基于肾脏SIRT1/自噬通路失衡在衰老中的作用探讨“肾藏精”的科学内涵

It has been reported that chronic kidney disease is an important factor to induce premature senility of the body and down-regulation of SIRT1 in kidney can induce the aggregation of aging-related proteins in multiple organs. Thus we hypothesise that: SIRT1/autophagy pathway in kidney, closely associated with aging, is the material basis of the anti-aging function of ”kidney storing essence”. To test this hypothesis, we first observe patients with Diabetic Kidney Disease (DKD), a typical aging-related disease, to investigate the relationship between the abnormality of SIRT1/autophagy pathway in kidney and the premature senility of the body, as well as the symptom score of "deficiency of kidney-YIN, deficiency of kidney-YANG, and deficiency of kidney-QI, collateral stasis and obstruction". Then the animal model of podocyte conditional knockout of SIRT1 will be used to confirm whether the treatment of "nourishing kidney yin, nourishing kidney yang and tonifying kidney activating collateral" can restore SIRT1/autophagy pathway to retard kidney and body premature aging from aging or diabetes. Finally, the cells with gene knockdown or drug intervention to disturb SIRT1/autophagy pathway will be used to find the specific nodes of anti-premature aging by containing serum or freeze-dried powder of above treatment. We thus can clear the inner mechanism of anti-aging effect laid in the treatment. The aim of this study is to explore the scientific connotation of "kidney essence". And we also try to explore SIRT1/autophagy pathway related to renal senescence is an important material basis of the aforementioned Chinese medicine theory.

前期研究发现：慢性肾脏病可导致机体早衰，肾脏SIRT1表达下降可诱发衰老相关蛋白在多器官聚集。基于此，我们提出与衰老关系密切的肾脏SIRT1/自噬通路是“肾藏精”抗衰老的重要物质基础的假说。为验证该假说，本研究首先以典型衰老相关疾病糖尿病肾病患者为研究对象，探讨肾脏SIRT1/自噬通路异常与机体早衰、以及与“肾阴亏虚、肾阳虚损、肾气不足伴络脉瘀阻”症状积分的关系；并借助足细胞条件性敲除SIRT1的动物模型，探讨“滋肾阴、益肾阳、补肾通络”法是否能够通过纠正SIRT1/自噬通路失衡延缓增龄所致的衰老以及糖尿病诱发的肾脏及机体早衰；最后采用化学药物及基因干预的肾脏固有细胞模型，筛查上述诸法含药血清或冻干粉对抗细胞早衰所干预的具体节点，从而明确其延缓衰老的内在机制。本研究旨在对“肾藏精”抗衰老的科学内涵进行有益探索，冀以阐明肾脏衰老相关SIRT1/自噬通路是上述传统中医理论的重要物质基础。

前期研究发现：慢性肾脏病可导致机体早衰，肾脏SIRT1表达下降可诱发衰老相关蛋白在多器官聚集。基于此，我们提出与衰老关系密切的肾脏SIRT1/自噬通路是“肾藏精”抗衰老的重要物质基础的假说。为验证该假说，本研究首先以典型衰老相关疾病糖尿病肾病患者为研究对象，探讨肾脏SIRT1/自噬通路异常与机体早衰、以及与“肾阴亏虚、肾阳虚损、肾气不足伴络脉瘀阻”症状积分的关系。基础研究方面，DN大鼠肾脏抗衰老蛋白SIRT1的检测结果显示，模型组大鼠肾脏SIRT1的表达水平较对照组明显降低，“滋肾阴、益肾阳、补肾通络”法可上调SIRT1表达。对自噬通路的检测发现，“滋肾阴、益肾阳、补肾通络”法的LC3-II、p62较模型组明显下降，提示上调SIRT1可以疏通自噬通路。体外培养的足细胞也进一步证实“滋肾阴、益肾阳、补肾通络”法治疗可以上调糖尿病状态下SIRT1的表达。同时发现，“滋肾阴、益肾阳、补肾通络”法也可以疏通足细胞中的自噬通路，减少LC3-II、p62堆积，中药治疗得到类似的作用结果，但是使用EX527抑制SIRT1后，中药对自噬通路的改善作用受到抑制，更加证明了“滋肾阴、益肾阳、补肾通络”法很可能通过上调SIRT1表达疏通自噬通路。另外，与模型组相比，“滋肾阴、益肾阳、补肾通络”法可以减弱SA-β-gal活性、降低p53和p21水平，改善细胞骨架结构，提高podocin、nephrin表达水平，中药治疗组也表现出相似的作用结果，但是这种改变在加入SIRT1抑制剂EX527后受到抑制，进一步佐证金蝉益肾通络方很可能通过上调SIRT1，发挥对抗DN足细胞衰老及损伤的作用。研究表明，“滋肾阴、益肾阳、补肾通络”法很可能通过上调SIRT1表达疏通DN肾脏细胞自噬通路，对抗糖尿病肾病引起的肾脏细胞衰老、减轻肾脏损伤，进而保护肾脏功能、延缓糖尿病肾病进展。上述研究结果已发表SCI论文6篇，中文核心期刊论文6篇。相关成果得到国内外同行认可，部分研究数据为支撑获中华中医药学会科学技术奖二等奖1项。