Purpose: We previously demonstrated increased villus height following Retinoblastoma protein (Rb) deletion in the intestinal epithelium (Rb-IKO). Here we determined the functional consequences of augmented mucosal growth on intestinal fat absorption. Methods: Mice with constitutively disrupted Rb expression in the intestinal epithelium (Rb-IKO) along with their floxed (WT) littermates were placed on a high fat diet (HFD, 42% kcal fat). Mice were weighed weekly, and fat absorption determined, along with indirect calorimetry and MRI body composition. Rb-IKO mice were also subjected to small bowel resections (SBR), followed by HFD feeding. In separate experiments, we examined intestinal fat absorption in mice with conditional (tamoxifen inducible) intestinal Rb (inducible Rb-IKO) deletion. Results: Constitutive Rb-IKO mice demonstrated greater mucosal surface area, yet manifested paradoxically impaired intestinal long chain triglyceride absorption and decreased cholesterol absorption. Despite attenuated lipid absorption, there were no differences in metabolic rate, body composition, and weight gain in Rb-IKO and WT mice at baseline as well as following SBR. We also confirmed fat malabsorption in inducible Rb-IKO mice. Conclusion: Despite an expanded mucosal surface area, Rb-IKO mice demonstrate impaired lipid absorption without compensatory alterations in energy homeostasis or body composition. These findings underscore the importance of delineating structural/functional relationships in the gut and suggest a previously unknown role for Rb in the regulation of intestinal lipid absorption.