Background: Type 1 diabetes (T1D) is characterized by selective destruction of the pancreatic β- cells. Advanced disease is also associated with dysfunction of α cells which results in life threatening hypoglycemic episodes and unawareness. The cause and the underlying mechanisms of α- cell dysfunction are unknown. Previous reports have suggested that β- cells regulate α cells via paracrine signaling. Extracellular vesicles such as exosomes are nanosized (30–300 nm) membrane vesicles secreted by most cell types to communicate with neighboring and distant cells. Alpha and beta cells also release exosomes, but the functional in response to physiological stimulation needs investigation. Thus, the main objective of this study is to investigate the crosstalk that takes place between α and β-cell via exosome.  

Methods: Mouse TC1- α and βTC6 cell lines were stimulated with low (2.5mM), basal (5.5mM) and high glucose (25mM) media for 24hrs after exosome depletion. Conditioned media was collected for isolation of exosomes using differential centrifugation method and kit-based method for higher yields. Exosome was confirmed using TEM, and western blot for exosomal markers. Quantification and characterization was done using ELISA, micro-BCA analysis, and Nanosight analysis. Exosomes were used to further cross-stimulate a and b cells under low and high glucose condition at various time points (6h, 12h, 24h, 48h), and to evaluate the role of exosomes in insulin and glucagon gene expression and secretion. ELISA was used to measure insulin and glucagon levels in the media, q-PCR was performed on insulin, glucagon and their regulating transcription factors.

Results: TEM analysis revealed that a and b cells released a homogenous mixture of cup-shaped, rounded vesicles within the size range of exosomes (30-300nm). The size of exosomes based on nanosight analysis was found to be 50 – 200nm. Western blot analysis for exosomal markers (CD9) further confirmed the presence of exosomes. Glucagon expression in a cells was significantly increased when stimulated with b cell exosomes under low glucose condition (6h: P = 0.0134; 24h: P=0.0455; 48h: P= 0.0532). The expression of glucagon in a cell was not affected by b cell exosomes under high glucose. MafB expression in α cells correlated with glucagon expression. Interestingly, expression of insulin in b cells was not significantly affected by a cell exosomes.

Conclusion: Exosomes from beta cells seem to play a role in the regulation of glucagon by a cells. Further investigation relating to production and release of insulin and glucagon in response to exosomes is warranted.