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1.
Signal detection of endothelin receptor subtypes in human cirrhotic liver by a new in situ hybridization method 总被引:1,自引:0,他引:1
Endothelin (ET) has been implicated in the regulation of hepatic microcirculation and development of portal hypertension, but the role of ET in cirrhosis of the liver has not yet been elucidated. We present here the application of peptide nucleic acid (PNA) probes in a fast and sensitive in situ hybridization (ISH) method for localizing the mRNA of endothelin receptor subtypes in formalin-fixed, paraffin-embedded sections of normal and cirrhotic human liver. The results of ISH using synthetic FITC-labeled PNA probes combined with the catalyzed signal amplification (CSA) system were compared with those using the standard detection system. It was indicated that the CSA-ISH protocol is more sensitive for the detection of mRNA target than the standard ISH protocol. Our results with CSA-ISH showed that the expression of mRNA for the endothelin B receptor was significantly upregulated in hepatic sinusoidal lining cells in cirrhotic human liver tissues compared to control normal liver tissue. Therefore, the CSA detection system may facilitate and enhance the use of in situ hybridization protocols, and CSA-ISH will be used as an important diagnostic technique in the future. 相似文献
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Comparison of matrix metalloproteinase expression in normal and cirrhotic human liver 总被引:16,自引:0,他引:16
Ralf Lichtinghagen Karin Breitenstein Burkhard Arndt Tanja Kühbacher K. H. W. Böker 《Virchows Archiv : an international journal of pathology》1998,432(2):153-158
To study the extend of ongoing tissue remodelling in end-stage cirrhosis, the expression of different matrix metalloproteinases
[interstitial collagenase (MMP-1), Mr 72000 gelatinase (MMP-2), stromelysin-1 (MMP-3) and stromelysin-3 (MMP-11)] and of TIMP-1 was studied in 13 cirrhotic livers
explanted at transplantation. The results were compared with those obtained in normal liver. Western blot, northern blot,
ELISA, RT-PCR and zymogram analysis were used. Proenzymes of stromelysin-1 and -3, interstitial collagenase and Mr 72000 gelatinase were positive in normal liver, while activated enzymes were not detectable in western blot analysis. In
cirrhosis proenzyme levels of the studied MMPs were reduced to a mean of 60–70%, but mRNA expression and gelatin-degrading
activity increased. TIMP-1 expression was detectable on mRNA level and by ELISA in normal liver and also increased in cirrhosis.
Our results show that mRNA expression of certain matrix metalloproteinases is increased in end-stage liver cirrhosis, while
the amount of proenzyme is decreased, indicating enhanced MMP proenzyme turnover. These data suggest that besides increased
TIMP-1 activity, altered MMP expression may also play a part in fibroproliferation in liver disease.
Received: 6 June 1997 / Accepted: 1 September 1997 相似文献
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Increased levels of endothelin-1 and differential endothelin type A and B receptor expression in scleroderma-associated fibrotic lung disease. 总被引:10,自引:0,他引:10 下载免费PDF全文
D. J. Abraham R. Vancheeswaran M. R. Dashwood V. S. Rajkumar P. Pantelides S. W. Xu R. M. du Bois C. M. Black 《The American journal of pathology》1997,151(3):831-841
In addition to their vasoactive action, endothelins are potent peptides in the regulation of both cell proliferation and the turnover of extracellular matrix. Using immunohistochemical, autoradiographic, and molecular analyses, we have studied the localization and expression of endothelin-1 and endothelin A (ETA) and B (ETB) receptors in scleroderma-associated fibrotic lung disease. Increased ET-1 immunoreactivity was found in sclerotic tissue compared with control and was associated with the vasculature, pulmonary interstitium, and bronchial and alveolar epithelium. Microautoradiographic analysis after 125I-labeled ET-1 binding showed a two- to threefold increase in the expression of total ET-1 receptors in scleroderma lung tissue localized to the alveolar epithelium and the pulmonary interstitium which was composed of mainly fibroblastic cells with macrophages and some microvessels. RNAse protection assay revealed significantly reduced ETA receptor and slightly raised ETB message levels in systemic sclerosis lung. Surface expression of functional ET receptors was examined by targeted receptor blocking using mixed and receptor-subtype-selective ligands. A consistent decrease in ETA receptor binding sites was noted primarily within the interstitium and vasculature, in contrast to a slight increase in ETB receptors. Elevated ET-1 and the cell-specific pattern of endothelin receptor expression suggest that the endothelins may represent important mediators that influence the pathology of scleroderma-associated lung disease and other fibrotic conditions. 相似文献
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Expression of the hepatic endothelin system in human cirrhotic livers 总被引:11,自引:0,他引:11
Ikura Y Ohsawa M Naruko T Muraguchi T Hirayama M Suekane T Fukushima H Sugama Y Shirai N Kayo S Yoshimi N Ehara S Tanzawa K Ueda M 《The Journal of pathology》2004,204(3):304-310
It is considered that endothelin-1 participates in the development of liver cirrhosis and it has been recognized that every component of the endothelin system is upregulated in cirrhotic livers. However, the expression pattern of this system, including interaction between its components, is not fully understood in human livers. In this study, the expression pattern of the endothelin system was examined. Immunohistochemical analysis for endothelin-1, endothelin receptors and endothelin-converting enzyme was performed in 16 cirrhotic and 17 normal human liver tissues. Peptides, proteins, and RNAs extracted from the livers were also investigated using quantitative assays for the components of the hepatic endothelin system. Hepatic endothelin-1 levels were significantly higher in cirrhotic livers (0.084 +/- 0.052 pg/mg wet liver) than in normal livers (0.041 +/- 0.032 pg/mg; p < 0.01), and were closely related to the severity of liver fibrosis and portal hypertension. Immunoreactivity for endothelin-1, endothelin receptors, and endothelin-converting enzyme was detected mainly in fibrous areas and in the hepatic vasculature, and was enhanced in cirrhosis. Although there was a negative correlation between the expression of receptor mRNA and the hepatic endothelin-1 level, the amounts of the mRNAs were greater in cirrhotic livers than in normal livers. However, expression of endothelin-converting enzyme in cirrhotic livers was increased at the protein level but was relatively reduced at the mRNA level. These findings suggest that the hepatic endothelin system is activated in human cirrhotic livers in association with worsening of the disease, but that the regulation of the components of this system in this disorder is complex. 相似文献
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Brigitte Le Bail Saadia Faouzi Liliane Boussarie Charles Balabaud Paulette Bioulac-sage Jean Rosenbaum 《The Journal of pathology》1997,181(3):330-337
Extracellular matrix (ECM) plays a major role in cell differentiation, proliferation, and gene expression, both in physiological and in pathological conditions. Immunohistochemistry has been used to investigate modifications of ECM and related receptors, the integrins, in 26 small nodular lesions developed in human cirrhotic livers, on the basis that these lesions could represent sequential steps of hepatocarcinogenesis: the lesions were 16 macroregenerative nodules (MRNs), either of ordinary (n=5) or atypical (n=11) type, and ten small (<15 mm) hepatocellular carcinomas (HCCs). Data were compared with those obtained in the surrounding cirrhotic tissue, in large HCCs, and in normal liver. The results indicate similarities between ordinary MRNs and cirrhosis, on the one hand, and between atypical MRNs and small HCCs, on the other. Strong and homogeneous deposition of collagen type IV and laminin in sinusoids and overexpression of α6 integrin by sinusoidal cells and hepatocytes were especially noticeable in dysplastic areas characteristic of atypical MRNs, as in small HCCs. In addition, the staining of α 2 and α6 integrins in MRNs revealed the presence of widespread atypical ductular proliferation expanding from periportal and perinodular areas, containing epithelial cells with transitional (hepato-biliary) phenotype. These findings suggest a transition from atypical MRNs to small HCCs and a possible role for liver epithelial precursor cells (‘stem cells’) in the development and evolution of MRNs. © 1997 John Wiley & Sons, Ltd. 相似文献
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目的 采用反转录聚合酶链反应 (RT -PCR)法获得人α型叶酸受体 (folatereceptoralpha,FOLR - 1,FR - 1)基因序列 ,构建融合表达载体 pGEXFR - 1并进行表达。 方法将RT -PCR法扩增的FOLR - 1基因 ,克隆入融合表达载体 pGEX - 4T - 2中 ,使其受控于Ptac启动子 ,转化大肠杆菌DH5α。经异丙基 - β -D -硫代半乳糖苷 (IPTG)诱导 ,用SDS -PAGE检测表达产物。结果SDS -PAGE显示 ,FOLR - 1表达产物的相对分子质量 (Mr)约为 5 5 0 0 0 ,与预期的结果相符。Western -blot证实 ,在相应Mr处 ,有GST-FR1融合蛋白的显示条带。结论成功地构建了FOLR -1基因的融合表达载体并获得表达 ,为进一步探讨卵巢癌的免疫治疗提供了实验依据 相似文献
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Nakatani T Tanabe S Han YS Kayo S Yoshimi N Hai E Shirai N Ikura Y Ohsawa M Ueda M 《International journal of molecular medicine》2003,11(2):153-156
Transplant renal arteriosclerosis (TRA), characterized by concentric neointimal thickening, is one of the major causes of chronic rejection in human kidney transplantation. Endothelin-1 is known to be a powerful vasoconstrictor and a vascular smooth muscle cell mitogen. Previous experimental studies have shown that endothelin-A receptor (ET(A) receptor) is expressed selectively in vascular smooth muscle cells, and is the major mediator of endothelin-1-induced effects. However, ET(A) receptor expression in human renal allografts has not been reported. In this study, we immunohistochemically investigated the expression of ET(A) receptor in relation to the development of TRA, using nine human renal allografts removed due to rejection and ten normal kidneys as controls. In intrarenal arteries of normal kidneys, medial smooth muscle cells showed weak expression of ET(A) receptor. In intrarenal arteries with TRA of human renal allografts, increased expression of ET(A) receptor was found in medial smooth muscle cells, and distinct expression of ET(A) receptor was also detected in smooth muscle cells within the neointima. These results suggest that enhanced expression of ET(A) receptor may induce an increase in the local proliferative and vasoconstrictive effects of endothelin-1 in human renal allografts. 相似文献
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Collagens of normal and cirrhotic human liver 总被引:2,自引:0,他引:2
The collagens of normal and end-stage alcoholic cirrhotic human liver were investigated. Mild conditions of pepsin digestion were employed to preserved the more pepsin sensitive type IV collagen molecules while increasingly more harsh conditions were used which solubilized the type I and type III hepatic collagens. The total hepatic collagen content was elevated from 1.8-2.1 in normal liver to 7.3-8.2 percent in the end-stage alcoholic liver. The graded pepsin digestion resulted in solubilization of 76-90 percent of the total hepatic collagen. The genetically distinct types of collagen were fractionated into types I, III, IV and V preparations using judicious salt precipitation from dilute acid and neutral pH solutions. The average distribution of collagen was 42.5, 39.5, 6.9 and 10.6 percent types I, III, IV and V collagen respectively in normal liver compared to 56.6, 28.0, 5.5 and 9.6 percent respectively in the end-stage alcoholic livers. An additional 0.6 and 0.5 percent in normal and end-stage cirrhotic liver respectively were located in a fraction separating out of the salt concentration required for type VI collagen. The type IV collagen of basement membrane was separated into the constituent 95Kda alpha 1(IV) and alpha 2(IV) polypeptide chains by Agarose A 5m gel filtration with subsequent (carboxymethyl) CM-cellulose chromatography. These chains were indistinguishable from similar chains of type IV collagen isolated previously from human placenta. The type V collagen alpha chains, alpha 1(V) and alpha 2(V) were purified by a combination of Diethylaminoethyl (DEAE) and CM-cellulose chromatography. The individual chains were fragmented with cyanogen bromide to yield 9 peptides from the alpha 1(V) chain and 10 peptides from the alpha 2(V) collagen chain. These were purified and analyzed for amino acid content and molecular weight. The study suggests that all collagen types were elevated in the end-stage alcoholic liver, but type I collagen was disproportionately increased over all other collagens. These results are consistent with previous studies which demonstrated an elevated type I collagen in other fibrotic conditions such as lung fibrosis and hypertrophic scar. They do not, however, agree with the suggested disproportionate elevation of type V collagen in human alcoholic liver. The reasons for this discrepancy are discussed. 相似文献
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The expression of neurokinin-1 receptor at striatal and pallidal levels in normal human brain 总被引:2,自引:0,他引:2
To further our knowledge of the site of action of substance P (SP) in the human basal ganglia, we applied single- and double-antigen localization methods to human postmortem tissue to compare the distribution of SP and its high affinity receptor neurokinin-1 (NK1R) at striatal and pallidal levels. The human striatum was found to harbor numerous heterogeneously distributed aspiny neurons that expressed NK1R. Most of them were of small size, but a moderate number of large-sized neurons and a small number of medium-sized neurons also expressed NK1R. The medium-sized NK1R-positive neurons coexpressed parvalbumin and appear to represent a hitherto unknown striatal interneuron. The three types of striatal NK1R-positive neurons were preferentially localized in the peripheral region of the striosomes, which were identified by their intense immunostaining for the limbic system-associated membrane protein. Numerous NK1R expressing neurons also occurred in both external (GPe) and internal (GPi) segments of the globus pallidus, as well as in the ventral pallidum (GPv). There was a marked decreasing rostrocaudal gradient in the number of these neurons in the GPe, but not in the GPi. A multitude of smooth and highly branched SP-immunoreactive fibers pervaded the entire pallidal complex and some of these fibers were in close contact with NK1R-positive neurons in the GPi, as well as in the rostral portion of the GPe. The latter result reveals that the so-called 'direct' striatofugal pathway provides SP-immunoreactive collaterals to the GPe, a finding that is at odd with the current model of basal ganglia organization. 相似文献
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Ornit Chiba-Falek Marshall Nichols Sunil Suchindran John Guyton Geoffrey S Ginsburg Elizabeth Barrett-Connor Jeanette J McCarthy 《BMC medical genetics》2010,11(1):9
Background
Several studies have noted that genetic variants of SCARB1, a lipoprotein receptor involved in reverse cholesterol transport, are associated with serum lipid levels in a sex-dependent fashion. However, the mechanism underlying this gene by sex interaction has not been explored. 相似文献13.
目的探讨人类细小病毒B19感染与结直肠癌的关系。方法运用巢式PCR检测50例结直肠癌患者的肿瘤组织和对应的癌周结直肠组织以及10例正常成人结肠组织中的B19DNA,免疫组化检测B19结构蛋白VP1/VP2。结果肿瘤组织B19DNA阳性率为96%,显著高于癌周(60%)及正常对照(60%)。免疫组化显示,腺癌组织82%VP1/VP2蛋白表达,显著高于癌周组织(30%)及正常对照(20%)。结论人细小病毒B19可能在结直肠癌的发生中起一定作用,为探讨结直肠癌的发病机制提供新思路。 相似文献
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Differential adrenomedullin release and endothelin receptor expression in distinct subpopulations of human airway smooth-muscle cells 总被引:2,自引:0,他引:2
Upton PD Wharton J Davie N Ghatei MA Smith DM Morrell NW 《American journal of respiratory cell and molecular biology》2001,25(3):316-325
Although adrenomedullin (ADM) is implicated in the control of airway tone, regulation of ADM release from airway smooth-muscle cells (ASMCs) has not been explored. Preliminary experiments have indicated that human ASMC populations were heterogeneous in their rate of ADM release and expression of endothelin (ET)(A) and ET(B) receptors. We isolated these phenotypically distinct ASMCs from explants derived from the same airway segment. ASMCs possessing exclusively ET(A) receptors appeared smaller and proliferated faster than ET(A)/ET(B) isolates. Macroautoradiographic analysis confirmed the presence of both receptors in human bronchi. ADM release and messenger RNA expression was greater in ET(A)/ET(B) isolates compared with ET(A) isolates. No measurable ET release was detected from ASMCs. Exogenous ET-1 (1 to 100 nM) more potently stimulated the release of ADM from ET(A)/ET(B) compared with ET(A) isolates. In addition, ET-3 (1 to 100 nM) stimulated ADM release only from ET(A)/ET(B) isolates, implicating the ET(B) receptor in this response. Exogenous ET-1 potentiated platelet- derived growth factor-stimulated [3H]thymidine uptake in ET(A)/ ET(B) but not ET(A) isolates. ET-3 did not affect [3H]thymidine uptake in either cell type. Possession of ET(A)/ET(B) receptors is associated with higher rates of ADM release and slower proliferation, but a capacity for ET-1 stimulated DNA synthesis via ET(A) receptors. These results support a paracrine role for the regulation of ADM release predominantly via the ET(B) receptor in human ASMCs. 相似文献
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Although a critical role of the endothelin (ET) system in differentiation of neural crest cells has been reported, implication
of the ET system in human neuroblastic tumors has not been fully elucidated. We immunohistochemically examined for localization
of ET-1, ET-3, ET-A receptor (ET-A), and ET-B receptor (ET-B) in 24 ganglioneuromas, 8 ganglioneuroblastomas, 37 neuroblastomas,
14 normal sympathetic ganglia, and 10 fetal adrenal glands with regard to neuroblastic cell differentiation. Neuroblasts in
fetal adrenal glands expressed ET-B (100%) alone. Immature ganglionic cells in sympathetic ganglia of fetus frequently expressed
ET-1 (86%) and ET-B (100%), while ET-A was occasionally detected (28%). Ganglionic cells of mature adult ganglia consistently
harbored ET-1 (100%) and, infrequently, ET-3 (21%) or ET-B (29%). Expression of ET-1 and ET-B was closely associated with
tumor cell differentiation: the expression frequency of ET-1 or ET-B was 16% or 46% in neuroblastomas; 100% or 88% in ganglioneuroblastomas;
and 96% or 92% in ganglioneuromas. In contrast, ET-3 and ET-A showed no association with tumor cell differentiation: the expression
frequency of ET-3 or ET-A was 26% or 14% in neuroblastomas; 63% or 13% in ganglioneuroblastomas; and 29% or 21% in ganglioneuromas.
In conclusion, ET-1 and ET-B are expressed with differentiation of neuroblastic tumors. 相似文献