Objective To explore effects and potential mechanisms of high insulin environment on high density lipoprotein (HDL) generation-related functional protein ABCA1.Methods [3 H] labeled cholesterol efflux from mature 3T3-L1 adipocytes was detected by liquid scintillation counting.ABCA1 mRNA and protein expression in mature 3T3-L1 adipocytes post stimulation with various concentrations of insulin was detected by real-time fluorescence-based quantitative techniques and Western blot,respectively,in the absence and presence of CHX (cycloheximide,CHX),calpeptin (calpain pathway inhibitor) or MG-132 (proteasome pathway inhibitor).Results Cholesterol efflux rates were reduced post insulin stimulation in a dose-dependent manner ((7.06 ± 0.27) ％,(6.59 ± 0.30) ％,(6.34 ± 0.24) ％,(5.07 ± 0.40) ％,and (4.71 ±0.40)％ at 0,1,10,102,and 103 nmol/L of insulin,P ＜0.05).Cholesterol efflux rates decreased in a time-dependent manner post 103 nmol/L insulin stimulation (6.52 ± 0.30) ％,(5.59 ± 0.71)％,(5.44 ± 0.37)％,(4.52 ± 0.32)％,and (4.38 ± 0.33)％ at 0,2,4,6,12 h,respectively).ABCA1mRNA levels were not affected by insulin(P ＞ 0.05).ABCA1 protein level was significantly downregulated in 103 nmol/L insulin group compared to 0 nmol/L insulin group (P ＜ 0.01).Compared with the 0 h group,ABCA1 protein level was significantly reduced in 6 h group (P ＜ 0.05) and further reduced in 12 h group (P ＜ 0.01).Both calpeptin and MG-132 could partly reduce insulin-induced degradation of ABCA1.Compared with the negative control group,ABCA1 protein levels were significantly upregulated by cotreatment with calpeptin and MG-132,respectively (both P ＜ 0.01).Conclusion Our data suggest that high insulin level could promote the ABCA1 protein degradation and reduce cholesterol efflux from mature 3T3-L1 adipocytes through calpain and proteasome pathway,thus,produce a circumference not suitable for nascent HDL formation in 3T3-L1 adipocytes.