Diabetes mellitus (DM), also called diabetes, is a chronic metabolic disorder, with complications that include failure of immune responses, such as immune cell proliferation and activation. The current study investigated changes in natural killer (NK) cell activity under diabetic conditions. To demonstrate the influence of hyperglycemia on NK cell activity, type 1 and 2 diabetes models were constructed using C57/BL6 mice. NK cells isolated from mice with either type 1 or 2 diabetes showed lower cytotoxic activity against cancer cells than NK cells isolated from healthy control mice, and the longer the duration since induction of diabetes, the lower was the cytotoxic activity. Moreover, in the diabetic mouse models, fewer NK cells expressed activation markers, such as NKG2D and NKp46, compared to NK cells from control mice. Production of the apoptosis inducer, granzyme B, was also lower in NK cells from diabetic mice than control mice. These results demonstrate that high glucose conditions influence NK cell activity.