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Objectives: The aim of this study was to determine whether high-protein and high-carbohydrate diets exert differential effects on serum cholesterol, triglyceride and fructosamine concentrations in healthy cats., Methods: A randomised, crossover diet trial was performed in 35 healthy shelter cats. Following baseline health assessments, cats were randomised into groups receiving either a high-protein or high-carbohydrate diet for 4 weeks. The cats were then fed a washout diet for 4 weeks before being transitioned to whichever of the two studied diets they had not yet received. Fasting serum cholesterol, triglyceride and fructosamine concentrations were determined at the end of each 4-week diet period., Results: Cats on the high-carbohydrate diet had significantly lower serum cholesterol ( P <0.001) concentrations compared with baseline measurements. Cats on the high-protein diet had significantly higher serum cholesterol ( P  <0.001) and triglyceride ( P  <0.001) concentrations, yet lower fructosamine ( P <0.001) concentrations compared with baseline measurements. In contrast, overweight cats (body condition score [BCS] >5) had lower cholesterol ( P  = 0.007) and triglyceride ( P  = 0.032) concentrations on the high-protein diet than cats within other BCS groups., Conclusions and Relevance: Diets higher in protein and lower in carbohydrates appear beneficial for short-term glucose control in healthy cats. A high-protein diet was associated with significantly elevated cholesterol and triglyceride concentrations in healthy cats, even though the increase was significantly less pronounced in cats with a BCS >5. This finding suggests that overweight cats process high-protein diets, cholesterol and triglycerides differently than leaner cats.