GENETICS OF GLOMERULAR DISEASE IN MICE

多种多样的证据表明， 人类的肾小球硬化受基因控制。Os基因突变 导致先天性肾单位数量减少50% 肾小球肥大，标记指数增加。这种突变 在易感品系(ROP小鼠)中诱导肾小球硬化(GS)，但 在耐GS(C57小鼠)品系中不存在(J Clin Invest 1996，97：1-8)。 本研究的目的是确定肾单位减少 出生后在易感小鼠中诱发GS，以及是否进一步 肾单位质量减少(75%)将在抗性品系中诱导GS。 成年ROP+/+、ROP Os/+、C57+/+和C57 Os/+小鼠接受单侧 肾切除术(NX)。两周后肾小球体积增加 ROP+/+、C57+/+和ROP Os/+小鼠肾切除，C57Os/+小鼠未见肾切除。 ROP+/+NX和C57+/+NX小鼠肾小球标记指数增加。 而C57+/+Nx和C57Os/+Nx小鼠的肾小球没有变化， ROP+/+NX小鼠皮损较轻，ROP-Os/+NX小鼠GS较重。 ROP+/+NX组肾小球IIV胶原基因表达高于C57+/+组 NX。肾小球胶原、层粘连蛋白1和肌腱蛋白mRNAs增加 在肾切除2个月后，ROP/OS小鼠而不是C57Os/+小鼠。因此， 对成年肾单位减少的反应取决于小鼠的品系，因为 肾切除仅对ROP小鼠造成损害。ROP+/+中的病变 Nx没有假手术ROP os/+小鼠严重，这表明 肾单位减少发生的时间在 确定硬化反应的幅度，即使在 敏感品系。为了进一步确定该案件的性质 与肾小球硬化有关的基因我们鉴定了一种小鼠品系 未发展为肾小球硬化(C3H)，并与 携带OS突变的易硬化症小鼠(ROP)品系。这个 突变被用来加速或使更明显的趋势 患上硬化症。F1动物与ROP背景的回交 结果显示，由此产生的F2代小鼠有不同程度的 3个月龄肾小球硬化。因此，我们可以演示 有一个明确的遗传易感性的发展 小鼠肾小球硬化。这次调查的下一步将是 试图对某些基因座进行定位和表征 这是那些损伤最严重的小鼠所特有的。自.以来 OS基因与肾脏发育有关，我们试图对其进行定位 使用40个SSLP的基于PCR的作图方法的这种突变 位于8号染色体中心的标记。根据 这些动物已经回交了几代，因为这种突变是 在1952年诱导的时候，我们预计会发现一个非常小的区域。令人惊讶的是， 我们的作图数据揭示了一个区域的连锁平衡失调 10厘米长的。通过随后的G-显带分析和荧光 原位杂交，我们发现无论是大倒位还是 染色体的这一部分似乎存在较大的缺失。我们 现在必须考虑到缺陷是由于一个或多个较小的 染色体异常。由于血脂异常被认为是 作为肾脏损伤的潜在来源，我们描述了肾脏 诱导性血脂异常小鼠的表型。我们选择了学习 转基因LCAT和Apo E基因敲除小鼠。vt.在.上 抗硬化症背景我们发现LCAT小鼠有脂质 肾小球内皮细胞和系膜细胞内的包裹体，但有 是非常轻微的硬化症。在载脂蛋白E中也有类似的发现 Ko老鼠。我们现在正在将这些异常现象转化为一种容易出现硬化症的 鼠标背景。

Multiple lines of evidence suggest that the propensity to develop glomerulosclerosis in man is under genetic control. The Os mutation induces a congenital 50% reduction in nephron number and both hypertrophy and increased labeling index in glomeruli. This mutation induced glomerulosclerosis (GS) in a susceptible strain (ROP mice), but not in a GS resistant (C57 mice) strain (J Clin Invest 1996, 97:1-8). The purposes of this study were to determine whether nephron reduction after birth induced GS in susceptible mice, and whether further reduction in nephron mass (75%) would induce GS in resistant strains. Adult ROP +/+, ROP Os/+, C57 +/+ and C57 Os/+ mice underwent unilateral nephrectomy (NX). Glomerular volume increased two weeks after nephrectomy in ROP +/+, C57 +/+ and ROP Os/+ but not in C57 Os/+ mice. Glomerular labeling index increased in ROP +/+ NX and C57 +/+ NX mice. Whereas C57 +/+ NX and C57 Os/+ NX mice showed no glomerular changes, ROP +/+ NX mice exhibited mild lesions and ROP Os/+ NX had severe GS. Glomerular 1IV collagen mRNA was higher in ROP +/+ NX than in C57 +/+ NX. 1IV collagen, laminin 1, and tenascin glomerular mRNAs increased in ROP/Os but not C57 Os/+ mice 2 months after nephrectomy. Thus, the response to adult nephron reduction depended on the mouse strain, since nephrectomy induced lesions only in ROP mice. The lesions in ROP +/+ NX were less severe than in sham operated ROP Os/+ mice, suggesting that the time at which nephron reduction occurs was critical in determining the amplitude of the sclerotic response, even in a susceptible strain. In order to further determine the nature of the genes implicated in glomerulosclerosis we identified a mouse strain that did not develop glomerulosclerosis (C3H), and crossed it with a sclerosis-prone mouse (ROP) strain carrying the Os mutation. The mutation was used to accelerate, or make more obvious, the tendency to develop sclerosis. A backcross of the F1 animals to the ROP background revealed that the resulting F2 generation mice had varying degrees of glomerulosclerosis at the age of 3 months. Thus, we could demonstrate that there was a clear-cut inheritance of the susceptibility to develop glomerulosclerosis in mice. The next steps in this investigation will be to attempt the localization and characterization of certain loci which are peculiar to those mice with the most severe lesions. Since the Os gene is of interest in renal development, we tried to localize this mutation using a PCR based mapping approach using forty SSLP markers in the center of chromosome 8. Based on the number of generations the animals have been backcrossed, since the mutation was induced in 1952, we expected to find a very small region. Surprisingly, our mapping data revealed a linkage dysequilibrium involving a region of 10 cM in length. By subsequent G-banding analysis and fluorescence in situ hybridization, we found that neither a large inversion nor a large deletion seems to be present in this part of the chromosome. We now have to consider that the defect is due to one or more smaller chromosomal abnormalities. Since lipid abnormalities are thought to be a potential source of renal injury, we characterized the renal phenotype of mice with induced lipid abnormalities. We chose to study mice transgenic for LCAT and Apo E knockout mice. On a sclerosis-resistant background we found that LCAT mice had lipid inclusions in glomerular endothelial and mesangial cells, but that there was very little sclerosis. Similar findings were present in the Apo E ko mice. We are now crossing these abnormalities onto a sclerosis-prone mouse background.