Downregulation of Renal G Protein–Coupled Receptor Kinase Type 4 Expression via Ultrasound‐Targeted Microbubble Destruction Lowers Blood Pressure in Spontaneously Hypertensive Rats

Abstract

BackgroundG protein–coupled receptor kinase type 4 (GRK4) plays a vital role in the long‐term control of blood pressure (BP) and sodium excretion by regulating renal G protein–coupled receptor phosphorylation, including dopamine type 1 receptor (D1R). Ultrasound‐targeted microbubble destruction (UTMD) is a promising method for gene delivery. Whether this method can deliver GRK4 small interfering RNA (siRNA) and lower BP is not known.Methods and ResultsBP, 24‐hour sodium excretion, and urine volume were measured after UTMD‐targeted GRK4 siRNA delivery to the kidney in spontaneously hypertensive rats. The expression levels of GRK4 and D1R were determined by immunoblotting. The phosphorylation of D1R was investigated using immunoprecipitation. The present study revealed that UTMD‐mediated renal GRK4 siRNA delivery efficiently reduced GRK4 expression and lowered BP in spontaneously hypertensive rats, accompanied by increased sodium excretion. The increased sodium excretion might be accounted for by the UTMD regulation of D1R phosphorylation and function in spontaneously hypertensive rats. Further analysis showed that, although UTMD had no effect on D1R expression, it reduced D1R phosphorylation in spontaneously hypertensive rats kidneys and consequently increased D1R‐mediated natriuresis and diuresis.ConclusionsTaken together, these study results indicate that UTMD‐targeted GRK4 siRNA delivery to the kidney effectively reduces D1R phosphorylation by inhibiting renal GRK4 expression, improving D1R‐mediated natriuresis and diuresis, and lowering BP, which may provide a promising novel strategy for gene therapy for hypertension.

Role of GRK4 in the regulation of the renal ETB receptor in hypertension

“…28 The persistent phosphorylation of GPCRs leads to their uncoupling from their G protein/effector complex and impairment of their function, D 1 R and D 3 R, for example. 25,28,[36][37][38][39] In the present study, we found that renal GRK4 expression was, indeed, increased in SHRs, which was consistent with previous studies. 25,36 Therefore, we speculated that increased GRK4 expression and/or activity may have contributed to the increased renal ETBR phosphorylation in the SHR and hGRK4γ142V transgenic mice.…”

“…As aforementioned, ETBR belongs to the family of GPCRs; their phosphorylation is a common mechanism that desensitizes their function. 25,27,31 Our present study found that ETBR phosphorylation is higher in kidneys from SHRs than WKY rats, which may explain the dysfunction of renal ETBR in hypertension. 17 However, this supposition needs to be confirmed, that is, decreasing the hyper-phosphorylation of ETBR, per se, should improve ETBR function and ameliorate the hypertension.…”

“…20 Poly-L-lysine (PLL) and the electrostatic adsorption method were used to prepare microbubbles carrying GRK4 siRNA, as previously described. 24,25 In brief, a PLL solution (1 mg/ mL) was prepared with sterile double-distilled water and mixed with equal volume of blank microbubbles followed by incubation at 4°C for 30 minutes. Phosphate-buffered saline (PBS) was used to wash the mixture twice to remove any unbound PLL.…”

“…Ultrasound-targeted microbubble destruction (UTMD) of GRK4 in the kidney of SHRs was performed following the method we have previously reported. 25 In brief,…”

Ultrasound Combined with Microbubbles Enhances the Effects of Methylprednisolone in Lipopolysaccharide-Induced Human Mesangial Cells

“…However, repeated and prolonged steroid therapy is associated with an excess of adverse events, some of which are dose related (Zhao et al, 2015a). Previous studies confirmed that US combined with MBs could enhance the delivery of genes or drug to target areas of tissues including the kidney (Ibsen et al, 2013;Aryal et al, 2014;Huang et al, 2016;Wei et al, 2016). A recent study showed that CoQ10-liposome in combination with US and MBs could improve the protective effect of the drug on the damaged kidney as seen in diabetic nephropathy (Yue et al, 2017).…”