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Abstract: Objective To explore the change of α cell mass and its mechanism with diabetic progress. Methods Diabetic mice were killed by exsanguinations after 4, 12 and 20 weeks of diabetes, respectively. Indirect double immunofluorescences for Insulin/Ki67, or BrdU, Cleaved-Caspase 3, TUNEL were used to evaluate pancreatic α cell mass, regeneration and apoptosis of α cells. Indirect triple immunofluorescences for Glucagon/Neurogenin 3/MafA and Western blot analysis for Neurogenin 3 were used to determine neogenesis of pancreatic α cells. Results Pancreatic α cell mass was gradually increased with diabetic progress. It was significantly different from that of controls. There weren’t any proliferation and apoptosis of α cell during diabetes, however, many Neurogenin 3+ cells appeared in the pancreatic islets of diabetic mice, and most of them were co-stained with MafA and Glucagon. Conclusion Pancreatic α cell mass is gradually increased with diabetic progress. It seems to be strongly associated with neogenesis of pancreatic α cells.