Xiang Tianya, Wang Xijian, Huang Shujiao, Zhou Kexin, Fei Shengnan, Zhou Bing, Yue Kun, Li Qingxin, Xue Shengnan, Dai Yongyi, Zhang Jing, Ni Haoran, Sun Cheng, Huang Xinzhong
Department of Nephrology, Affiliated Hospital of Nantong University, 20 Xisi Road, Nantong, Jiangsu, 226001, China; Medical School of Nantong University, Nantong, 226001, China.
Xinglin College, Nantong University, Nantong, 226001, China.
Phytomedicine. 2025 Jan;136:156324. doi: 10.1016/j.phymed.2024.156324. Epub 2024 Dec 15.
Macrophage-myofibroblast transition (MMT) plays a significant role in the progression of renal fibrosis in chronic kidney disease (CKD), making inhibition of MMT a promising therapeutic strategy. Pyruvate kinase M2 (PKM2) and its metabolite lactate are implicated in the pathogenesis of renal fibrosis; however, the mechanisms through which they contribute to this process remain poorly understood.
To investigate the effects of PKM2 inhibition by shikonin on renal fibrosis and the underly mechanisms.
Mice were subjected to unilateral ureteral obstruction (UUO) to establish a CKD model. Renal fibrosis was assessed using histochemistry and western blotting. The MMT and histone lactylation levels were evaluated by immunofluorescence and western blotting. The interaction between the Tgfb1 promoter and lactylated histone H3 (K18) was examined using chromatin Immunoprecipitation (ChIP).
PKM2 expression was significantly elevated in the renal tubular cells of UUO mouse kidneys, resulting in increased pyruvate and lactate production. Similarly, lactate levels were elevated in TGF-β1-treated TCMK-1 cells and in the serum of CKD patients. In UUO mice, treatment with shikonin, a potent PKM2 inhibitor, effectively reduced lactate production, alleviated renal fibrosis, decreased TGF-β1 expression, and suppressed the MMT process. Mechanistic studies revealed that lactate treatment stimulates Tgfb1 expression in TCMK-1 cells. Consequently, TGF-β1 in conditioned media from lactate-treated TCMK-1 cells promoted M2 macrophage polarization and upregulated fibrotic gene expression in RAW264.7 cells. Pharmacological intervention demonstrated that TGF-β1 activates the Smad3 pathway to drive the MMT process. In TCMK-1 cells, both lactate treatment and PKM2 overexpression induced Tgfb1 expression by promoting histone H3K18 lactylation.
Our findings indicate that PKM2-induced excessive lactate production renal tubular cells contributes to renal fibrosis. Lactate promotes histone lactylation, leading to TGF-β1 expression in these cells, which subsequently activates the Smad3 pathway in macrophages, driving the MMT and fibrosis in the kidney. Therefore, targeting PKM2, as with shikonin treatment, may represent an effective therapeutic strategy for managing renal fibrosis in CKD.
巨噬细胞-肌成纤维细胞转变（MMT）在慢性肾脏病（CKD）肾纤维化进展中起重要作用，抑制MMT成为一种有前景的治疗策略。丙酮酸激酶M2（PKM2）及其代谢产物乳酸与肾纤维化发病机制有关；然而，它们促成这一过程的机制仍知之甚少。
研究紫草素抑制PKM2对肾纤维化的影响及其潜在机制。
对小鼠进行单侧输尿管梗阻（UUO）以建立CKD模型。使用组织化学和蛋白质印迹法评估肾纤维化。通过免疫荧光和蛋白质印迹法评估MMT和组蛋白乳酸化水平。使用染色质免疫沉淀（ChIP）检测Tgfb1启动子与乳酸化组蛋白H3（K18）之间的相互作用。
PKM2在UUO小鼠肾脏肾小管细胞中的表达显著升高，导致丙酮酸和乳酸生成增加。同样，在TGF-β1处理的TCMK-1细胞和CKD患者血清中乳酸水平也升高。在UUO小鼠中，用强效PKM2抑制剂紫草素治疗可有效降低乳酸生成，减轻肾纤维化，降低TGF-β1表达，并抑制MMT过程。机制研究表明，乳酸处理可刺激TCMK-1细胞中Tgfb1表达。因此，来自乳酸处理的TCMK-1细胞条件培养基中的TGF-β1促进RAW264.7细胞中M2巨噬细胞极化并上调纤维化基因表达。药理学干预表明，TGF-β1激活Smad3信号通路以驱动MMT过程。在TCMK-1细胞中，乳酸处理和PKM2过表达均通过促进组蛋白H3K18乳酸化诱导Tgfb1表达。
我们的研究结果表明，PKM2诱导肾小管细胞中过量乳酸生成促成肾纤维化。乳酸促进组蛋白乳酸化，导致这些细胞中TGF-β1表达，随后激活巨噬细胞中的Smad3信号通路，驱动肾脏中的MMT和纤维化。因此，如紫草素治疗那样靶向PKM2可能是管理CKD肾纤维化的有效治疗策略。
