Hydroxyfasudiattenuates Lipopolysaccharide Induced Endothelial Inflammation Via Suppressing The Expression Of Rho-associated Coiled-coil Protein Kinase 1,Connexin43 And Caveolin1

Objective:Toinvestigatetheeffectandmechanismofhydroxyfasudi(HF),aspecific Rho kinase inhibitor,on lipopolysaccharide(LPS)induced endothelial inflammatory reaction.Methods:A total of 24 male Sprague Dawley ratswereintocontrolgroup(n=12),HF group(n=12),LPS group(n=12)and LPS + HF group(n=12)with randomly divided with random number table method.There was no special treatment in control group.HF(30mg/kg)wasinjectedintraperitoneallyin HFgroup.LPS(1mg/kg)wereinjectedintravenouslyin LPS group.In LPS+HF group,HF(30 mg/kg)was injected intraperitoneally,followed by intravenous LPSinjection(1mg/kg)30minuteslater..24ratsweresacrificedafter8 hours,and aortictissuewasextracted..RT-PCRwasperformedtodetectmRNAlevelsof Rho-associated coiled-coil protein kinase(ROCK)1,connexin(Cx)43 and caveolin(Cav)1.Theprotein levers of ROCK1,Cx43 and Cav-1 were assessed by Western blot and immunohistochemicalstainingrespectively.Results:(1)RT-PCRexperimentsshowedthatmRNAlevelsof ROCK1(2.67±0.03),Cx43(1.73±0.03)and Cav1(1.85±0.04)in LPS group weresignificantlyhigher thanmRNAlevelsof ROCK1(1.0±0.04),Cx43(1.00±0.08)and Cav1(1.0±0.03)incontrolgroupandmRNAlevelsof ROCK1(0.77±0.04),Cx43(0.91±0.01)and Cav1(0.82±0.03)in HF guoup.(all P<0.05).The mRNAlevelsof ROCK1(0.38±0.02),Cx43(0.58±0.02),and Cav1(0.27±0.01)in LPS + HF group were significantly lower than in LPS group(all P<0.05).(2)Western blot analysis showedthat proteinlevelsof ROCK1(3.46±0.82),Cx43(0.33±0.09)and Cav1(3.45±0.74)in LPS group were significantly higher than protein levels of ROCK1(2.19±0.56),Cx43(0.21±0.09)and Cav1(2.25±0.91)in controlgroupand proteinlevelsof ROCK1(1.57±0.38),Cx43(0.18±0.07)and Cav1(2.06±0.40)in HF group(all P<0.05).Proteinlevelsof ROCK1(1.09±0.52),Cx43(0.11±0.06),and Cav1(2.06±0.40)in LPS + HF group weresignificantlylower thanin LPSgroup(all P<0.05).(3)Immunohistochemical staining showed that protein levels of ROCK1(84.1±0.9),Cx43(99.1±2.1),and Cav1(167.0±6.4)in LPS group were significantlyhigherthan proteinlevelsof ROCK1(53.7±2.9),Cx43(46.2±0.8),and Cav1(84.9±1.0)incontrolgroup and proteinlevelsof ROCK1(40.1±0.9),Cx43(35.1±0.6),and Cav1(74.4±0.5)in HF group(all P<0.05).Proteinlevelsof ROCK1(30.4±0.6),Cx43(21.4±1.3),and Cav1(55.8±2.8)in LPS + HF group weresignificantlylower thanin LPSgroup(all P<0.05).Conclusion: HF attenuates LPS induced endothelial inflammatory reaction probably via suppressing the expression of ROCK1,Cx43 and Cav1.