Fibronectin Type ? Domain Containing 5 Inhibits Phenotypic Transformation And Inflammasome Activation Of Adventitial Fibroblasts In SHR

BackgroundHypertension is a cardiovascular disease endangering the human health seriously.Vascular remodeling plays an important role in the occurrence and development of hypertension.Vascular adventitial fibroblasts?AFs?is the major component of the adventitia.If the blood vessels are injured,AFs preserves phenotype alterations and differentiates into myofibroblasts?MFs?.MFs has stronger ability of proliferation and migration,leading to neointimal formation and vascular remodeling.Nucleotide-binding oligomerization domain-like receptors family pyrin domain containing 3?NLRP3?inflammasome is a macromolecule protein complex composed of NLRP3,apoptosis-associated speck-like protein containing a caspase recruitment domain?ASC?and procaspase-1.The levels of interleukin-1??IL-1??and interleukin-18?IL-18?in plasma were significantly higher in patients with hypertension than in normal people.The levels of inflammatory factors are highly correlated with blood pressure.NLRP3 inflammasome is involved in the pathogenesis of hypertension.Fibronectin type ? domain containing 5?FNDC5?is a I type membrane protein participating in the regulation of aerobic exercise mediated metabolic changes.Our previous studies showed that FNDC5 overexpression could improve the obese mice hyperlipidemia and promote fat cell lipolysis.However,its role in cardiovascular diseases is still unknown.Our study focuses on the effects of FNDC5 in phenotypic transformation and NLRP3 inflammation activation and and their downstream signal pathway in AFs from SHR.Objective1.To determine the effects of FNDC5 overexpression on phenotype transformation,NLRP3 inflammation activation and its molecular mechanism in AFs from aortic adventitia of SHR;2.To determine the therapeutic effect of FNDC5 overexpression on adventitial inflammation and vascular remodeling in SHR.MethodsPrimary AFs were isolated from thoracic aorta of WKY and SHR by enzyme digestion.After the rats were anesthetized,the thoracic aorta was removed and placed in sterile PBS.Simply,thoracic aorta was isolated,and perivascular adipose tissues of the aorta were removed.The aorta was cut open and stripped of the intima and media.The adventitia was treated with 0.2%Type 1A collagenase in PBS for digestion for15min.Cells were re-suspended in DMEM with 10%fetal bovine serum and 1%penicillin-streptomycin maintained at 37?in a humidified atmosphere containing5%CO₂.WKY and SHR rats were randomly divided into two groups,six rats in each group.The two groups of rats were injected with empty lentivirus virus or FNDC5overexpressing lentivirus virus?1.0×10⁸ plaque-forming units?via tail vein respectively.After three weeks,the thoracic aorta was removed for detection.Immunohistochemical staining was used for detecting the expression of FNDC5in aortic adventitia of rats.ROS production in aortic adventitia and AFs was evaluated with dihydroethidium?DHE?staining.Masson staining was used for detecting rat aortic adventitia fibrosis and vascular remodeling.FNDC5,NOX2,NOX4,collagen-I,collagen-III,fibronectin,?-SMA,NLRP3,ASC,caspase-1,IL-1?,CTGF,TGF-?1,EGFP and GAPDH in AFs and aortic adventitia were determined with Western blot analysis.FNDC5,NOX2,NOX4,collagen-I,collagen-III,fibronectin,?-SMA,NLRP3,ASC,caspase-1,IL-1?,CTGF,TGF-?₁,EGFP and GAPDH were analyzed by real-time quantitative PCR.Results1 In aortic adventitia of SHR,FNDC5 mRNA and protein levels were downregulated,while NLRP3 inflammasome was activated.2 FNDC5 overexpression inhibited the phenotypic transformation,fibrosis,NLRP3 inflammasome activation and NOX2-derived ROS production in AFs of SHR,but had no significant effect on NOX4.3 FNDC5 knockdown promoted phenotype transformation,fibrosis,inflammation and oxidative stress in AFs of SHR,but had no significant effect on NOX4.4 FNDC5 overexpression inhibited NOX2-derived ROS production,NLRP3 inflammasome activation,phenotype transformation and fibrosis in aortic adventitia of SHR.5 FNDC5 overexpression reduced SBP and alleviated vascular remodeling in SHR.Conclusions1.FNDC5 inhibited NOX2-derived ROS production,NLRP3 inflammasome activation,phenotypic transformation and fibrosis in AFs of SHR.2.FNDC5 overexpression alleviated oxidative stress,inflammation and vascular remodeling in aortic adventitia of SHR.