Department of endocrinology, Taihe Hospital of traditional Chinese Medicine, No. 59 Tuanjie West Road, Taihe County, Fuyang, 236600, Anhui, China.
J Bioenerg Biomembr. 2024 Jun;56(3):273-284. doi: 10.1007/s10863-024-10001-6. Epub 2024 Mar 1.
Tetramethylpyrazine (TMP) is one of the active ingredients of Chuan Xiong that has been reported to have effects on numerous diseases, including diabetic nephropathy (DN). Whereas, related molecular mechanisms are not fully elucidated. We aimed to explore circACTR2's role in TMP-mediated protective effects on DN. In vitro DN condition was established in human kidney cells (HK-2) by treating high glucose (HG). CCK-8 assay and flow cytometry assay were used to observe cell viability and survival. Oxidative stress was determined by the associated markers using kits. The release of inflammatory factors was detected using ELISA kits. Quantitative real-time PCR (qPCR) and western blot were utilized for expression analysis of cricACTR2, miR-140-5p, and GLI pathogenesis-related 2 (GLIPR2). The binding between miR-140-5p and circACTR2 or GLIPR2 was confirmed by dual-luciferase, RIP, and pull-down studies. HG largely induced HK-2 cell apoptosis, oxidative stress, and inflammation, which were alleviated by TMP. CircACTR2's expression was enhanced in HG-treated HK-2 cells but attenuated in HG + TMP-treated HK-2 cells. CircACTR2 overexpression attenuated the functional effects of TMP and thus restored HG-induced cell apoptosis, oxidative stress, and inflammation. CircACTR2 bound to miR-140-5p to enhance the expression of GLIPR2. MiR-140-5p restoration or GLIPR2 inhibition reversed the role of circACTR2 overexpression. CircACTR2 attenuated the protective effects of TMP on HG-induced HK-2 cell damages by regulating the miR-140-5p/GLIPR2 network, indicating that circACTR2 was involved in the functional network of TMP in DN.
川芎嗪（TMP）是川芎的一种有效成分，已被报道对许多疾病具有作用，包括糖尿病肾病（DN）。然而，相关的分子机制尚未完全阐明。我们旨在探讨 circACTR2 在 TMP 介导的对 DN 的保护作用中的作用。通过用高葡萄糖（HG）处理，在人肾细胞（HK-2）中建立体外 DN 条件。使用 CCK-8 测定法和流式细胞术测定法观察细胞活力和存活。使用试剂盒测定相关标记物以确定氧化应激。使用 ELISA 试剂盒检测炎症因子的释放。定量实时 PCR（qPCR）和 Western blot 用于分析 circACTR2、miR-140-5p 和 GLI 发病相关蛋白 2（GLIPR2）的表达。通过双荧光素酶、RIP 和下拉研究证实了 miR-140-5p 与 circACTR2 或 GLIPR2 的结合。HG 极大地诱导了 HK-2 细胞凋亡、氧化应激和炎症，而 TMP 减轻了这些作用。HG 处理的 HK-2 细胞中 circACTR2 的表达增强，而 HG+TMP 处理的 HK-2 细胞中 circACTR2 的表达减弱。circACTR2 的过表达减弱了 TMP 的功能作用，从而恢复了 HG 诱导的细胞凋亡、氧化应激和炎症。circACTR2 与 miR-140-5p 结合以增强 GLIPR2 的表达。miR-140-5p 的恢复或 GLIPR2 的抑制逆转了 circACTR2 过表达的作用。circACTR2 通过调节 miR-140-5p/GLIPR2 网络减弱了 TMP 对 HG 诱导的 HK-2 细胞损伤的保护作用，表明 circACTR2 参与了 TMP 在 DN 中的功能网络。
