3 in imprint smears resulted in a significant increase in sensitivity (0.99 vs 0.88 [conventional stain]) and in negative predictive value (0.99 vs 0.85) (P<0.01; t-test). Micrometastases in particular were detected more readily. Specificity (0.93 vs 0.88) and positive predictive value (0.95 vs 0.90) did not differ significantly between the two techniques. Immunolabelling with KL1 was associated with lower sensitivity and negative predictive value, indicating lack of cross-reactivity of this antibody with canine epithelial cells. In fine needle aspirates the detection of lymph node metastases, especially micrometastases, was more efficient by mean of immunolabelling with AE1/AE3 than by conventional staining. The study indicated the value of immunocytological labelling for the detection of metastases in cytological specimens of canine lymph node preparations.