Division of Nephrology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, Taiwan.
Division of Nephrology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, Taiwan.
Life Sci. 2020 Nov 1;260:118280. doi: 10.1016/j.lfs.2020.118280. Epub 2020 Aug 12.
Vascular calcification is a common complication in patients with chronic kidney disease and associated with increased morbidity and mortality. The role of TRPM7 in vascular smooth muscle cell (VSMC) transformation during vascular calcification is not clear. We aim to investigate the effects of phosphate and indoxyl sulphate on the expression of TRPM7 and calcification-related molecules in VSMC.
Human aortic smooth muscle cells (HASMC) were treated with phosphate (3.3 mM) or indoxyl sulphate (500 μM and 1000 μM). 2-APB, a channel blocker of TRPM7 was added simultaneously in blocking experiment. Cells were then examined grossly and alizarin red solution was employed for calcification assessment. Lastly, cells were harvested for gene expression and protein abundance analysis.
Phosphate treatment induced significant increase in BMP2, RUNX2, BMP7, vitamin D receptor (VDR), calcium sensing receptor (CaSR) and TRPM7, but 1-alpha hydroxylase, klotho, DKK1 and sclerostin were not changed. The addition of 2-APB prevented increase of BMP2, RUNX2, BMP7, VDR, CaSR and TRPM7. Indoxyl sulphate treatment was associated with decrease in TRPM7 and DKK1, but increase in RUNX2, BMP2 and VDR were noted. There were no significant alterations in BMP7, CaSR, klotho,1-alpha hydroxylase and sclerostin. Co-treatment with 2-APB reversed the increase in VDR.
Both phosphate and indoxyl sulphate induced calcification in VSMC but it was more prominent in phosphate. TRPM7 was upregulated by phosphate but downregulated in indoxyl sulphate treatment. Vascular calcification was reduced by blocking TRPM7 with 2-APB and there was partial anti-calcification effect in indoxyl sulphate.
血管钙化是慢性肾脏病患者的常见并发症，与发病率和死亡率的增加有关。TRPM7 在血管平滑肌细胞（VSMC）血管钙化过程中的转化中的作用尚不清楚。我们旨在研究磷酸盐和吲哚硫酸对 VSMC 中 TRPM7 表达和钙化相关分子的影响。
用磷酸盐（3.3mM）或吲哚硫酸（500μM 和 1000μM）处理人主动脉平滑肌细胞（HASMC）。在阻断实验中同时添加 2-APB，一种 TRPM7 的通道阻断剂。然后对细胞进行大体检查，并使用茜素红溶液进行钙化评估。最后，收集细胞进行基因表达和蛋白丰度分析。
磷酸盐处理诱导 BMP2、RUNX2、BMP7、维生素 D 受体（VDR）、钙敏感受体（CaSR）和 TRPM7 显著增加，但 1-α羟化酶、klotho、DKK1 和 Sclerostin 没有变化。添加 2-APB 可防止 BMP2、RUNX2、BMP7、VDR、CaSR 和 TRPM7 的增加。吲哚硫酸处理与 TRPM7 和 DKK1 的减少有关，但 RUNX2、BMP2 和 VDR 的增加。BMP7、CaSR、klotho、1-α羟化酶和 Sclerostin 没有明显变化。用 2-APB 共同处理可逆转 VDR 的增加。
磷酸盐和吲哚硫酸均可诱导 VSMC 钙化，但磷酸盐诱导作用更明显。磷酸盐上调 TRPM7，但吲哚硫酸处理下调 TRPM7。用 2-APB 阻断 TRPM7 可减少血管钙化，吲哚硫酸处理具有部分抗钙化作用。
