The Effect And Mechanism Of CTRP9 On The Cholesterol-reversal Transport Of RAW264.7 Source Foam Cells

Objective:First,we cultured RAW264.7,then induced it to become a foam cell,and observed the effect of CTRP9 on the reverse transport of RAW264.7 derived foam cells,and further explored its possible mechanism.Methods:1.First,RAW264.7 was cultured between cells and then treated with 50mg/L oxidized low-density lipoprotein(ox-ldl),then incubated for 24 h,and induced into foam cells.2.The experimental group:(1)The effect of control time on all indexes and cholesterol levels of RAW264.7 source foam cells was measured.1 hour experimental group;3 hours experimental group;For 6hours in the experimental group.The above is treated with 3ug/ml CTRP9 for different time.Real-time fluorescent quantitative method of Real-Time PCR assay each cell ABCA1,ABCG1,LXRa mRNA expression of relative quantity,Western blot-determination of each cell ABCA1,ABCG1,LXRa protein content,high performance liquid chromatography(HPLC)determination of cholesterol in the cell.To observe the effect of CTRP9 on ABCA1,ABCG1 and LXRa in RAW264.7 cell source foam cells.The best time points for the index and cholesterol level of the cells of RAW264.7 cells were screened out.(2)The effects of different concentrations of CTRP9 on the various indexes and cholesterol levels of RAW264.7 source foam cells were determined.(2)0.3 ug/ml CTRP9group;3.1 ug/ml CTRP9 group;(4)3 ug/ml CTRP9 group.The culture medium with no control of CTRP9 was cultured for 3 hours.The culture medium containing the corresponding concentration of CTRP9 was cultured for 3 hours.Real-time fluorescentquantitative method of Real-Time PCR assay each cell ABCA1,ABCG1,LXRa mRNA expression of relative quantity,Western blot-determination of each cell ABCA1,ABCG1,LXRa protein content,high performance liquid chromatography(HPLC)determination of cholesterol in the cell.To observe the effect of CTRP9 on ABCA1,ABCG1 and LXRa in RAW264.7 cell source foam cells.The optimal concentration of the control index and cholesterol level of the control of RAW264.7 source foam cells was screened out.Result(1)Using ox-ldl to successfully induce RAW264.7 to become foam cells;(2)RT PCR and Western blot results shows: under the same concentration,each index in the normal RAW264.7 mRNA and protein expression level is very low,in the index level of CTRP9 intervention in 1 hour begins to rise,with the extension of time,the content of each index increased significantly,and reached a peak in 3 hours,6 hours were decreased,but than content is still high with the control group(P < 0.05);High performance liquid chromatographic results showed that the cholesterol level was very high in normal RAW264.7 cells,and the cholesterol level began to decrease after 1 hour of intervention of CTRP9.The cholesterol level decreased significantly with time,and reached the lowest level in 3 hours,the highest in 6 hours,but still lower than the control group(P<0.05).The above results indicated that the three hours experimental group had the most significant effect on the indexes and cholesterol levels of RAW264.7 source foam cells.(3)RT PCR and Western blot results shows: under the same time,with the increasing concentration of CTRP9,the index of cell m RNA and protein expression level increased gradually,gradually reduce cholesterol levels in cells,cells in CTRP9 concentration reaches 3 ug/ml each index of the highest amount of m RNA and protein expression,intracellular cholesterol content is the lowest,comprehensive the above results show that the 3 ug/ml CTRP9 group of RAW264.7 source sex each index of the foam cells and cholesterol effect the most significant(P < 0.05);Conclusion:1.CTRP9 increased the expression of RAW264.7 source foam cells ABCA1,ABCG1 and LXRa through mRNA and protein levels,while increasing the rate of cholesterol flow and lowering the cholesterol level in the cells.2.CTRP9 increased the expression of RAW264.7 source foam cells ABCA1,ABCG1 and LXRa through mRNA and protein levels,while increasing the rate of cholesterol flow and lowering the cholesterol level in the cells.It has time and concentration dependence.