Transgenic mice expressing the amyloidogenic human islet amyloid polypeptide (hIAPP) in their islet β-cells are a model of islet amyloid formation as it occurs in type 2 diabetes. Our hIAPP transgenic mice developed islet amyloid when fed a breeder chow but not regular chow. Because the breeder chow contained increased amounts of fat, we hypothesized that increased dietary fat enhances islet amyloid formation. To test this hypothesis, we fed male hIAPP transgenic and nontransgenic control mice diets containing 15% (low fat), 30% (medium fat), or 45% (high fat) of calories derived from fat for 12 months, and we measured islet amyloid, islet endocrine cell composition, and β-cell function. Increased dietary fat in hIAPP transgenic mice was associated with a dose-dependent increase in both the prevalence (percentage of islets containing amyloid deposits; 34 ± 8, 45 ± 8, and 58 ± 10%, P < 0.05) and severity (percentage of islet area occupied by amyloid; 0.8 ± 0.5, 1.0 ± 0.5, and 4.6 ± 2.5%, P = 0.05) of islet amyloid. In addition, in these hIAPP transgenic mice, there was a dose-dependent decrease in the proportion of islet area comprising β-cells, with no significant change in islet size. In contrast, nontransgenic mice adapted to diet-induced obesity by increasing their islet size more than twofold. Increased dietary fat was associated with impaired insulin secretion in hIAPP transgenic (P = 0.05) but not nontransgenic mice. In summary, dietary fat enhances both the prevalence and severity of islet amyloid and leads to β-cell loss and impaired insulin secretion. Because both morphologic and functional defects are present in hIAPP transgenic mice, this would suggest that the effect of dietary fat to enhance islet amyloid formation might play a role in the pathogenesis of the islet lesion of type 2 diabetes in humans.