Nangaku M, Miyata T, Suzuki D, Umezono T, Hashimoto T, Wada T, Yagi M, Nagano N, Inagi R, Kurokawa K
Molecular and Cellular Nephrology, Institute of Medical Sciences Tokai University School of Medicine, Kanagawa, Japan.
Kidney Int. 2001 Aug;60(2):641-52. doi: 10.1046/j.1523-1755.2001.060002641.x.
We recently cloned a new human mesangium-predominant gene, megsin. Megsin is a novel member of the serine protease inhibitor (serpin) superfamily. To elucidate functional roles of this gene, we cloned megsin in rodents and investigated its role in a rat nephritis model.
Megsin homologues were cloned from cultured rat and mouse mesangial cDNAs utilizing polymerase chain reaction (PCR) with degenerative primers. Expression of megsin in three different types of resident glomerular cells was investigated by PCR. Levels of megsin mRNA expression at various time points in the anti-Thy1 rat nephritis model were studied by semiquantitative PCR and Northern blotting analysis. In order to investigate megsin protein expression in anti-Thy1 nephritis rats, we raised antibody against rat megsin-specific synthetic peptide, with which immunohistochemical studies were performed.
Rat and mouse megsins were composed of 380 amino acids, which revealed 75.3 and 73.9% identity, respectively, with human megsin at the amino acid level. Characteristic features as an inhibitory serpin were conserved in both rat and megsin megsins. PCR analysis revealed expression of megsin in cultured mesangial cells but not in glomerular epithelial or endothelial cells. In anti-Thy1 nephritis rats, semiquantitative PCR and Northern blotting showed that expression of megsin mRNA was up-regulated in glomeruli at day 8. Immunohistochemical studies demonstrated the prominent accumulation of megsin in glomeruli at the same time point. Megsin was mainly localized in mesangial area. The megsin expression level returned to the basal level at day 28.
Sequences of megsin were well conserved among different species. Rat megsin was also predominantly expressed in mesangial cells. Expression of megsin was up-regulated at the peak of hypercellularity and matrix accumulation in the mesangioproliferative glomerulonephritis model, suggesting that megsin may participate in the process of glomerulosclerosis by modulating extracellular matrix deposition or cell survival.
我们最近克隆了一个新的人类系膜优势基因——megsin。Megsin是丝氨酸蛋白酶抑制剂（serpin）超家族的一个新成员。为阐明该基因的功能作用，我们在啮齿动物中克隆了megsin，并在大鼠肾炎模型中研究了其作用。
利用聚合酶链反应（PCR）和简并引物从培养的大鼠和小鼠系膜cDNA中克隆Megsin同源物。通过PCR研究Megsin在三种不同类型的肾小球固有细胞中的表达。通过半定量PCR和Northern印迹分析研究抗Thy1大鼠肾炎模型中不同时间点Megsin mRNA的表达水平。为研究抗Thy1肾炎大鼠中Megsin蛋白的表达，我们制备了针对大鼠Megsin特异性合成肽的抗体，并进行了免疫组织化学研究。
大鼠和小鼠Megsin均由380个氨基酸组成，在氨基酸水平上与人Megsin的同源性分别为75.3%和73.9%。大鼠和小鼠Megsin均保留了作为抑制性serpin的特征。PCR分析显示Megsin在培养的系膜细胞中表达，但在肾小球上皮细胞或内皮细胞中不表达。在抗Thy1肾炎大鼠中，半定量PCR和Northern印迹显示，在第8天肾小球中Megsin mRNA的表达上调。免疫组织化学研究表明，在同一时间点Megsin在肾小球中显著积聚。Megsin主要定位于系膜区。在第28天，Megsin表达水平恢复到基础水平。
Megsin序列在不同物种间保守性良好。大鼠Megsin也主要在系膜细胞中表达。在系膜增生性肾小球肾炎模型中，Megsin的表达在细胞增多和基质积聚的高峰期上调，提示Megsin可能通过调节细胞外基质沉积或细胞存活参与肾小球硬化过程。
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