Experimental study of the effect of oxidized low density lipoprotein on macrophages

Abstract

Objective To investigate the effect of oxidized low density lipoprotein(oxLDL) on macrophages and their secretion of macrophage migration inhibitory factor(MMIF/MIF). Methods (1)Preparation of monocytes derived macrophages(MDMs), co-culture with oxLDL at concentrations of 25, 50, 75 mg/L, respectively.MDMs cultured without ox LDL were normal controls(NC).MDM proliferation was detected using cell counting kit- 8 assay(CCK- 8 assay); supernatant MIF was examined using enzyme linked immunosorbent assay(ELISA).(2)Preparation of MDMs derived from THP- 1 cell lines which were transfected with nuclear factor-κB(NF-κB)- LUC plasmids,co- culture with ox LDL as above- mentioned, NF-κB pathway activity was tested by luciferase test.(3) MDMs containing NF-κB - LUC plasmids were co- cultured with ox LDL, self- control study was performed at the same concentration of ox LDL, with or without adding NF-κB pathway inhibitor BAY 11- 7082 at the concentration of 10μmol/L to culture medium.Supernatant MIF was detected. Results (1) Compared with NC, significantly increased MDM proliferation occurred 12 hours after co-culture with oxLDL, the proliferation increased by 17.7%,27.8% and 41.2%, respectively at 48 hours(P<0.01);supernatant MIF level increased simultaneously, reaching 1.49, 1.67 and 2.09 folds, respectively at 48 hours, as compared with NC(P< 0.01).(2) Compared with NC, significant activation of NF-κB pathway was detected 12 hours after co- culture with ox LDL, the activation was increased by 38.1%, 60.3% and 61.1%, respectively at 48 hours(P< 0.01).(3)Compared with NF-κB pathway inhibitor BAY 11- 7082- free group, supernatant MIF level decreased significantly at 12 hours in BAY 11- 7082 group,it decreased by 58.6%,69.3%,69.7% and 73.5%, respectively at 48 hours (P<0.01). Conclusion oxLDL at concentrations of 25 to 75 mg/L could promote MDM proliferation and MIF secretion, this effect was enhanced with prolonged co-culture time. The increased MIF secretion induced by oxLDL might be achieved through activation of the NF-κB pathway. Key words: Oxidized low density lipoprotein; Macrophage; Proliferation; Macrophage migration inhibitory factor; Nuclear factor-κB pathway