Mechanism of oxidative stress-AMPK-Cx43-NLRP3 pathway regulating extracellular matrix remodeling of rat gastric smooth muscle cells under high glucose condition
AIM:To investigate the regulatory effect of oxidative stress-AMP-activated protein kinase(AMPK)-connexin 43(Cx43)-nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3)pathway on extracellular matrix(ECM)remodeling of gastric smooth muscle cells under high glucose(HG)condition.METHODS:Primary rat smooth muscle cells cultured in vitro were divided into normal glucose group,HG group,HG+NLRP3 inhibitor MCC950(15 nmol/L)group,HG+Cx43 hemichannel blocker GAP19(100 μmol/L)group,HG+AMPK inhibitor Compound C(CC;10 μmol/L)group,and high glucose+antioxidant α-lipoic acid(α-LA;100 μmol/L)group,which were all cultured for 48 h for detection.The protein levels of caspase-1,matrix metalloproteinase-2(MMP-2),NLRP3,Cx43,transforming growth factor-β1(TGF-β1),TGF-β3,tissue inhibitor of metalloproteinase-1(TIMP-1),purinergic P2X7 receptor(P2X7R)and phosphorylated AMPK(p-AMPK)in the cells were detected by Western blot.The levels of adenosine tri-phosphate,interleukin-1β,collagen type Ⅰ(Col I)and collagen type Ⅲ(Col Ⅲ)in the cell culture medium were deter-mined by ELISA.RESULTS:Compared with HG group,the levels of MMP-2 and TGF-β3 in HG+MCC950 group were decreased(P<0.01),while the levels of TGF-β1 and TIMP-1 were increased(P<0.01).The levels of NLRP3 and cas-pase-1 in HG+GAP19 group were decreased(P<0.01),while the expression of P2X7R was not changed.The levels of NLRP3,caspase-1,P2X7R,total Cx43 and membrane Cx43,and the ratio of membrane Cx43 to cytoplasmic Cx43 were decreased in HG+CC group(P<0.01).The levels of p-AMPK,caspase-1,NLRP3,P2X7R,total Cx43 and membrane Cx43,and the ratio of membrane Cx43 to cytoplasmic Cx43 were decreased in HG+α-LA group(P<0.05).CONCLU-SION:High glucose regulates the content of Col I and Col Ⅲ through the oxidative stress-AMPK-Cx43-NLRP3 pathway,thereby participating in the extracellular matrix remodeling of gastric smooth muscle cells.
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维普
