Salusin-α Inhibits THP-1 Derived Foam Cell Formation Via PPARγ

Background: Atherosclerosis(AS)is the common pathophysiological basis of cardiovascular and cerebrovascular diseases.The newly discovered bioactive peptide Salusin-α has anti-atherosclerosis effect,it mainly inhibits foam cell formation by inhibiting the expression of Acyl-co A: cholesterol acyltransferase 1(ACAT1).Peroxisome pro-liferator-activated receptor gamma(PPARγ)is a class of ligand-activated nuclear transcription factors,which plays an anti-atherosclerotic role in atherosclerosis after activation.Objective: To observe the changes of PPARγ and ACAT1 expression in the THP-1 derived foam cell formation induced by Salusin-α,and to preliminarily investigate whether Salusin-α inhibits the THP-1 derived foam cells formation by downregulating ACAT1 expression through PPARγ pathway.Methods:(1)To induce monocyte to macrophage differentiation,THP-1cells were cultured with 160 n M Phorbol 12-myristate 13-acetate(PMA)for 48 h.THP-1 derived macrophage were divided into control group,which were incubated with 50 mg/L oxidized low density lipoprotein(ox-LDL)for 48 h,and intervention group,which were incubated with 50 mg/L ox-LDL and Salusin-α at different concentrations(0.2,0.4,0.6,0.8,1.0 n M)for 48 h.The free cholesterol(FC)and total cholesterol(TC)were measured by enzyme linked photometry.The difference between TC and FC was cholesterol ester(CE).The ratio of CE to TC(CE/TC)in each group was compared.The changes of cytoplasmic lipid droplets were observed by oil red O staining.RT-PCR and Western blot were used to detect relative m RNA and protein expression level of ACAT1 and PPARγ in each group.(2)To induce monocyte to macrophage differentiation,THP-1 cells were cultured with 160 n M PMA for 48 h.THP-1derived macrophage were divided into control group,Salusin-α group,Salusin-α+T0070907 group,and T0070907 group.T0070907 is a specific PPARγantagonist.The control group was cultured with 50 mg/L ox-LDL for 48 h to induce into foam cell,the Salusin-α group was cultured with 50 mg/L ox-LDL and 0.6 n M Salusin-α at the same time for 48 h,the Salusin-α+T0070907 groupwas pretreated with 10 μM T0070907 for 2 h before incubation with ox-LDL and Salusin-α,the T0070907 group was pretreated with 10 μM T0070907 for 2 h before incubation with ox-LDL.The observation indexes of each group were the same as those of the first part.Results:1.The CE/TC of the control group was 67.13±1.88%.Oil red O staining showed more lipid droplets in cytoplasm,which indicated macrophage-derived foam cell model of THP-1 cell line was established successfully.2.The CE/TC of the intervention group were 9.15±2.71%,52.99±2.51%,45.38±2.99%,52.18±2.95%,55.99±2.10%,respectively.With the increase of Salusin-α concentration,the CE/TC first decreased and then slightly increased,which indicated the best concentration was 0.6 n M(all P<0.05).The CE/TC of control group,Salusin-α group,Salusin-α+T0070907 group,and T0070907 group were 67.13±1.88%,45.38±2.99%,59.07±2.54%,68.23±1.60%,respectively(all P<0.05).T0070907 partially reversed the decrease of CE/TC induced by Salusin-α.3.Oil red O staining showed that,Salusin-a inhibited the formation of THP-1 derived foam cell,while T0070907 attenuated the inhibitory effect of Salusin-α on THP-1 derived foam cell formation.4.The outcomes of RT-PCR showed that,with the increase of Salusin-αconcentration,the relative expression level of ACAT1 m RNA decreased,while the relative expression level of PPARγ m RNA increased(all P<0.05).T0070907 partially weaken the effect of Salusin-α on the relative expression level of ACAT1 m RNA and PPARγ m RNA(all P<0.05).5.The outcomes of Western blot showed that,with the increase of Salusin-αconcentration,the relative expression level of ACAT1 protein decreased,while the relative expression level of PPARγ protein increased(all P<0.05).T0070907 partially attenuated the effect of Salusin-α on the relative expression level of ACAT1 protein and PPARγ protein(all P<0.05).Conclusion: Salusin-α inhibits THP-1 derived foam cell formation bydownregulating ACAT1 expression via PPARγ-dependent pathway,which contributes to its antiatherogenic properties.