An Adenovirus Vector-Mediated RNA Interference Efficiently Inhibits Rat Stim1 Expression in Vascular Smooth Muscle Cells and Rat Carotid Artery

Objective: RNA interference (RNAi) is a widely used mechanism to knock down target gene expression. Successful application of RNAi requires efficient delivery of siRNA into cells. This study demonstrates an efficient and specific knockdown of stim1, a protein essential for store-operated calcium (SOC) influx and cell proliferation, in vascular smooth muscle cells (VSMCs) and rat carotid artery, highlighting a promising application of adenovirus systems in vascular physiological studies. Method: Two target sequences for stim1 were chemically synthesized, and recombinant adenoviruses for RNAi-mediated stim1 knockdown were produced. Rat aortic VSMCs were primarily cultured and transfected with adenoviral vectors. Angioplasty of the rat left carotid artery was performed using a balloon embolectomy catheter followed by transfection with adenoviral vectors. Stim1 protein levels were measured by western blot. Main results: An adenovirus vector for stim1 knockdown was successfully constructed. At 1, 3, and 5 days after infection with Ad-si/stim1 at an MOI of 10 pfu/cell, stim1 protein levels significantly decreased compared to the Ad-empty group. Infection with Ad-si/stim1 virus (3 days post-infection at 10 pfu/cell MOI) resulted in a robust decrease in SOC. Ad-mediated siRNA effectively infected the rat carotid artery and significantly inhibited stim1 expression. Conclusion: This study establishes a model system for in vivo and in vitro studies of stim1 function, demonstrating that siRNA expression and specific gene knockdown can be achieved using adenoviral vectors.