Glomerular mesangial cell (MC) proliferation is one of the causative factors of glomerular diseases and one of their prominent pathological features. Rapamycin can inhibit MC proliferation and slow the progression to chronic renal fibrosis. The present study was designed to observe the role of rapamycin in MC proliferation and to explore the mechanism by which rapamycin acts on Akt and MAPK/ERK1/2 pathways in mesangial cells. MTT assay and flow cytometry were used to evaluate the proliferation and the cell cycle phase of glomerular mesangial cells respectively. The mRNA expression level of p70S6K was detected by RT-qPCR. Western blotting was performed to determine p70S6K, PI3K/Akt, and PI3K/MAPK protein expression. We found that rapamycin could reduce mesangial cell proliferation and arrest the cell cycle in the G1 phase, however the inhibition effect of 1000 nmol/L rapamycin was not higher than that in the 100 nmol/L group. The results of western blotting showed that 1000 nmol/L rapamycin more significantly inhibited the phosphorylation of p70S6K than 100 nmol/L, suggesting there should be another signaling pathway that activates the proliferation of MCs. Moreover, our results revealed that 1000 nmol/L rapamycin led to Raf1-MEK1/2-ERK pathway activation through a p70S6K-PI3K-mediated feedback loop in MCs. This study demonstrated that high-dose rapamycin leads to ERK1/2 activation through a p70S6K/PI3K/MAPK feedback loop in rat MCs, thus reducing the inhibitory effect of rapamycin on MC proliferation.

肾小球系膜细胞（MC）增殖是肾小球疾病的致病因素之一，也是其突出的病理特征之一。雷帕霉素可抑制 MC 增殖并减缓慢性肾纤维化的进展。本研究旨在观察雷帕霉素在MC增殖中的作用，并探讨雷帕霉素作用于系膜细胞Akt和MAPK/ERK1/2通路的机制。MTT法和流式细胞仪分别用于评价肾小球系膜细胞的增殖和细胞周期阶段。通过 RT-qPCR 检测 p70S6K 的 mRNA 表达水平。进行蛋白质印迹以确定 p70S6K、PI3K/Akt 和 PI3K/MAPK 蛋白表达。我们发现雷帕霉素可以减少系膜细胞增殖并将细胞周期阻滞在G1期，但1000 nmol/L雷帕霉素的抑制作用不高于100 nmol/L组。Western blotting结果显示，1000 nmol/L雷帕霉素比100 nmol/L更显着抑制p70S6K的磷酸化，提示应该有另一个信号通路激活MCs的增殖。此外，我们的结果显示，1000 nmol/L 雷帕霉素通过 MCs 中 p70S6K-PI3K 介导的反馈回路导致 Raf1-MEK1/2-ERK 通路激活。本研究表明，大剂量雷帕霉素通过 p70S6K/PI3K/MAPK 反馈环在大鼠 MCs 中导致 ERK1/2 激活，从而降低雷帕霉素对 MC 增殖的抑制作用。Western blotting结果显示，1000 nmol/L雷帕霉素比100 nmol/L更显着抑制p70S6K的磷酸化，提示应该有另一个信号通路激活MCs的增殖。此外，我们的结果显示，1000 nmol/L 雷帕霉素通过 MCs 中 p70S6K-PI3K 介导的反馈回路导致 Raf1-MEK1/2-ERK 通路激活。本研究表明，大剂量雷帕霉素通过 p70S6K/PI3K/MAPK 反馈环在大鼠 MCs 中导致 ERK1/2 激活，从而降低雷帕霉素对 MC 增殖的抑制作用。Western blotting结果显示，1000 nmol/L雷帕霉素比100 nmol/L更显着抑制p70S6K的磷酸化，提示应该有另一个信号通路激活MCs的增殖。此外，我们的结果显示，1000 nmol/L 雷帕霉素通过 MCs 中 p70S6K-PI3K 介导的反馈回路导致 Raf1-MEK1/2-ERK 通路激活。本研究表明，大剂量雷帕霉素通过 p70S6K/PI3K/MAPK 反馈环在大鼠 MCs 中导致 ERK1/2 激活，从而降低雷帕霉素对 MC 增殖的抑制作用。我们的结果表明，1000 nmol/L 雷帕霉素通过 MCs 中 p70S6K-PI3K 介导的反馈回路导致 Raf1-MEK1/2-ERK 通路激活。本研究表明，大剂量雷帕霉素通过 p70S6K/PI3K/MAPK 反馈环在大鼠 MCs 中导致 ERK1/2 激活，从而降低雷帕霉素对 MC 增殖的抑制作用。我们的结果表明，1000 nmol/L 雷帕霉素通过 MCs 中 p70S6K-PI3K 介导的反馈回路导致 Raf1-MEK1/2-ERK 通路激活。本研究表明，大剂量雷帕霉素通过 p70S6K/PI3K/MAPK 反馈环在大鼠 MCs 中导致 ERK1/2 激活，从而降低雷帕霉素对 MC 增殖的抑制作用。