Department of Pharmacology, Experimental Therapy & Toxicology, Eberhard-Karls-University of Tübingen, Tübingen, Germany.
Institute of Preventive Veterinary Medicine, Key Laboratory of Animal Disease and Human Health of Sichuan Province, Sichuan Agricultural University, Chengdu, China.
J Mol Med (Berl). 2021 Mar;99(3):373-382. doi: 10.1007/s00109-020-02016-4. Epub 2021 Jan 7.
Vascular calcification may result from stimulation of osteogenic signalling with upregulation of the transcription factors CBFA1, MSX2 and SOX9, as well as alkaline phosphatase (ALPL), which degrades and thus inactivates the calcification inhibitor pyrophosphate. Osteogenic signalling further involves upregulation of the Ca-channel ORAI1. The channel is activated by STIM1 and then accomplishes store-operated Caentry. ORAI1 and STIM1 are upregulated by the serum & glucocorticoid inducible kinase 1 (SGK1) which is critically important for osteogenic signalling. Stimulators of vascular calcification include vasopressin. The present study explored whether exposure of human aortic smooth muscle cells (HAoSMCs) to vasopressin upregulates ORAI1 and/or STIM1 expression, store-operated Ca entry and osteogenic signalling. To this end, HAoSMCs were exposed to vasopressin (100 nM, 24 h) without or with additional exposure to ORAI1 blocker MRS1845 (10 μM) or SGK1 inhibitor GSK-650394 (1 μM). Transcript levels were measured using q-RT-PCR, cytosolic Ca-concentration ([Ca]) by Fura-2-fluorescence, and store-operated Ca entry from increase of [Ca] following re-addition of extracellular Ca after store depletion with thapsigargin (1 μM). As a result, vasopressin enhanced the transcript levels of ORAI1 and STIM1, store-operated Ca entry, as well as the transcript levels of CBFA1, MSX2, SOX9 and ALPL. The effect of vasopressin on store-operated Ca entry as well as on transcript levels of CBFA1, MSX2, SOX9 and ALPL was virtually abrogated by MRS1845 and GSK-650394. In conclusion, vasopressin stimulates expression of ORAI1/STIM1, thus augmenting store-operated Caentry and osteogenic signalling. In HAoSMCs, vasopressin (VP) upregulates Ca channel ORAI1 and its activator STIM1. VP upregulates store-operated Caentry (SOCE) and osteogenic signalling (OS). VP-induced SOCE, OS and Ca-deposition are disrupted by ORAI1 inhibitor MRS1845. VP-induced SOCE, OS and Ca-deposition are disrupted by SGK1 blocker GSK-650394. KEY MESSAGES: • In HAoSMCs, vasopressin (VP) upregulates Ca channel ORAI1 and its activator STIM1. • VP upregulates store-operated Caentry (SOCE) and osteogenic signalling (OS). • VP-induced SOCE, OS and Ca-deposition are disrupted by ORAI1 inhibitor MRS1845. • VP-induced SOCE, OS and Ca-deposition are disrupted by SGK1 blocker GSK-650394.
血管钙化可能是由于成骨信号的刺激导致的，其转录因子 CBFA1、MSX2 和 SOX9 的上调，以及碱性磷酸酶（ALPL）的上调，ALPL 可降解并使钙化抑制剂焦磷酸失活。成骨信号还涉及钙通道 ORAI1 的上调。该通道被 STIM1 激活，然后完成储存操作的钙内流。ORAI1 和 STIM1 被血清和糖皮质激素诱导激酶 1（SGK1）上调，SGK1 对成骨信号至关重要。血管钙化的刺激物包括血管加压素。本研究探讨了血管加压素是否会使人类主动脉平滑肌细胞（HAoSMCs）上调 ORAI1 和/或 STIM1 表达、储存操作的钙内流和成骨信号。为此，HAoSMCs 暴露于血管加压素（100 nM，24 小时）而不暴露于 ORAI1 阻滞剂 MRS1845（10 μM）或 SGK1 抑制剂 GSK-650394（1 μM）。使用 q-RT-PCR 测量转录水平，使用 Fura-2 荧光测量细胞质 Ca 浓度([Ca])，并通过在储存耗尽后用他普西坦（1 μM）重新添加细胞外 Ca 来测量储存操作的 Ca 内流来测量[Ca]的增加。结果表明，血管加压素增强了 ORAI1 和 STIM1、储存操作的 Ca 内流的转录水平，以及 CBFA1、MSX2、SOX9 和 ALPL 的转录水平。MRS1845 和 GSK-650394 几乎消除了血管加压素对储存操作的 Ca 内流以及 CBFA1、MSX2、SOX9 和 ALPL 转录水平的影响。总之，血管加压素刺激 ORAI1/STIM1 的表达，从而增强储存操作的 Ca 内流和成骨信号。在 HAoSMCs 中，血管加压素（VP）上调钙通道 ORAI1 和其激活剂 STIM1。VP 上调储存操作的 Ca 内流（SOCE）和成骨信号（OS）。VP 诱导的 SOCE、OS 和 Ca 沉积被 ORAI1 抑制剂 MRS1845 破坏。VP 诱导的 SOCE、OS 和 Ca 沉积被 SGK1 阻断剂 GSK-650394 破坏。关键信息：• 在 HAoSMCs 中，血管加压素（VP）上调钙通道 ORAI1 和其激活剂 STIM1。• VP 上调储存操作的 Ca 内流（SOCE）和成骨信号（OS）。• ORAI1 抑制剂 MRS1845 破坏了 VP 诱导的 SOCE、OS 和 Ca 沉积。• SGK1 阻断剂 GSK-650394 破坏了 VP 诱导的 SOCE、OS 和 Ca 沉积。
