Diabetes Metab J. 2025;49(6):1229-1241.

Anti-Senescence Effect of Inhibiting Sodium-Glucose Cotransporter 2 and α-Glucosidase in a Type 2 Diabetes Mellitus Animal Model.

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제1저자 홍세린, 공병수, 이현석 · 교신저자 조영민

PMID 40403762 PMC12620679 DOI 10.4093/dmj.2024.0339 IF 8.2 내분비대사내과

소속 (3)

Department of Internal Medicine, Seoul National University College of Medicine, Seoul, Korea.
Department of Translational Medicine, Seoul National University College of Medicine, Seoul, Korea.
Genomic Medicine Institute, Medical Research Center, Seoul National University College of Medicine, Seoul, Korea.

Abstract

BACKGRUOUND The prevalence of type 2 diabetes mellitus (T2DM) increases with age, and cellular senescence of pancreatic β-cells plays a key role in T2DM pathogenesis. As canagliflozin and acarbose have been shown to increase lifespan in mice, we investigated the effect of sodium-glucose cotransporter 2 (SGLT2) inhibitor, α-glucosidase inhibitor or both on the cellular senescence of β-cells in a T2DM mouse model.

METHODS Enavogliflozin (0.3 mg/kg), acarbose (10 mg/kg), or vehicle was orally administered daily to db/db mice for 6 weeks. The levels of senescence markers (p16, p21, and p53) in the pancreas and kidney were measured through real-time polymerase chain reaction (PCR), immunofluorescence staining, and Western blot. In an in vitro analysis, isolated pancreatic islets were exposed to H2O2 to induce cellular senescence, then treated with β-hydroxybutyrate (β-HB), and subsequently assessed for levels of senescent markers.

RESULTS Enavogliflozin alone or combined with acarbose effectively lowered blood glucose levels in db/db mice. The combined treatment resulted in the greatest increase in β-cell function calculated using insulinogenic index and homeostasis model assessment of β-cell function compared to the vehicle. Additionally, the combined treatment significantly reversed the increase in p16, with a similar trend observed in p21 and p53 in the islets. Treatment increased circulating β-HB and in vitro analysis suggested the activation of nuclear factor erythroid 2-related factor 2 (Nrf2) by β-HB in reducing senescence in the islets.

CONCLUSION The combined administration of enavogliflozin and acarbose significantly reduced blood glucose, improved β-cell function, and reduced senescent β-cells in db/db mice. This combination therapy holds potential as a senotherapeutic strategy for managing T2DM.

Keywords

AcarboseCellular senescenceDiabetes mellitus, type 2Sodium-glucose transporter 2 inhibitors

Figures

**Fig. 1.** The effect of enavogliflozin and/or acarbose on body weight, blood glucose, cumulative food intake, oral glucose tolerance test and plasma insulin levels. (A) Body weight, (B) blood glucose, (C) cumulative food intake of mice in db/m+ (n=10), *db/db*+ vehicle (*db/db*+V), *db/db*+enavogliflozin (*db/db*+S), *db/db*+acarbose (*db/db*+A) (n=10), and *db/db*+S+A (n=10) groups during 6 weeks of treatment. (D) Blood glucose concentrations and (E) the corresponding area under the curve (AUC) during the oral glucose tolerance test after 4 weeks of treatment (n=10 per group). (F) Plasma insulin levels during oral glucose tolerance test. Measured at 0 and 15 minutes after administration of glucose (n=10–11 per group). ELISA, enzyme-linked immunosorbent assay. ^aP<0.01 vs. *db/db*+V, ^bP<0.0001 vs. db/m+ (one-way analysis of variance [ANOVA]).

**Fig. 2.** The effect of enavogliflozin and/or acarbose on insulinogenic index, homeostasis model assessment of β-cell function (HOMA-β), homeostasis model assessment of insulin resistance (HOMA-IR), and the relative insulin-positive β-cell area of the pancreas sections. (A) Insulinogenic index in an oral glucose tolerance test (OGTT) (n=7–8 per group). (B) HOMA-β% indexes. (C) HOMA-IR (n=5 per group). (D) Relative insulin-positive β-cell area of immunofluorescence staining in the pancreas sections. Scale bars=500 μm. (E) Insulin-positive β-cell area. SGLT2i, sodium-glucose cotransporter 2 inhibitor; AGI, α-glucosidase inhibitor. ^aP<0.05 *db/db*+enavogliflozin (*db/db*+S) vs. *db/db*+enavogliflozin+acarbose (*db/db*+S+A) (t-test), ^bP<0.05, ^cP<0.01, ^dP<0.001 vs. *db/db*+vehicle (*db/db*+V) (analysis of variance [ANOVA]), ^eP<0.05 vs. db/m+, ^fP<0.01 vs. db/m+ (ANOVA).

**Fig. 3.** Relative mRNA expression of *p16, p21*, and *p53* in pancreatic islets and the immunofluorescence staining of insulin, p16 and p21 in the pancreas sections by treatment groups. (A) Relative mRNA expression of *p16, p21*, and *p53* in pancreatic islet cells. (B) Immunofluorescence staining of insulin (green) and p16 (red). Scale bars=150 μm. (C) Immunofluorescence staining of insulin (green), p21 (red) in the pancreas sections. Scale bars=150 μm. (D) Immunofluorescence staining of insulin (green), γ H2A. X variant histone (γH2AX) (red) in the pancreas sections. Scale bars=150 μm. SGLT2i, sodium-glucose cotransporter 2 inhibitor; AGI, α-glucosidase inhibitor; S, enavogliflozin; A, acarbose. ^aP<0.05, ^bP<0.01 vs. *db/db*+vehicle (analysis of variance [ANOVA]).

**Fig. 4.** Plasma β-hydroxybutyrate (β-HB) concentrations by treatment groups, the relative mRNA expression of *p16, p21*, and *p53* in pancreatic islets and the protein level of nuclear factor erythroid 2-related factor 2 (NRF2) in the pancreatic islets after treatment with H₂O₂ followed by β-HB. (A) Plasma β-HB concentrations (n=9–10 per group) (analysis of variance [ANOVA]). (B) Relative expression of *p16, p21*, and *p53* in the H₂O₂ treated pancreatic islet cells. (C) Western blot and band intensity for NRF2 in the pancreatic islet cells. (D) Western blot analysis of p53BP1, p16, p21, phosphor-adenosine monophosphate-activated protein kinase (p-AMPK), and NRF2 in whole pancreas tissue. V, vehicle; S, enavogliflozin; A, acarbose; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; SGLT2i, sodium-glucose cotransporter 2 inhibitor; AGI, α-glucosidase inhibitor; HSP90, heat shock protein 90. ^aP<0.05, ^bP<0.01, ^cP<0.001 (ANOVA).

**Fig. 5.** The effect of enavogliflozin and/or acarbose on mesangial expansion, glomerulus size and cellular senescence of the kidney. (A) Representative images of periodic acid-Schiff (PAS)-stained section of the kidney. (B) The mesangial area (scale bars=50 μm) and (C) glomerular area after treatment. (D) Western blot for p53BP1, p16, phosphor-adenosine monophosphate-activated protein kinase (p-AMPK), and nuclear factor erythroid 2-related factor 2 (NRF2) in the kidney. (E) Immunohistochemical staining of γ H2A.X variant histone (γH2AX) in the kidney. Scale bars=50 μm. S, enavogliflozin; A, acarbose; SGLT2i, sodium-glucose cotransporter 2 inhibitor; AGI, α-glucosidase inhibitor; HSP90, heat shock protein 90. ^aP<0.05 vs. *db/db*+vehicle, ^bP<0.01 vs. db/m+ (analysis of variance [ANOVA]), ^cP<0.01 (ANOVA).

그림 출처: PubMed Central 공개 논문 (PMC12620679)

Conflict of interest statement

CONFLICTS OF INTEREST . Young Min Cho is an outside director of Daewoong Pharmaceutical and received consultation fees from LG Chemicals.

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