Functional genomics for renal diseases

肾脏疾病的功能基因组学

• 批准号: 11307016
• 负责人: KUROKAWA Kiyoshi
• 金额: $ 24.45万
• 依托单位: Tokai University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A).
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11307016/
• 关键词: 3'-directed cDNA library; mesangial cells; mesangium-specific transcription factor; megsin; mesangioproliferative glomenilonephlitis; molecular biology; メサンギウム細胞特異的遺伝子; 3'-directed; cDNA; library法

Hemodialysis due to end-stage renal diseases is a major social and economic problem in Japan because about 170,000 patients are treated at present, and the number has been increasing by about 13,000 every year. and because the medical expenses for the treatment of dialysis patients has risen up to more than a trillion U.S.dollars per year which takes up approximately four percent of total medical expenses in Japan. Chronic glomerulonephritis and diabetic nephropathy are two major underlying diseases which account for about 80% of the total number of patients with renal insufficiency. However, their exact mechanisms are still unknown at the molecular levels and therefore there is no effective therapy. The purposes of the projects in progress in our laboratory is 1) to elucidate the molecular pathogenesis of renal diseases, such as chronic glomerulonephritis and diabetic nephropathy, and 2) to develop novel therapeutic approaches to prevent or retard the progression of renal diseases.Mes … More angial cells play an important role in maintaining a structure and function of the glomerulus and in the pathogenesis of glomerular diseases. The proliferation of mesangial cells and the accumulation of extracellular mesangial matrix are primary events leading to the progression to glomerulosclerosis in patients with a variety of glomerular disorders such as chronic glomerulonephritis and diabetic nephropathy, two major causes of end-stage renal failure. The purposes of the present study were 1) to perform a molecular biological quantification of genes expressed in cultured human mesangial cells and 2) to identify specific genes expressed in mesangial cells. To obtain quantitative information of expressed genes in mesangial cells, we employed a 3'-directed regional cDNA library from human mesangial cells. With this approach, we can avoid variable cloning efficiencies reflecting the size of cDNA.we randomly chose almost 2000 transformant colonies, amplified their cDNA moieties by PCR and determined their cDNA sequences. Since the sequences at the 3'-region are unique, sequencing data from about 150-300 nucleotides were sufficient to characterize the gene. Redundancy of the same sequence represents the abundance of corresponding transcript in cells. The "expression profiles" in mesangial cells were apparently different from those obtained from other libraries, which confirmed specificity of "expression profiles" in different cells and organs. And, we identified 5 unknown genes, termed megsin, meg-1, -2, -3, and -4, specifically expressed in mesangial cells.Megsin is a new member of the serine protease inhibitor (serpin) superfamily. Our previous studies strongly suggest the role of megsin in the pathogenesis of human glomerular diseases, but its exact biological significance has not been clear. We produce human megsin transgenic mice. Overexpression of megsin leads to progressive mesangial matrix expansion and to an increase in the number of mesangial cells. Megsin has thus a biologically relevant influence on mesangial function. Less

终末期肾病导致的血液透析是日本的一个重大社会和经济问题，目前约有17万名患者接受治疗，并且这一数字每年以约1.3万人的速度增加。因为每年用于治疗透析患者的医疗费用已高达上万亿美元，约占日本总医疗费用的百分之四。慢性肾小球肾炎和糖尿病肾病是两大基础疾病，约占肾功能不全患者总数的80%。然而，它们的确切机制在分子水平上仍然未知，因此没有有效的治疗方法。我们实验室正在进行的项目的目的是 1) 阐明肾脏疾病的分子发病机制，如慢性肾小球肾炎和糖尿病肾病，2) 开发新的治疗方法来预防或延缓肾脏疾病的进展。 Mes … More 血管细胞在维持肾小球的结构和功能以及肾小球肾病的发病机制中发挥着重要作用。 肾小球疾病。系膜细胞的增殖和细胞外系膜基质的积累是导致患有多种肾小球疾病（例如慢性肾小球肾炎和糖尿病肾病）的患者进展为肾小球硬化的主要事件，这是终末期肾衰竭的两个主要原因。本研究的目的是 1) 对培养的人系膜细胞中表达的基因进行分子生物学定量，2) 鉴定系膜细胞中表达的特定基因。为了获得系膜细胞中表达基因的定量信息，我们使用了来自人系膜细胞的3'定向区域cDNA文库。通过这种方法，我们可以避免反映 cDNA 大小的可变克隆效率。我们随机选择近 2000 个转化体菌落，通过 PCR 扩增其 cDNA 部分并确定其 cDNA 序列。由于 3' 区域的序列是独特的，因此约 150-300 个核苷酸的测序数据足以表征该基因。相同序列的冗余代表细胞中相应转录本的丰度。系膜细胞中的“表达谱”与其他文库中获得的表达谱明显不同，这证实了不同细胞和器官中“表达谱”的特异性。并且，我们鉴定了 5 个未知基因，称为 megsin、meg-1、-2、-3 和 -4，专门在系膜细胞中表达。Megsin 是丝氨酸蛋白酶抑制剂 (serpin) 超家族的新成员。我们之前的研究强烈表明megsin在人类肾小球疾病发病机制中的作用，但其确切的生物学意义尚不清楚。我们生产人 megsin 转基因小鼠。 megsin 的过度表达导致进行性系膜基质扩张和系膜细胞数量增加。因此，Megsin 对系膜功能具有生物学相关的影响。较少的

项目成果

Miyata T et al: "Apolipoprotein E2/E5 variants in lipoprotein glomerulopathy recurred in transplanted kidney."J Am Soc Nephrol. 10(7). 1590-1595 (1999)

Miyata T 等人：“移植肾中复发了脂蛋白肾小球病中的载脂蛋白 E2/E5 变异。”J Am Soc Nephrol。

Miyata T, Sugiyama S, Nangaku M, Suzuki D, Uragami K, Inagi R, Kurokawa K: "Apolipoprotein E2/E5 variants in lipoprotein glomerulopathy recurred in transplanted kidney"J Am Soc Nephrol. 10 (7). 1590-1595 (1999)

Miyata T、Sugiyama S、Nangaku M、Suzuki D、Uragami K、Inagi R、Kurokawa K：“移植肾中复发的脂蛋白肾小球病中的载脂蛋白 E2/E5 变异”J Am Soc Nephrol。

Kawada N et al.: "Increased oxidative stress in mouse kidneys with unilateral ureteral obstruction"Kidney Int. 56. 1004-1013 (1999)

Kawada N 等人：“单侧输尿管梗阻导致小鼠肾脏氧化应激增加”Kidney Int。

Suzuki D et. al: "Immunohistochemical evidence for an increased oxidative stress and carbonylmodification of proteins in diabetic glomerular lesions."J Am Soc Nephrol. 10(4). 822-832 (1999)

铃木 D 等.

Kawada N et. al: "Increased oxidative stress in mouse kidneys with unilateral ureteral obstruction."Kidney Int. 56(3). 1004-1013 (1999)

川田奈等人。