MicroRNA‐335‐5p suppresses lower extremity deep venous thrombosis by targeted inhibition of PAI‐1 via the TLR4 signalingpathway

This study aims to investigate the effects of microRNA‐335‐5p (miR‐335‐5p) on lower‐extremity deep vein thrombosis (LEDVT) by targeting PAI‐1 through the TLR4 signaling pathway in rat models. siRNA, mimic, and inhibitor were used for transfection. The miR‐335‐5p expression was detected by in situ hybridization. CCK‐8 assay and flow cytometry were adopted to detect proliferation, cell cycle, and apoptosis, respectively. Scratch test and Matrigel‐based tube formation assay were used to detect the effect of miR‐335‐5p on cell migration ability and tube formation ability. A miR‐335‐5p lentivirus plasmid was constructed and injected into LEDVT rats. The length and weight of thrombus were measured, changes of thrombus recanalization were observed by CD34 immunohistochemistry, and levels of PAI‐1 and inflammatory factors in femoral vein blood were detected by ELISA. LEDVT rats showed a higher AOD value of PAI‐1, higher expression of PAI‐1, NF‐κB, Rac1, IL‐1β, and TLR4 and a lower miR‐335‐5p expression. PAI‐1 and miR‐335‐5p were negatively correlated. Compared to the blank and siRNA‐NC groups, the miR‐335‐5p mimic and siRNA‐PAI‐1 groups showed declined expression of PAI‐1, TLR4, NF‐κB, Rac1, and IL‐1β, increased proliferation and tube formation abilities, less cells in G0/G1 phase, and decreased apoptosis, decreased length and weight of thrombus, organized thrombus, increased new blood vessels, and decreased levels of PAI‐1, IL‐1, IL‐6, and Tnf‐a. miR‐335‐5p may suppress the occurrence and development of LEDVT in rats by repressing the activation of the TLR4 signaling pathway by targeted inhibition of PAI‐1. miR‐335‐5p may suppress the occurrence and development of LEDVT in rats by repressing the activation of the TLR4 signaling pathway by targeted inhibition of PAI‐1.