Background: Angiopoietin-like protein 3 (ANGPTL3) is produced by hepatocytes. Once secreted, it inhibits endothelial lipase activity and regulates angiogenesis. We previously reported the positive association between serum ANGPTL3 and HDL function in nondiabetic participants. Hereby, we investigated whether ANGPTL3 directly modulates HDL function. Methods: HDLs were isolated from plasma. ANGPTL3 in HDLs was studied by western blot and ELISA. Cholesterol efflux to HDLs was analyzed in macrophages preloaded with fluorescent cholesterol. Recombinant human ANGPTL3 was subjected to endothelial cells treated with TNF-α and ICAM-1 expression was analyzed by FACS. Similar experiments were repeated in db mice injected with AAV encoding human ANGPTL3 cDNA or shRNA. Results: ANGPTL3 was present in human HDLs, which were lower in T2DM patients than controls (p=0.04). ANGPLT3 in HDLs was positively associated with cholesterol efflux in nondiabetic controls (r=0.33, p=0.044) but not in T2DM patients (r=0.05, p=0.77). Recombinant ANGPTL3 reduced ICAM-1 expression in endothelial cells exposed to TNF-α (p=0.0001). Similarly, ANGPTL3 levels in HDLs were lower in db mice at 18 weeks old than 8 weeks old, which was accompanied with reduced cholesterol reflux to HDLs and rendered inhibition of ICAM-1 expression in endothelial cells (p<0.for all). Following AAV-mediated ANGPTL3 cDNA transfer, ANGTL3 levels were increased in HDLs of db mice, which increased cholesterol efflux and decreased ICAM-1 expression compared with db mice with vector transfer. Vice versa, decreased ANGPTL3 levels in HDLs by AAV-mediated shRNA transfer abrogated HDL function mentioned above (p<0.for both). When placed on high fat diet for 10 weeks, atherosclerotic plaque was larger in db mice with ANGPTL3 shRNA transfer than other gene transferred groups (p<0.05). Conclusion: ANGPTL3 is a HDL component and direct regulates HDL function. Decrease of ANGPTL3 expression may reduce HDL function and accelerate atherosclerosis in T2DM. Disclosure L. Yang: None. Y. Wang: None. Y. Fu: None. J. Lang: None. D. Zhao: None. Y. Feng: None.