ABSTRACT
AIM:To observe the effect of angiotensin-converting enzyme 2(ACE2)deletion on vasoconstric-tion reactivity of aortic segments in ACE2 knockout(KO)mice with tourniquet shock(TS).METHODS:The 8-month-old male mice with C57BL/6 background were divided into wild-type(WT)control group,WT-TS group,KO group and KO-TS group,with 10 mice in each group,of which five were used for determination of vascular reactivity,and the other five for the other assays.The hindlimbs of the mice in WT-TS group and KO-TS group were ligated with tourniquet for 2 h and loosened for 4 h.The mice in WT group and KO group were subjected to the same treatment except for tourniquet liga-tion.The vasoconstriction reactivity of the aorta was measured on tensiometer.The morphological damage of the aorta was evaluated by vascular histopathology.Western blot was used to detect the expression of AT1,MAS,ACE and ACE2 pro-teins in aorta.The serum levels of angiotensin(Ang)Ⅱ and Ang-(1-7)were determined by enzyme-linked immunosorbent assay.RESULTS:Compared with WT group,the mice in WT-TS group had lower vascular reactivity to norepinephrine(NE)and obvious vascular lesions.The expression of ACE protein increased significantly(P＜0.01),while the expres-sion of ACE2 decreased(P＜0.05).The expression of AT1 protein in aorta decreased significantly,the expression of MAS protein increased significantly,and the AT1/MAS ratio decreased(P＜0.01).Serum Ang II level increased,serum Ang-(1-7)level decreased,and Ang Ⅱ/Ang-(1-7)ratio increased(P＜0.05).Compared with WT group,vascular reactivity in KO group increased at low concentration of NE(＜10-7 mol/L),and decreased at high concentration(＞10-7 mol/L)without vascular lesion.The expression levels of aortic AT1,MAS and ACE were all elevated(P＜0.05).The serum level of Ang Ⅱ increased(P＜0.05),but the level of Ang-(1-7)had no obvious change.Compared with KO and WT-TS groups,the aortic reactivity in KO-TS group subtracted apparently(P＜0.05),representing its curve shifting to the right obviously.The morphological damage aggravated slightly,and the expression of AT1 and ACE increased slightly in KO-TS group com-pared with WT-TS group(P＜0.05).However,the expression of MAS increased significantly in vascular tissue(P＜0.01).The serum levels of Ang Ⅱ and Ang-(1-7)further increased and decreased,respectively,and the Ang Ⅱ/Ang-(1-7)ratio increased(P＜0.01).CONCLUSION:Deficiency of ACE2 induces severe aortic hyporeactivity to NE during TS,which may be related to the increased imbalance of renin-angiotensin system in ACE2 gene knockout mice.
ABSTRACT
AIM:To observe the cyclic adenosine monophosphate(cAMP)transfer across myoendothelial gap junctions(MEGJ)in the regulatory effect of angiopoietin-2(Ang2)on hyporeactivity after hypoxia in vascular smooth mus-cle cells(VSMCs ).METHODS:The double-sided cell co-culture model of vascular endothelial cells(VECs )and VSMCs was set up.The protein expression of inducible nitric oxide synthase(iNOS)was determined by Western blot.The contraction of VSMCs was detected via the leakage of FITC-labeled bovine serum albumin.Alexa Fluor 488-cAMP was used as the tracer to observe the cAMP transfer across MEGJ from VECs to VSMCs.RESULTS:In cultured VECs and VSMCs alone ,the cAMP concentrations were both significantly increased after exogenous Ang 2 treatment and hypoxia ,and more in VECs than that in VSMCs(P<0.05).In the double-sided cell co-culture model,the difference was weakened,and the increase in cAMP concentration in VSMCs after exogenous Ang 2 treatment and hypoxia was antagonized by connexin 43(Cx43)small interfering RNA(siRNA)(P<0.05).Alexa Fluor 488-cAMP in VECs transfered into VSMCs after exoge-nous Ang2 treatment and hypoxia,which was also antagonized by Cx43 siRNA(P<0.05).The cAMP antagonist inhibited the protein expression of iNOS in the VSMCs and the hyporeactivity of the VSMCs after exogenous Ang 2 treatment and hy-poxia(P<0.05).CONCLUSION:Ang2 may regulate the protein expression of iNOS in VSMCs and the hyporeactivity of VSMCs after hypoxia through the cAMP transfer across MEGJ.
ABSTRACT
Objective To investigate the relationship of the beneficial effect of CsA on vascular reactivity to Rho-kinase,protein kinase C ( PKC) and mitochondrial permeability transition pore ( MPTP) in traumatic hemorrhagic shock rats. Methods With traumatic hemor-rhagic shock rats and hypoxia-treated vascular smooth muscle cells ( VSMCs) ,the role of Rho-kinase and PKC in CsA-regulating vascular re-activity following shock and their relationship to MPTP was observed. Meanwhile,the effects of CsA on inflammatory mediators including TNF-α, IL-1β and IL-6 were also studied. Results CsA significantly improved the vascular reactivity of superior mesenteric artery following hemor-rhagic shock. Rho-kinase inhibitor Y27632 significantly antagonized CsA-induced increase of vascular reactivity,while PKC inhibitor stauros-porine had no significant influences on the effects of CsA. Further studies showed that CsA and Rho-kinase agonist U46619 inhibited the o-pening of MPTP in hypoxia-treated VSMCs. In addition,shock induced a significant increase of TNF-αand IL-1β,but CsA did not show a sig-nificant inhibitory effect on their level. Conclusion CsA-induced restoration of vascular reactivity following traumatic hemorrhagic shock via inhibiting MPTP opening,and Rho-kinase participate in this process.