Adaptor protein APPL1 counteracts streptozotocin-induced diabetes and ß cells loss in mice

Abstract

Loss of pancreatic β cells mass and function are the major contributors to the pathogenesis of type 1 and type 2 diabetes mellitus. Inflammation is a key culprit in mediating β-cell dysfunction and subsequent apoptosis in diabetes. APPL1, an adaptor protein containing a NH2-terminal Bin1/Amphiphysin/Rvs167 domain, a central pleckstrin homology domain and C-terminal phosphotyrosine binding domain, regulates insulin and adiponectin signaling cascades, thereby maintaining glucose homeostasis. Human with loss-of-function mutations in APPL1 gene develop diabetes. We previously demonstrated that APPL1 promotes insulin sensitivity and insulin secretion by fine-tuning Akt activity. In type 2 diabetic mice, β cell APPL1 is decreased, leading to defective insulin secretion and subsequent glucose intolerance. In this study, we aimed to explore whether APPL1 involves in β-cell mass maintenance and β-cell inflammation in type 1 diabetes using in vivo and ex vivo approaches.

Like in type 2 diabetes, APPL1 expression was dramatically decreased in pancreatic β cells of streptozotocin (STZ)-induced and genetically inherited type 1 diabetic mouse models. The reduction of APPL1 was associated with diminished insulin content and increased insulitis in type 1 diabetic mice. Interestingly, treatment with diabetogenic cytokine cocktail (tumor necrosis factor α [TNFα], interleukin 1 β [IL1 β] and interferon 1-γ [IFN γ]) reduced APPL1 mRNA expression in pancreatic islets. To directly examine whether APPL1 plays a role in type 1 diabetes, we injected APPL1 global knockout (KO) mice and their wild-type (WT) littermates with STZ to induce β cell apoptosis and diabetes. Genetic ablation of APPL1 exacerbated STZ-induced hyperglycemia, glucose intolerance, impaired glucose-stimulated insulin secretion. These changes were associated with reduced β-cell mass and insulin content and increased insulitis in APPL1 KO mice. The reduced β cell mass was due to elevated β cell apoptosis and reduced β cell proliferation in APPL1 KO mice.

To test whether β-cell specific overexpression of APPL1 counteracts STZ-induced diabetes, we created two distinct mouse models that expressed human APPL1 under the control of rat insulin promoter and modified mouse insulin promoter using transgenic and adeno-associated viral (AAV) gene transfer approaches, respectively. Immunoblotting and immunohistochemical analysis revealed that exogenous APPL1 were detected in pancreatic β cells of the above APPL1 overexpression mouse models. AAV-mediated overexpression of APPL1 in β cells conferred resistance to diabetes induced by STZ. STZ-induced reduction in pancreatic β-cell mass and insulin content were partially reversed by APPL1 overexpression, and these changes were due to decreased β-cell apoptosis and increased β-cell proliferation. Likewise, transgenic expression of APPL1 in β cells prevented STZ-induced diabetes. Further analysis revealed that APPL1 regulated both β-cell apoptosis and inflammation in β cells using ex vivo and in vitro approaches. APPL1 deficiency potentiated STZ- and the cytokine cocktail-induced inflammatory response in β cells, as evidenced by increased expression of inducible nitric oxide synthase, NF-κB activity and inflammatory chemokines.

These results collectively suggest that APPL1 is a key common signaling molecule protects β cell from apoptosis and inflammation, and it might represent a potential target for the future development of new strategies for treating and preventing type 1 diabetes.