Effects Of Na+/H+ Exchanger NHE 1 On The Proliferation Of Vascular Smooth Muscle Cells And The Ca²⁺ Regulation Induced By High Salt

Objective: The aim of this study was to observe the expression of NHE1 in the vascular smooth muscle cell（VSMC） proliferation induced by high salt,and to determine the mechanism of NHE1 mediated intracellular regulation of p H value and Ca²⁺ in VSMC.Methods: Rat vascular smooth muscle cells were cultured by tissue block method. The MTT assays were employed to determine action concentration of NHE1 inhibitor（amiloride）. The experiment was divided into NS group（Na + 139mmol/L）, HS group（Na+159mmol/L）, HS+amiloride group（Na+ 159mmol/ L +amiloride 10Μm, 20μM, 30μM and 40μM）. After synchronization with free-serum medium for 24 h, VSMC were divided into NS group（Na+139mmol/L）,NS+amiloride group（Na+139mmol/L+amiloride 30μM）, HS group（Na+159mmol/L）, HS+amiloride group（Na+ 159mmol/ L +amiloride 30μM）,then continued to culture for 72 h.The activity of Ca²⁺-ATPase was determined by spectrophotography. The m RNA expression of NHE1, NCX-1, PMCA4 were measured by real time-PCR. Immunofluorescence and western blot assay were further performed to detect the location and protein level of NHE1, NCX-1, PMCA4. The fluorescence ratio assay was applied to detect intracellular p H（p Hi）value and Ca²⁺ concentration in VSMC.Results:The result of MTT showed that the proliferative tendency of VSMC induced by high-salt inhibited by amiloride in a dose-dependent manner. Compared with NS group, the m RNA and protein expression of NHE1 and NCX-1 increased, the protein level of PMCA4 decreased, and the intracellular calcium concentration and p Hi increased significantly（P<0.05）.The m RNA and protein expression of NHE1, NCX-1 was down-regulated in HS +amiloride group（P<0.05）, and the intracellular calcium concentration and p Hi were significantly decreased（P<0.05） compared to HS group. There was no significant difference in the protein and m RNA expression of PMCA4（P>0.05）.Conclusion:I. In cultured VSMCs, high-salt promoted cellular proliferation, and the increase was attenuated by amiloride in a dose-dependent manner.II. High salt increases the expression of NHE1. Amiloride inhibited NHE1 expression, thus attenuated high salt-accelerated NCX-1 expression, intracellular free calcium and p H value.