Pharmacological data on Combination of Astragaloside Ⅳ with Tanshinone ⅡA inhibits the Angiotensin II-stimulated proliferation and migration of vascular smooth muscle cells via the STIM1, TRPC1/6, and NFATc3 signaling pathway

Published: 21-07-2019| Version 4 | DOI: 10.17632/m5f5tzdwjv.4
Contributor:
Xiaoxia Qi

Description

Objective: Astragaloside Ⅳ and Tanshinone ⅡA combined application is widely used to clinically treat coronary heart disease and atherosclerosis in China. However, it is not clear if a combined application is better than a single application of either drug alone. This study aimed to evaluate the role of TRPC/NFATc-mediated calcium signaling in the angiotensin II-induced migration and proliferation of vascular smooth muscle cells and the inhibitory effect of astragaloside Ⅳ combined with tanshinone ⅡA that occurs via the STIM1, TRPC1/ 6, and NFAT3 signaling pathway. Materials and methods: Primary VSMCs were obtained from the abdominal aortas of SD rats, cultured by tissue block adherence and identified by immunocytochemistry. VSMC migration and proliferation were investigated by transwell migration and MTT assays, respectively. Intracellular Fluo-3/AM [Ca2+]i fluorescence was detected using a laser confocal microscope. The protein expression of STIM1, TRPC1, TRPC6, calcineurin and NFATc3 was determined using a Western blot. Results: Astragaloside Ⅳ combined with tanshinone ⅡA could significantly inhibit angiotensin II-induced VSMC proliferation, migration and [Ca2+]i increases, and this combination was superior to either drug alone and was not significantly different from the TRPC blocker SKF96365. Western blotting revealed that Ang II induced obvious increases in VSMC STIM1, TRPC1, TRPC6, CaN and NFATc3 protein expression and that astragaloside Ⅳ combined with tanshinone ⅡA and SKF96365 significantly inhibited the STIM1, TRPC1, TRPC6, CaN and NFATc3 protein expression induced by angiotensin II, indicating that astragaloside Ⅳ combined with tanshinone ⅡA has the same blocking effect on TRPC-NFATc calcium signaling as SKF96365 and is better than the application of a single drug alone. Conclusions: Astragaloside Ⅳ combined with tanshinone ⅡA can inhibit the proliferation and migration of angiotensin II-induced VSMCs and is superior to the single application of either astragaloside Ⅳ or tanshinone ⅡA, and the mechanism is related to the inhibition of the STIM1, TRPC1/6, CaN, and NFATc3 calcium signaling pathway.

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Primary VSMCs were obtained from the abdominal aortas of SD rats, cultured by tissue block adherence and identified by immunocytochemistry. VSMC migration and proliferation were investigated by transwell migration and MTT assays, respectively. Intracellular Fluo-3/AM [Ca2+]i fluorescence was detected using a laser confocal microscope. The mRNA and protein expression was determined using qPCR and Western blot, respectively. The statistical comparisons among the groups were performed with one-way analysis of variance (ANOVA) followed by a least significant difference (LSD) test.