Effects And Mechanism Of Dickkopf-related Protein 1 On The Urinary Sodium Excretion In Spontaneously Hypertensive Rats

Objective:To explore the effect and mechanism of DKK1 on urinary sodium excretion and blood pressure in spontaneously hypertensive rats.Method:1.The role of Wnt-?-catenin signaling in blood pressure regulation has been recognized,however,DKK1 which is Wnt-?-catenin signaling classic inhibitor in hypertension is still unclear.In the present study,109 hypertension patients and 30 normotensive subjects were recruited.The circulating bone-derived factors including Dickkopf-related protein 1(DKK1),osteoprotegerin(OPG),osteocalcin(OC),osteopontin(OPN),osteonectin and osteocrin were measured by a protein liquid-chip assay.2.Effect and mechanism of DKK1on urinary sodium excretion.2.1.We observe expression of DKK1 in Wistar Kyoto rats(WKY)and spontaneously hypertensive rats(SHR),as well as renal proximal tubule epithelial(RPT),by western blot and immunohistochemical.2.2.12 WKY rats were randomly divided into WKY+DKK1 group and WKY control group(n=6).12 SHR rats were randomly into SHR+DKK1 group and SHR group(n=6);WKY+DKK1 group and SHR+DKK1 group were pumped with the recombinant human protein DKK1,blood pressure of the rats were measured by non-invasive tail-cuff method.And the urine was collected by metabolic cages for 24-hour urinary volume and urinary sodium,correcting with weight.Observing 24-hour urinary volume,urinary sodium and blood pressure,determine whether DKK1 has an effect on sodium urine and blood pressure.2.3.The clarify the effect of DKK1 on Na~+-K~+-ATPase.Recombinant DKK1 protein was added to WKY-RPT and SHR-RPT cells,or using siDKK1 interfered the expression of DKK1 to observe Na~+-K~+-ATPase activity.3.To clarify that DKK1 regulated the activity of Na~+-K~+-ATPase through Wnt/?-catenin pathway.3.1.Recombinant DKK1 protein and the inhibitor of GSK3?(BIO)was added into RPT cells,to test Na~+-K~+-ATPase activity.3.2.Using siDKK1 interfered RPT cells and added?-catenin nuclear transcription inhibitor(ICTR 14),to observed the activity of Na~+-K~+-ATPase.Result:1.The data revealed that serum DKK1,osteonectin and osteocrin concentrations derived from hypertensive participants were considerably lower than those of controls.Hypertension was independently associated with both DKK1 and osteonectin levels after adjustment for age,BMI,FBG,HbA1c,HDL-C and eGFR.Decreased DKK1 and osteonectin levels in circulation were observed in hypertension patients,which are associated with risk of hypertension.2.DKK1 can reduce Na~+-K~+-ATPase activity and increase urinary sodium excretion.2.1.Study showed that the expression level of DKK1 in WKY rats'kidney and WKY-RPT cell was higher than that in SHR.2.2.Exogenous DKK1 protein increased urinary sodium excretion and decreased blood pressure in SHR,but had no effect on WKY.2.3.The activity of Na~+-K~+-ATPase in group WKY+DKK1 and group SHR+DKK1decreased after adding recombinant DKK1 protein,compared with group WKY and group SHR.After transfection of RPT cells with siDKK1,the activity of Na~+-K~+-ATPase was detected.It was found that the activity of Na~+-K~+-ATPase in WKY+siDKK1 and SHR+siDKK1was higher than that in group WKY and SHR..3.DKK1 regulated Na~+-K~+-ATPase activity dependence of?-catenin.3.1.GSK3?inhibitor(BIO)reverses the decreasing of Na~+-K~+-ATPase activity induced by DKK1 protein.3.2.Inhibitor of?-catenin nuclear transcription(ICRT 14)reversed increasing of Na~+-K~+-ATPase activity induced by siDKK1.Conclusion:Low serum DKK1 is associated with hypertension.And in spontaneously hypertensive rats,DKK1 regulates the activity of Na~+-K~+-ATPase in a Wnt/?-catenin manner to affect urinary sodium excretion,which regulates blood pressure.