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Weak Staining (weak + staining)
Selected AbstractsMyocardial lipofuscin-laden lysosomes contain the apoptosis marker caspase-cleaved cytokeratin-18EUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 10 2008A. Soleiman ABSTRACT Background Acute coronary syndrome is related to increased circulatory concentration of soluble apoptosis specific caspase-cleaved cytokeratin-18 (ccCK-18). Potential cardiac sources of this intermediate filament derivative have not been investigated to date. Materials and methods Paraffin embedded tissue of normal myocardium, and chronically damaged samples of ischaemic, congestive and hypertrophic cardiomyopathy were analysed by histology and by CK-8, CK-18, ccCK-18 immunohistochemistry (each group, n = 15). Antibody specificity of the ccCK-18 antibody M30 was checked by immunoblotting on lysed myocardium and enriched myocardial lysosomes. Results ccCK-18 and CK-18 but not CK-8 were present in all forms of cardiomyopathy, most prominently in ischaemic cardiomyopathy while only traces were detectable immunohistochemically in normal myocardium. Weak CK-18 and strong ccCK-18 staining co-localized to lysosomes with cardiac age pigment lipofuscin. Weak staining of CK-18 was detected in the cytoplasm of coronary endothelia. Conclusion Our study reveals that cardiac lipofuscin-laden lysosomes contain ccCK-18, a marker of apoptosis and its precursor CK-18. This ccCK-18 pool might contribute to increased systemic levels of ccCK-18 in acute coronary syndrome thus monitoring myocardial damage. [source] mRNA expression of urokinase and plasminogen activator inhibitor-1 in human crescentic glomerulonephritisHISTOPATHOLOGY, Issue 2 2001H S Lee mRNA expression of urokinase and plasminogen activator inhibitor-1 in human crescentic glomerulonephritis Aims:,Weak staining for urokinase-plasminogen activator (uPA), tissue type plasminogen activator (tPA), or plasminogen activator inhibitor-1 (PAI-1) confined to crescents has been described in a few cases of severe crescentic glomerulonephritis. We evaluated the molecular mechanism by which these proteins are increased or induced within crescents. Methods and results:,We examined uPA, tPA and PAI-1 mRNA expression in 12 renal biopsies with crescentic glomerulonephritis, and in six control renal biopsies with no detectable abnormalities by RNA in-situ hybridization. The expressions of uPA, tPA and PAI-1 proteins were also assessed by immunofluorescence. To better determine the cellular origin of uPA and PAI-1 transcripts, CD68 protein was studied by immunohistochemistry on the same sections on which in-situ hybridization had been performed. In controls, there were very low level signals of uPA and PAI-1 mRNAs in a few glomerular epithelial cells (GECs). Specific signals of uPA and PAI-1 mRNAs were detected in the cells forming crescents in all the cases with crescentic glomerulonephritis. However, weak expression of mRNA for tPA was detected in two cases only. Immunostaining for uPA and PAI-1 was positive in some but not all, cases of crescentic glomerulonephritis. A double-labelling study showed that the signal for PAI-1 and uPA mRNAs was mainly in CD68, cells. Conclusions:,Local accumulation of uPA or PAI-1 in crescents is associated with enhanced mRNA expression of these proteins. The up-regulation of PAI-1 mRNA by GECs, in particular, could play a major role in the formation of persistent fibrin deposits and progression of the lesions in crescents. Whether up-regulation of uPA is an epiphenomenon or plays a pathogenic role in the formation of crescents remains to be clarified. [source] Trefoil factor family 3 expression in the oral cavityEUROPEAN JOURNAL OF ORAL SCIENCES, Issue 6 2009T. Storesund This study examined the expression, in oral keratinocytes and in the major and minor salivary glands, of Trefoil factor family 3 (TFF3) peptide. Trefoil factor family 3 messenger RNA (mRNA) and peptide were detected in cultures of normal oral keratinocytes by quantitative real-time polymerase chain reaction (PCR) and western blotting, respectively. Trefoil factor family 3 was found, by immunohistochemical analyses, to be expressed in the basal layers of the oral mucosal epithelium. In salivary glands, immunohistochemical staining showed that TFF3 peptide expression was strongest in the mucous acini of the submandibular and the small salivary glands. Serous cells in the same glands showed weak staining. In the parotid gland, many serous acini showed weak positive staining, while other areas did not. In all glands examined, the intercalated, striated, and collecting ducts were moderately TFF3-positive. Double immunostaining confirmed that mucous (MUC5B positive) cells were moderately or strongly positive for TFF3 and that some serous (MUC7 positive) cells showed restricted TFF3 expression, mostly in a granular pattern. The prevalence of the TFF3 peptide in the salivary secretions of healthy volunteers was detected by western blotting of saliva from minor salivary glands (four of five) and the parotid gland (one of five) and of mixed submandibular/sublingual saliva (five of five). In conclusion, the submandibular and small salivary glands appear to be the major producers of oral TFF3, but duct cells of all glands and keratinocytes of the oral mucosa may also contribute as sources of TFF3 in the oral cavity. [source] Ultrastructural and histochemical study on gills and skin of the Senegal sole, Solea senegalensisJOURNAL OF APPLIED ICHTHYOLOGY, Issue 6 2004J. M. Arellano Summary This study was undertaken to identify the normal ultrastructural features of gills and skin of the Senegal sole, Solea senegalensis, for a comparative measure to morphological alterations caused by environmental stressors such as reduced water quality and diseases. In the Senegal sole skin, four morphologically distinct layers were identified: cuticle, epidermis, dermis and hypodermis. The epidermis was composed of stratified epithelium containing three cellular layers: the outermost or mucosa layer, the middle or fusiform layer and the stratum germinativum or the basal layer. In the mucosa, two mucous cell types were differentiated: type A cells containing several round vesicles of different electron density and type B cells containing mucosomes of uniform electron density. Senegal sole have five pairs of gill arches, each containing two rows of well-developed and compactly organized primary filaments and secondary lamellae. Fingerprint-like microridges were observed on the surface of epithelial cells. The branchial lamellae epithelium consisted of different cell types: pavement, mucous and chloride. Between the chloride cells and the larger pavement cells, accessory cells were observed. Complexes of tight junctions and desmosomes were frequently observed between adjacent chloride and epithelial cells. Neutral mucosubstances and/or glycoconjugates were observed in the epidermis, dermis and hypodermis of S. senegalensis skin. Proteins rich in different amino acids, such as arginine and cysteine, reacted negatively or weakly positive in the epidermis, dermis and hypodermis. In gills, some mucous cells responded weakly positive to periodic acid-Schiff (PAS) reaction but were strongly stained with Alcian Blue at pH 0.5, 1 and 2.5. When Alcian Blue pH 2.5,PAS reaction was performed, most mucous cells were stained blue (carboxylated mucins) and some mucocytes stained purple, indicating a combination of neutral and acid mucins. Proteins rich in cysteine-bound sulphydryl (-SH-) and cystine disulphide (-S-S-) groups were strongly detected in branchial and epidermal mucous cells, whereas lysine, tyrosine and arginine containing proteins showed very weak staining in both epidermal and branchial mucous cells. Protein reactions were strongly positive in the pillar cells, except for those rich in tryptophan, whereas the branchial cartilaginous tissue did not show an important reaction. The performed lipid reactions were negative in goblet and chloride cells. It is concluded from this study that ultrastructural and cytohistochemical features of the Senegal sole skin and gills may serve as control structures in both natural and aquaculture systems to monitor or detect environmental stress responses at the histological level. [source] Elevated expression of bisecting N -acetylglucosaminyltransferase-III gene in a human fetal hepatocyte cell line by hepatitis B virusJOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY, Issue 12 2004JAE-KYOUNG SHIM Abstract Background and Aim:, UDP-N-acetylglucosamine: ,-D-mannoside ,-1,4 N-acetylglucosaminyltransferase III (GnT-III) is a key enzyme in N-glycan biosytnesis. Human GnT-III enzyme activity was found to be elevated in the serum of patients with hepatomas and liver cirrhosis and in hepatocellular carcinoma tissues. Therefore, to understand the relationship between the elevation in GnT-III activity and hepatitis B viral (HBV) hepartocarcinogenesis, we investigated GnT-III gene expression in the HBV-infected cells. Methods:, A cell line, HFH-T1, producing HBV was produced by natural infection of human fetal hepatocytes. A 170-bp band corresponding to the pre-S1 region of HBV was detected in the culture medium by polymerase chain reaction. Virions were also isolated from the culture medium by sucrose density gradient centrifugation. The synthesis of both ,-fetoprotein and albumin as an indicator that these cells were functional hepatocytes and the extent of differentiation was examined. Polymerase chain reaction and Western blot analysis using a monoclonal antibody, GT273, which was prepared using human aglycosyl recombinant GnT-III were used for HBV DNA and GnT-III detection. Results:, Two types of HBV-related particles were secreted into the culture medium; one was a Dane particle (40 nm in size) containing HBV DNA and the other was a subviral hepatitis B surface antigen particle (20 nm in size) that did not contain the viral genome. The secretion from the cell line was diminished by the number of passages and, thus, this cell was renamed as HFH-T2. A decreased level of the HBV was secreted from the cells after a rest period. HFH-T2 cells showed a weak staining for ,-fetoprotein and a moderate staining for albumin in the cytoplasm around the nucleus. High levels of a 0.7 kb DNA fragment originating from GnT-III DNA were detected in HFH-T2 cells. Western blot analysis using a monoclonal antibody, GT273, whixh was prepared using human aglycosyl recombinant GnT-III showed a single band, corresponding to Mr 63 kDa, whereas aglycosyl GnT-III showed a band at Mr 53 kDa, with a molecular weight difference of about 10 kDa. This indicates that HFH-T2 cells express glycosylated GnT-III. GnT-III activities were 347.2 ± 53.6 pmol/mg of protein/h in HFH-T2, 276 ± 26.3 in Hep3B, 252.5 ± 23.3 in HepG2 and 30.7 ± 3.4 in NIH-3T3. GnT-III activity was higher in HFH-T2 cells than in the hepatoma cell lines, Hep3B and HepG2. Conclusion:, A human fetal hepatocyte cell line was transformed by infection with HBV and the cell line expressed high levels of GnT-III as the levels of secretion of HBV decreased. The decrease in HBV secretion from HFH-T2 cells could be due to a high level of expression of GnT-III. Such a cell line could be used to investigate relationships between HBV infection and glycosyltransferase gene expression. Furthermore, this cell line will be useful in future studies on the effect of the expression of GnT-III on other glycosyltransferase. [source] Expression of zinc finger protein 105 in the testis and its role in male fertilityMOLECULAR REPRODUCTION & DEVELOPMENT, Issue 6 2010Huaxin Zhou Using an in silico approach, we identified a putative zinc finger domain-containing transcription factor (zinc finger protein 105, ZFP105) enriched in the adult mouse testis. RT-PCR analyses showed that Zfp105 was indeed highly expressed in adult mouse testis and that its expression was regulated during postnatal development. To further characterize Zfp105 expression, we generated a Zfp105:,-galactosidase (LacZ) knock-in reporter mouse line (Zfp105LacZ/+) in which a Zfp105:LacZ fusion gene was expressed. Whole-mount LacZ analyses of adult Zfp105LacZ/+ tissues showed robust LacZ staining in the testis, very weak staining in the ovary, and no staining in the spleen, liver, kidney, heart, lung, thymus, adrenal gland, uterus, or oviduct. Sectional LacZ staining showed that ZFP105 was highly expressed in pachytene spermatocytes. ZNF35, the human ortholog of Zfp105, was also highly expressed in human testis. Immunofluorescence analysis showed that ZNF35 was located primarily in the cytoplasm of male germ cells. More importantly, reduced male fertility was observed in adult Zfp105LacZ/LacZ mice. Histological studies showed the presence of undifferentiated spermatogenic cells in the lumen of seminiferous tubules at stage VII and in the epididymal lumen of adult Zfp105LacZ/LacZ mice. Taken together, our results suggest that ZFP105 is a male germ-cell factor and plays a role in male reproduction. Mol. Reprod. Dev. 77: 511,520, 2010. © 2010 Wiley-Liss, Inc. [source] Receptor for advanced glycation endproduct (RAGE),mediated nuclear factor-,B activation in vasculitic neuropathyMUSCLE AND NERVE, Issue 6 2004Karl-Matthias Haslbeck MD Abstract Binding of ligands to the receptor for advanced glycation endproducts (RAGE) results in activation of the proinflammatory transcription factor nuclear factor-kappaB (NF-,B) and subsequent expression of NF-,B,regulated cytokines. In order to determine whether engagement of RAGE contributes to the pathogenesis of vasculitic neuropathy, we studied the presence of the RAGE ligand N, -(carboxymethyl)lysine (CML), the receptor itself, NF-,B, and interleukin-6 (IL-6) in sural nerve biopsies of 12 patients with vasculitic neuropathies and 12 controls. In the patients, CML, RAGE, NF-,B, and IL-6 were localized in mononuclear cells, epineurial and endoneurial vessels and the perineurium. CML, RAGE, NF-,B, and IL-6 were expressed by CD4+, CD8+, and CD68+ cells invading the nerves. Controls showed only weak staining. These data suggest that the RAGE pathway plays a critical proinflammatory role in vasculitic neuropathy. Muscle Nerve 29: 853,860, 2004 [source] Immunohistochemical analysis of receptor tyrosine kinase signal transduction activity in chordomaNEUROPATHOLOGY & APPLIED NEUROBIOLOGY, Issue 1 2008J. H. Fasig Aims: Currently, there are no effective chemotherapeutic protocols for chordoma. Reports of receptor tyrosine kinase (RTK) expression in chordoma suggest that these tumours may respond to kinase inhibitor therapy. However, RTK signalling activity has not been extensively investigated in chordoma. Methods: A tissue microarray containing 21 cases of chordoma was analysed for expression of a number of proteins involved in signal transduction from RTKs by immunohistochemistry. Results: Platelet-derived growth factor receptor-,, epidermal growth factor receptor (EGFR), KIT and HER2 were detected in 100%, 67%, 33% and 0% of cases, respectively. Platelet-derived growth factor receptor-, staining was of moderate-to-strong intensity in 20 of 21 cases. In contrast, KIT immunoreactivity was weak and focal in each of the seven positive cases. Total EGFR staining was variable; weak staining for phosphorylated EGFR was detected in nine cases. Phosphorylated isoforms of p44/42 mitogen-activated protein kinase, Akt and STAT3, indicative of tyrosine kinase activity, were detected in 86%, 76% and 67% of cases, respectively. Conclusions: Chordomas commonly express RTKs and activated signal transduction molecules. Although there were no statistically significant correlations between the expression of any of the markers studied and disease-free survival or tumour location, the results nonetheless indicate that chordomas may respond to RTK inhibitors or modulators of other downstream signalling molecules. [source] Immunophenotypic Comparison of Salivary Gland Oncocytoma and Metastatic Renal Cell Carcinoma,THE LARYNGOSCOPE, Issue 6 2005John A. Ozolek MD Abstract Objectives/Hypothesis: The differential diagnosis of oncocytic neoplasms of salivary glands includes both primary and metastatic tumors, one of which is renal cell carcinoma. This study compared immunohistochemical staining characteristics of oncocytomas arising from salivary gland to metastatic renal cell carcinoma using a panel of markers. Study Design: Immunohistochemistry for cytokeratin 7 (CK7), cytokeratin 20 (CK20), epithelial membrane antigen (EMA), vimentin, CD10, and renal cell carcinoma marker (RCC) was performed on 10 oncocytomas and compared with ten metastatic renal cell carcinomas. Results: There were overlapping histologic findings in the oncocytomas and metastatic renal cell carcinomas, with oncocytomas displaying clear cell changes in 2 of 10 cases. CK7 was positive in 9 of 10 oncocytomas and CK20 in 8 of 10 (7/10 stained for both), and vimentin was only weakly positive in 4 of 10 oncocytomas. All oncocytomas were EMA positive, with membranous staining, and all were negative for CD10 and RCC. Metastatic renal cell carcinoma was strongly positive for vimentin, EMA, and CD10 in most cases. RCC and CK7 were variably positive in metastatic renal cell carcinomas (4/10), and only 1 of 10 showed weak staining with CK20. Conclusions: Salivary gland oncocytomas and metastatic renal cell carcinomas share some similar histologic and immunohistochemical characteristics. CD10 and CK20 were the most useful markers to distinguish metastatic renal cell carcinoma from oncocytomas in the salivary gland, whereas RCC, EMA, CK7, and vimentin are not as useful. [source] Antibodies produced by clonally expanded plasma cells in multiple sclerosis cerebrospinal fluid,ANNALS OF NEUROLOGY, Issue 6 2009Gregory P. Owens PhD Objective Intrathecal IgG synthesis, persistence of bands of oligoclonal IgG, and memory B-cell clonal expansion are well-characterized features of the humoral response in multiple sclerosis (MS). Nevertheless, the target antigen of this response remains enigmatic. Methods We produced 53 different human IgG1 monoclonal recombinant antibodies (rAbs) by coexpressing paired heavy- and light-chain variable region sequences of 51 plasma cell clones and 2 B-lymphocyte clones from MS cerebrospinal fluid in human tissue culture cells. Chimeric control rAbs were generated from anti-myelin hybridomas in which murine variable region sequences were fused to human constant region sequences. Purified rAbs were exhaustively assayed for reactivity against myelin basic protein, proteolipid protein, and myelin oligodendrocyte glycoprotein by immunostaining of transfected cells expressing individual myelin proteins, by protein immunoblotting, and by immunostaining of human brain tissue sections. Results Whereas humanized control rAbs derived from anti-myelin hybridomas and anti-myelin monoclonal antibodies readily detected myelin antigens in multiple immunoassays, none of the rAbs derived from MS cerebrospinal fluid displayed immunoreactivity to the three myelin antigens tested. Immunocytochemical analysis of tissue sections from MS and control brain demonstrated only weak staining with a few rAbs against nuclei or cytoplasmic granules in neurons, glia, and inflammatory cells. Interpretation The oligoclonal B-cell response in MS cerebrospinal fluid is not targeted to the well-characterized myelin antigens myelin basic protein, proteolipid protein, or myelin oligodendrocyte glycoprotein. Ann Neurol 2009;65:639,649 [source] Overexpression of S100A4 is closely related to the aggressiveness of gastric cancerAPMIS, Issue 5 2003YONG GU CHO Elevated levels of the calcium-binding protein S100A4 cause metastasis of benign rat mammary tumor cells. To investigate whether S100A4 plays an important role in the invasion and metastasis of gastric cancers, we examined the gene mutations in the coding regions and expression patterns of the S100A4 in gastric adenocarcinoma in Korea. Moderate to strong expression of S100A4 was found in 53 (68.8%) of the 77 gastric adenocarcinomas, whilst normal gastric epithelium either failed to stain or showed weak staining. Interestingly, S100A4 expression was more frequently observed in gastric cancer patients with advanced gastric cancer (p=0.039), positive lymph node metastasis (p=0.001), and peritoneal dissemination (p=0.022). No gene mutations were found in the analyzed genomic area in 77 gastric adenocarcinomas and 15 gastric cancer cell lines. We found one single nucleotide polymorphism without an amino acid change, A99G, in two cases. These data suggest that the overexpression of S100A4 may be closely related to the aggressiveness of gastric cancer in Korea. [source] | |||||||||||||||||||