GLP-1 receptor agonist attenuates endoplasmic reticulum stress-mediated β-cell damage in Akita mice

Shunsuke Yamane, Yoshiyuki Hamamoto, Shin-Ichi Harashima, Norio Harada, Akihiro Hamasaki, Kentaro Toyoda, Kazuyo Fujita, Erina Joo, Yutaka Seino, Nobuya Inagaki, Shunsuke Yamane, Yoshiyuki Hamamoto, Shin-Ichi Harashima, Norio Harada, Akihiro Hamasaki, Kentaro Toyoda, Kazuyo Fujita, Erina Joo, Yutaka Seino, Nobuya Inagaki

Abstract

Aims/Introduction: Endoplasmic reticulum (ER) stress is one of the contributing factors in the development of type 2 diabetes. To investigate the cytoprotective effect of glucagon-like peptide 1 receptor (GLP-1R) signaling in vivo, we examined the action of exendin-4 (Ex-4), a potent GLP-1R agonist, on β-cell apoptosis in Akita mice, an animal model of ER stress-mediated diabetes.

Materials and methods: Ex-4, phosphate-buffered saline (PBS) or phlorizin were injected intraperitoneally twice a day from 3 to 5 weeks-of-age. We evaluated the changes in blood glucose levels, bodyweights, and pancreatic insulin-positive area and number of islets. The effect of Ex-4 on the numbers of C/EBP-homologous protein (CHOP)-, TdT-mediated dUTP-biotin nick-end labeling (TUNEL)- or proliferating cell nuclear antigen-positive β-cells were also evaluated.

Results: Ex-4 significantly reduced blood glucose levels and increased both the insulin-positive area and the number of islets compared with PBS-treated mice. In contrast, there was no significant difference in the insulin-positive area between PBS-treated mice and phlorizin-treated mice, in which blood glucose levels were controlled similarly to those in Ex-4-treated mice. Furthermore, treatment of Akita mice with Ex-4 resulted in a significant decrease in the number of CHOP-positive β-cells and TUNEL-positive β-cells, and in CHOP mRNA levels in β-cells, but there was no significant difference between the PBS-treated group and the phlorizin-treated group. Proliferating cell nuclear antigen staining showed no significant difference among the three groups in proliferation of β-cells.

Conclusions: These data suggest that Ex-4 treatment can attenuate ER stress-mediated β-cell damage, mainly through a reduction of apoptotic cell death that is independent of lowered blood glucose levels. (J Diabetes Invest, doi: 10.1111/j.2040-1124.2010.00075.x, 2010).

Keywords: Apoptosis; Endoplasmic reticulum stress; Glucagon‐like peptide‐1.

Figures

Figure 1
Figure 1
Ex‐4 significantly reduced blood glucose levels in Akita mice. (a) Blood glucose concentration and (b) bodyweight were measured in wild‐type C56BL/6 mice (closed diamond, n = 10), Akita mice treated with PBS alone (closed circle, n = 10), Ex‐4 (closed square, n = 12) and phlorizin (closed triangle, n = 10). Each symbol represents mean ± SE. *P < 0.05, **P < 0.01 vs PBS‐treated Akita mice.
Figure 2
Figure 2
Ex‐4 treatment increased insulin‐positive areas, number of islets and insulin content. (a–d) Representative mouse pancreata at 5 weeks‐of‐age stained with insulin. (a) Wild, (b) Akita mice treated with PBS, (c) Ex‐4 or (d) phlorizin. (e) Insulin‐positive areas and (f) number of islets were evaluated as described in Materials and Methods (n = 5 for each group). (g) Pancreatic insulin content was measured as described in Materials and Methods, and expressed as ng/islet (n = 5 for each group). Each column represents mean ± SE. *P < 0.05, **P < 0.01.
Figure 3
Figure 3
Ex‐4 treatment resulted in a significant decrease in the expression levels of C/EBP‐homologous protein (CHOP) mRNA and Bip mRNA in Akita mice. (a) mRNA expression levels of CHOP were evaluated by quantitative real‐time polymerase chain reaction (PCR). (b) mRNA expression levels of BiP were evaluated by quantitative real‐time PCR. Data are expressed as the ratio to that of glyceraldehyde 3‐phosphate dehydrogenase in the same sample (n = 5 for each group). Each column represents mean ± SE. *P < 0.05, **P < 0.01.
Figure 4
Figure 4
Ex‐4 treatment resulted in a significant decrease in the ratio of C/EBP‐homologous protein (CHOP)‐positive β‐cells in Akita mice. (a) Representative mouse pancreata at 5 weeks‐of‐age stained with insulin (red), CHOP (green) and DAPI (blue). (b) The number of CHOP‐positive β‐cells normalized per insulin‐positive area was quantified as described in Materials and Methods. Each column represents mean ± SE. **P < 0.01.
Figure 5
Figure 5
Ex‐4 treatment decreased the ratio of TUNEL‐positive β‐cells. (a) Representative mouse pancreata at 5 weeks‐of‐age stained with insulin (red) and TUNEL (green). (b) The number of TUNEL‐positive β‐cells normalized per insulin‐positive area was quantified as described in Materials and Methods. Each column represents mean ± SE. **P < 0.01.
Figure 6
Figure 6
Ex‐4 treatment did not significantly increase the ratio of PCNA‐positive β‐cells. (a) Representative mouse pancreata at 5 weeks‐of‐age stained with insulin (red) and PCNA (green). (b) The number of PCNA‐positive β‐cells normalized per insulin‐positive area was quantified as described in Materials and Methods. Each column represents mean ± SE.

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