TRPC6 and TRPC4 Heteromultimerization Mediates Store Depletion-Activated NCX1 Reversal in Proliferative Vascular Smooth Muscle Cells

Store depletion has been shown to induce Ca²⁺ entry by Na+/Ca+ exchange (NCX) 1 reversal in proliferative vascular smooth muscle cells (VSMCs). The study objective was to investigate the role of transient receptor potential canonical (TRPC) channels in store depletion and NCX1 reversal in proliferative VSMCs. In cultured VSMCs, expressing TRPC1, TRPC4, and TRPC6, the removal of extracellular Na⁺ was followed by a significant increase of cytosolic Ca²⁺ concentration that was inhibited by KBR, a selective NCX1 inhibitor. TRPC1 knockdown significantly suppressed store-operated, channel-mediated Ca²⁺ entry, but TRPC4 knockdown and TRPC6 knockdown had no effect. Separate knockdown of TRPC1, TRPC4, or TRPC6 did not have a significant effect on thapsigargin-initiated Na⁺ increase in the peripheral regions with KBR treatment, but knockdown of both TRPC4 and TRPC6 did. Stromal interaction molecule (STIM)1 knockdown significantly reduced TRPC4 and TRPC6 binding. The results demonstrated that TRPC4–TRPC6 heteromultimerization linked Ca²⁺ store depletion and STIM1 accumulation with NCX reversal in proliferative VSMCs.

已有研究证实，在增殖性血管平滑肌细胞（vascular smooth muscle cells, VSMCs）中，钙库耗竭可通过钠/钙交换体1（Na+/Ca+ exchange, NCX1）的反向转运诱导钙离子内流。本研究旨在探究瞬时受体电位经典型（transient receptor potential canonical, TRPC）通道在增殖性VSMCs的钙库耗竭及NCX1反向转运过程中的作用。在体外培养的VSMCs中，当细胞表达TRPC1、TRPC4与TRPC6时，去除细胞外钠离子后，胞质钙离子浓度显著升高，该效应可被选择性NCX1抑制剂KBR所阻断。敲低TRPC1可显著抑制钙库操纵性通道介导的钙离子内流，但敲低TRPC4或TRPC6则无明显影响。单独敲低TRPC1、TRPC4或TRPC6，均未对毒胡萝卜素（thapsigargin）诱导的、经KBR处理的外周区域钠离子升高产生显著作用，但同时敲低TRPC4与TRPC6则可产生显著抑制效果。基质相互作用分子1（stromal interaction molecule, STIM1）的敲低可显著降低TRPC4与TRPC6之间的结合。本研究结果表明，在增殖性VSMCs中，TRPC4与TRPC6的异源多聚体化将钙库耗竭、STIM1聚集与NCX1反向转运三者关联起来。