Effect of Sodium-Glucose Co-Transporter 2 Inhibitor, Dapagliflozin, on Renal Renin-Angiotensin System in an Animal Model of Type 2 Diabetes

Seok Joon Shin, Sungjin Chung, Soo Jung Kim, Eun-Mi Lee, Young-Hye Yoo, Ji-Won Kim, Yu-Bae Ahn, Eun-Sook Kim, Sung-Dae Moon, Myung-Jun Kim, Seung-Hyun Ko, Seok Joon Shin, Sungjin Chung, Soo Jung Kim, Eun-Mi Lee, Young-Hye Yoo, Ji-Won Kim, Yu-Bae Ahn, Eun-Sook Kim, Sung-Dae Moon, Myung-Jun Kim, Seung-Hyun Ko

Abstract

Background: Renal renin-angiotensin system (RAS) activation is one of the important pathogenic mechanisms in the development of diabetic nephropathy in type 2 diabetes. The aim of this study was to investigate the effects of a sodium-glucose co-transporter 2 (SGLT-2) inhibitor, dapagliflozin, on renal RAS in an animal model with type 2 diabetes.

Methods: Dapagliflozin (1.0 mg/kg, OL-DA) or voglibose (0.6 mg/kg, OL-VO, diabetic control) (n = 10 each) was administered to Otsuka Long-Evans Tokushima Fatty (OLETF) rats for 12 weeks. We used voglibose, an alpha-glucosidase inhibitor, as a comparable counterpart to SGLT2 inhibitor because of its postprandial glucose-lowering effect without proven renoprotective effects. Control Long-Evans Tokushima Otsuka (LT) and OLETF (OL-C) rats received saline (n = 10, each). Changes in blood glucose, urine albumin, creatinine clearance, and oxidative stress were measured. Inflammatory cell infiltration, mesangial widening, and interstitial fibrosis in the kidney were evaluated by histological analysis. The effects of dapagliflozin on renal expression of the RAS components were evaluated by quantitative RT-PCR in renal tissue.

Results: After treatment, hyperglycemia and urine microalbumin levels were attenuated in both OL-DA and OL-VO rather than in the OL-C group (P < 0.05). The urine angiotensin II (Ang II) and angiotensinogen levels were significantly decreased following treatment with dapagliflozin or voglibose, but suppression of urine Ang II level was more prominent in the OL-DA than the OL-VO group (P < 0.05). The expressions of angiotensin type 1 receptor and tissue oxidative stress markers were markedly increased in OL-C rats, which were reversed by dapagliflozin or voglibose (P < 0.05, both). Inflammatory cell infiltration, mesangial widening, interstitial fibrosis, and total collagen content were significantly increased in OL-C rats, which were attenuated in OL-DA group (P < 0.05).

Conclusion: Dapagliflozin treatment showed beneficial effects on diabetic nephropathy, which might be via suppression of renal RAS component expression, oxidative stress and interstitial fibrosis in OLETF rats. We suggest that, in addition to control of hyperglycemia, partial suppression of renal RAS with an SGLT2 inhibitor would be a promising strategy for the prevention of treatment of diabetic nephropathy.

Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Fig 1. Changes in body weight during…
Fig 1. Changes in body weight during the experimental period in LETO (LT) and OLETF rats with saline (OL-C), dapagliflozine (OL-DA) or voglibose (OL-VO) treatment.
*P < 0.05, OL-C vs. other groups; †P < 0.05, LT vs. other groups; ‡P < 0.05, OL-DA vs. OL-VO. Values are expressed as means ± SE.
Fig 2. Intraperitoneal glucose tolerance test at…
Fig 2. Intraperitoneal glucose tolerance test at 12 weeks in LETO (LT) and OLETF rats with saline (OL-C), dapagliflozin (OL-DA) or voglibose (OL-VO) treatment.
*P < 0.05, OL-C vs. other groups; †P < 0.05, LT vs. other groups. Values are expressed as means ± SE.
Fig 3. Systemic and intrarenal RAS activation…
Fig 3. Systemic and intrarenal RAS activation at 12 weeks.
Plasma renin activity (A), serum aldosterone (B), urinary angiotensin II (C) and angiotensinogen (D) in LETO (LT) and OLETF rats with saline (OL-C), dapagliflozin (OL-DA) or voglibose (OL-VO) treatment. *P < 0.05, OL-C vs. other groups; †P < 0.05, LT vs. other groups; ‡P < 0.05, OL-DA vs. OL-VO. Values are expressed as means ± SE.
Fig 4. The expressions of RAS and…
Fig 4. The expressions of RAS and antioxidant enzymes in renal tissues.
(A) Representative immunoblots of AT1R, Cu/ZnSOD, MnSOD and catalase. Quantitative analyses of the expression of AT1R (B), Cu/ZnSOD (C), MnSOD (D), catalase (E) in LETO (LT) and OLETF rats with saline (OL-C), dapagliflozine (OL-DA) or voglibose (OL-VO) treatment. *P < 0.05, OL-C vs. other groups; †P < 0.05, LT vs. other groups; ‡P < 0.05, OL-DA vs. OL-VO. Values are expressed as means ± SE. AT1R, Ang II type I receptor
Fig 5. Oxidative stress markers in renal…
Fig 5. Oxidative stress markers in renal tissues.
The levels of renal H2O2 (A) and malondialdehyde (MDA) (B), 8-OHdG immunostaining (x 200, C), quantitative measurement of 8-OHdG staining (D) in LETO (LT) and OLETF rats with saline (OL-C), dapagliflozin (OL-DA) or voglibose (OL-VO) treatment. *P < 0.05, OL-C vs. other groups; †P < 0.05, LT vs. other groups; ‡P < 0.05, OL-DA vs. OL-VO. Values are expressed as means ± SE. 8-OHdG, 8-hydroxy-2-deoxy guanosine.
Fig 6. Changes in glomerular or tubulointerstitial…
Fig 6. Changes in glomerular or tubulointerstitial phenotypes in LETO or OLETF rats.
Representative sections for assessing the inflammatory cell infiltration (immunostaining for ED-1, x 200), glomerular expansion (PAS, x 400) and tubulointerstitial fibrosis (Masson trichrome, x 200) in LETO (LT) and OLETF rats with saline (OL-C), dapagliflozine (OL-DA) or voglibose (OL-VO) treatment.
Fig 7
Fig 7
(A) Semi-quantitative analysis of the number of ED-1 positive cells, (B) fractional mesangial area, and (C) fibrotic area in kidney tissue. *P < 0.05, OL-C vs. other groups; †P < 0.05, LT vs. other groups; ‡P < 0.05, OL-DA vs. OL-VO. Values are expressed as means ± SE.
Fig 8. The expression of type IV…
Fig 8. The expression of type IV collagen and total collagen content in renal tissues.
(A) Representative immunoblot of type IV collagen, (B) quantitative analyses of the expression of type IV collagen and (C) the amount of hydroxyproline in renal tissue in LETO (LT) and OLETF rats with saline (OL-C), dapagliflozin (OL-DA) or voglibose (OL-VO) treatment. *P < 0.05, OL-C vs. other groups; †P < 0.05, LT vs. other groups; ‡P < 0.05, OL-DA vs. OL-VO; §P < 0.05, OL-C vs. OL-DA. Values are expressed as means ± SE.

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