Effect of tissue factor pathway inhibitor on the pyroptosis of vascular smooth muscle cells induced by angiotensin II.

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In recent years, a mass of studies have shown that pyroptosis plays an important role in the proliferation of vascular smooth muscle cells (VSMCs). We investigated whether angiotensin II (Ang II) induces the pyroptosis of rat aortic VSMCs and the role of NOD-like receptor family pyrin domain containing 3 (NLRP3) in this process. Additionally, we explored the effect and related mechanism of recombinant tissue factor pathway inhibitor (rTFPI) in Ang II-induced VSMC pyroptosis. Cultured VSMCs were divided into five groups: control group, Ang II group (1×10-5 mol/L), MCC950 group (NLRP3 inhibitor, 15 nmol/L), Ang II + MCC950 group and Ang II + rTFPI (50 µg/L) group. Cell viability was measured by cell counting kit-8 (CCK8) assays and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assays. Propidium iodide (PI) staining and immunofluorescence were performed to determine the pyroptosis of VSMCs. Changes in VSMC ultrastructure were evaluated through transmission electron microscopy. The expression levels of NLRP3, pro-caspase-1, gasdermin D-N (GSDMD-N), and interleukin-1β (IL-1β) were determined by western blot analysis. The cell viability, the positive rate of PI staining, and the expression level of GSDMD detected by immunofluorescence in the Ang II group were higher than that in the control group, whereas they all decreased in Ang II + MCC950 group and Ang II + rTFPI group compared with Ang II group (P<0.05). Electron microscopy analysis revealed less extracellular matrix, increased myofilaments, and decreased endoplasmic reticulum, Golgi complex, and mitochondria in Ang II + rTFPI-treated VSMCs than in Ang II-treated VSMCs. The protein expression levels of the pyroptosis-related molecules NLRP3, pro-caspase-1, GSDMD-N, and IL-1β in Ang II group showed an increasing trend compared with those in control group (P<0.05); however, these expression levels in Ang II + MCC950 and Ang II + rTFPI groups were significantly lower than those in Ang II group (P<0.05). Ang II may induce pyroptosis in VSMCs by activating NLRP3. rTFPI can inhibit Ang II-induced VSMC pyroptosis. Furthermore, rTFPI might exert this effect by inhibiting the NLRP3 pathway and therefore play an important role in the treatment of vascular remodeling induced by hypertension.