Koide H, Nakamura T, Kimura K, Ebihara I, Nagai R, Tomino Y
Department of Medicine, Juntendo University School of Medicine, Tokyo, Japan.
Nephrol Dial Transplant. 1996 Sep;11(9):1769-74.
We reported that an embryonic type of non-muscle-type myosin heavy-chain isoform (SMemb) may be a molecular marker for phenotypic alteration in initial glomerular injury and that methyl-prednisolone has no effect on SMemb expression in glomeruli of rats with puromycin aminonucleoside (PAN) nephrosis. The present study was designed to assess whether SMemb mRNA and protein expression in glomeruli are affected by a low-protein diet in rats with PAN-induced nephrosis and in control rats.
Rats were divided into four groups: group 1, PAN-injected rats fed a standard diet containing 22% protein; group 2, PAN-injected rats fed a low-protein diet containing 6% protein, starting on the day of PAN injection; group 3, control rats fed a standard diet; group 4, control rats fed a low-protein diet for the same period. We prepared glomerular RNA and performed Northern blot analysis and immunohistochemistry in all groups.
Glomerular SMemb mRNA increased on days 2 and 4 (prior to and soon after the onset of proteinuria), but declined on day 8 (the peak of proteinuria). Myosin heavy-chain protein expression was evaluated immunohistochemically by use of three antibodies against SM1, SM2, and SMemb. SM1 and SM2 were absent from the glomeruli of rats with PAN nephrosis until day 20. The SMemb isoform was barely detectable in normal glomeruli, but substantial amounts of SMemb were demonstrated in the glomeruli of rats with PAN nephrosis. In the latter condition, the number of SMemb-positive glomerular epithelial cells increased on days 3 and 4, then decreased in subsequent days. Moreover, some mesangial cells became SMemb-positive transiently, returning to barely detectable levels on day 20. In addition, alpha-smooth-muscle actin, type I and III collagens were absent from the glomeruli of rats with PAN nephrosis until day 20. Urinary protein excretion was markedly suppressed by the 6% protein diet in PAN nephrosis. The low-protein diet reduced the increased mRNA expression of SMemb as well as the increased number of SMemb-positive cells in the glomeruli of rats with PAN nephrosis. However, the low-protein diet did not affect SMemb mRNA and protein levels in the glomeruli of control rats.
In rats with PAN nephrosis, findings suggest that restriction of dietary protein leads to a reduction in glomerular SMemb expression.
我们曾报道，一种胚胎型非肌肉型肌球蛋白重链异构体（SMemb）可能是初始肾小球损伤中表型改变的分子标志物，且甲基强的松龙对嘌呤霉素氨基核苷（PAN）肾病大鼠肾小球中SMemb的表达无影响。本研究旨在评估PAN诱导肾病大鼠及对照大鼠中，低蛋白饮食是否会影响肾小球中SMemb mRNA和蛋白的表达。
将大鼠分为四组：第1组，注射PAN的大鼠喂食含22%蛋白质的标准饮食；第2组，自注射PAN之日起，注射PAN的大鼠喂食含6%蛋白质的低蛋白饮食；第3组，对照大鼠喂食标准饮食；第4组，对照大鼠同期喂食低蛋白饮食。我们制备了所有组的肾小球RNA，并进行了Northern印迹分析和免疫组织化学检测。
肾小球SMemb mRNA在第2天和第4天（蛋白尿出现之前及之后不久）增加，但在第8天（蛋白尿峰值）下降。使用针对SM1、SM2和SMemb的三种抗体通过免疫组织化学评估肌球蛋白重链蛋白表达。直到第20天，PAN肾病大鼠的肾小球中均未检测到SM1和SM2。正常肾小球中几乎检测不到SMemb异构体，但PAN肾病大鼠的肾小球中显示出大量的SMemb。在后一种情况下，SMemb阳性肾小球上皮细胞数量在第3天和第4天增加，随后几天减少。此外，一些系膜细胞短暂地变为SMemb阳性，在第20天恢复到几乎检测不到的水平。此外，直到第20天，PAN肾病大鼠的肾小球中均未检测到α-平滑肌肌动蛋白、I型和III型胶原蛋白。PAN肾病中6%的蛋白质饮食可显著抑制尿蛋白排泄。低蛋白饮食降低了PAN肾病大鼠肾小球中SMemb mRNA表达的增加以及SMemb阳性细胞数量的增加。然而，低蛋白饮食并未影响对照大鼠肾小球中SMemb mRNA和蛋白水平。
在PAN肾病大鼠中，研究结果表明饮食蛋白的限制导致肾小球SMemb表达降低。
