Effects Of LDL And OxLDL On Expression Of PCSK9 And LDLR In THP-1 Macrophages

Members of the PC family play a central role in the processing of various protein precursors ranging from hormones and growth factors to bacterial toxins and viral glycoproteins. PCSK9 gene code Neural apoptosis- regulated convertase 1 (NARC-1) .As a member of PC family,PCSK9 gene can reduce the amount of LDLR on hepatic cells, affect internalization of LDL and thus induce hypercholesterolemia. Recent studies indicate that the mutation of PCSK9 gene is the third gene association with Autosomal dominant hypercholesterolemia (ADH) after mutations in the genes encoding the LDL receptor and apoB100. Studies on PCSK9 including its relationship to LDL and the role in hypercholesterolemia and atherosclerosis may help to clarify the mechanism, and put new sights into prevention of hypercholesterolemia and atherosclerosis. In our study, we investigated the effect of low density lipoprotein, oxidized low density lipoprotein on PCSK9 and LDLR, try to find the association between PCSK9 and LDLR on THP-1 cells, and support the role of PCSK9 on the development of atherosclerosis.AIM: To study the effects of LDL and oxLDL on expression of PCSK9 and LDLR in THP-1 macrophages and find the relationship between them.METHODS: THP-1 cells were induced to differentiation into macrophages by PMA treatment. The cells were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum. The experiments were designed as follows: cells were co-incubated with LDL and oxLDL with a concentration of 0μg/ml,10μg/ml,20μg/ml,30μg/ml for 24hr separately. Cellular lipid accumulation was determined by Oil Red O staining . The localization and Semiquantitation of PCSK9 was confirmed by Immunofluorescence assay,the expression of PCSK9 and LDLR was analyzed by reverse transcription polymerase chain reaction (for mRNA expression) and Western blotting (for protein expression).RESULTS: Oil Red O staining showed that a mumber of visualized lipid droplet accumulated within THP-1 macrophages both in treating with oxLDL and LDL. The lipid droplet is big and accumulating in cells co-incubated with oxLDL but small and dispersing when co-incubated with LDL. Matured PCSK9 are expressed on the cell surface and increased with the increasing of LDL concentration through Immunofluorescence assay. RT-PCR,Western blot show that, in THP-1 macrophages, LDLR was downregulated while PCSK9 was upregulated especially when teated with LDL , but the treatment of low concentration of oxLDL seemed had no effect on expression of LDLR and PCSK9.CONCLUSIONS: The lipid accumulation in THP-1 macrophages effected the expression of PCSK9. The expression of LDLR was decreased by LDL and the variety of PCSK9 was conversed. Low concentration of oxLDL had no effect on the expression of PCSK9 and LDLR.