Scavenger receptor SR-BI has been implicated in HDL-dependent atheroprotective mechanisms. We report the generation of an SR-BI conditional knockout mouse model in which SR-BI gene targeting by loxP site insertion produced a hypomorphic allele (hypomSR-BI). Attenuated SR-BI expression in hypomSR-BI mice resulted in 2-fold elevation in plasma total cholesterol (TC) levels. Cre-mediated SR-BI gene inactivation of the hypomorphic SR-BI allele in hepatocytes (hypomSR-BI–KO^liver) was associated with high plasma TC concentrations, increased plasma free cholesterol/TC (FC/TC) ratio, and a lipoprotein-cholesterol profile typical of SR-BI^–/– mice. Plasma TC levels were increased 2-fold in hypomSR-BI and control mice fed an atherogenic diet, whereas hypomSR-BI–KO^liver and SR-BI^–/– mice developed severe hypercholesterolemia due to accumulation of FC-rich, VLDL-sized particles. Atherosclerosis in hypomSR-BI mice was enhanced (2.5-fold) compared with that in controls, but to a much lower degree than in hypomSR-BI–KO^liver (32-fold) and SR-BI^–/– (48-fold) mice. The latter models did not differ in either plasma lipid levels or in the capacity of VLDL-sized lipoproteins to induce macrophage cholesterol loading. However, reduced atherosclerosis in hypomSR-BI–KO^liver mice was associated with decreased lesional macrophage content as compared with that in SR-BI^–/– mice. These data imply that, in addition to its major atheroprotective role in liver, SR-BI may exert an antiatherogenic role in extrahepatic tissues.