Sacubitril/Valsartan Improves Progression of Early Diabetic Nephropathy in Rats Through Inhibition of NLRP3 Inflammasome Pathway

Abstract

Purpose: Diabetic nephropathy (DN), a global disease, is the leading cause of end-stage renal disease. There is a lack of specific treatment for this disease, and early intervention in disease progression is essential. In this paper, we used a rat model of early diabetic nephropathy to explore the therapeutic mechanism of sacubitril/valsartan in rats with early diabetic nephropathy.

Materials and methods: Rats were grouped into 1 normal group; 2. Model group (DN group): STZ (45 mg/kg/d) induced early diabetic nephropathy rats; 3. Sac group: DN rats + Sac group (orally, 60 mg/kg/d) for 6 weeks. After 6 weeks, the levels of serum albumin (ALB), glucose (GLU), creatinine (Cr), urea nitrogen (BUN) and 24-h urinary protein excretion were measured. In renal tissue homogenates, NLRP3 inflammasome, proinflammatory factors IL1-β and TNF-α, oxidative stress MDA and pro-fibrotic cytokine TGF-β1 were performed. Histological analysis of kidneys by hematoxylin and eosin (HE), PAS and Masson trichrome staining.

Results: 1. Sacubitril/valsartan (Sac) significantly improved renal hypertrophy, proteinuria and serum albumin levels in rats with early diabetic nephropathy (P < 0.001), and decreased GLU, Scr (P<0.001), and BUN levels (P < 0.01).2. Light microscopy of renal tissues showed glomerular hypertrophy and interstitial inflammatory cell infiltration, and mean glomerular area (MGA) and mean glomerular volume (MGV) were crucially increased in early diabetic nephropathy (P < 0.001), and the Sac group showed reduced renal pathology and improved MGA and MGV (P < 0.001).3. Kidney tissue homogenate levels of NLRP3, Caspase-1, IL1-β, TNF-α, MDA and TGF-β1 were critically, increased in DN rats (P < 0.001), and SOD was significantly decreased. All these indicators above were improved after treatment (P < 0.0001).

Conclusion: Nlrp3-inflammasome promote progression of diabetic nephropathy through inflammation, fibrosis and oxidative stress; sacubitril/valsartan ameliorated early diabetes-induced renal damage by inhibiting NLRP3 pathway activation.

Keywords: Nlrp3-inflammasome; diabetic nephropathy; fibrosis; inflammation; oxidative stress; sacubitril/valsartan.

Figure 1

Comparison of general conditions and biochemical indices in three groups of rats. Sac can improve renal hypertrophy, proteinuria, serum albumin, glucose metabolism, Scr and DUN levels in early DN rats. (A) Body weights of rats in the three groups.(B) Kidney weights of the three groups of rats. (C) Ratio of kidney weight/body weight in the three groups of rats. (D) Glucose of the three groups of rats. (E) Urine protein levels in the three groups of rats. (F) Albumin of the three groups of rats. (G) Creatinine levels in the three groups of rats. (H) Urea nitrogen levels in the three groups of rats. Data are expressed as mean ± standard deviation, *P<0.05; **P<0.01; ***P<0.001; ns: P>0.05.

Figure 2

Comparison of renal pathological features in three groups of rats. (A) Sac ameliorates glomerular hypertrophy and interstitial inflammatory cell infiltration in early diabetic nephropathy. (B) Mean glomerular area of rats in the three groups. (C) Mean glomerular volume of the three groups of rats. Data are expressed as mean ± standard deviation; ***P<0.001; ns: P>0.05.

Figure 3

Comparison of the expression levels of inflammation, oxidative stress and fibrosis in the kidney tissues of three groups of rats. Sac down-regulates TNF-a, MDA and TGF-β1 expression levels through inhibition of the NLRP3 inflammasome pathway, thus exerting to improve the inflammatory response, oxidative stress and fibrotic response in the diabetic kidney. (A) NLRP3 of rats in the three groups.(B) Caspase-1 of the three groups of rats. (C) IL-1β of the three groups of rats. (D) TNF-α of the three groups of rats. (E) MDA of the three groups of rats. (F) SOD of the three groups of rats. (G) TGF-β1 of the three groups of rats. Data are expressed as mean ± standard deviation, *P<0.05; ***P<0.001; ns: P>0.05.