Inflammation Aggravates Intracellular Cholesterol Accumulation And Injury In Human Mesangial Cell Loaded By Low-Density Lipoprotein Via Up-Regulating NPC1 Expression

Objective: To investigate the effect of IL1-βon NPC1(Niemann-pick Protein C1)expression in human mesangial cell(HMCs)loaded by low-density lipoprotein(LDL)and explore whether NPC1-mediated cholesterol accumulation causes cell injury.Methods: HMCs were divided into control group,LDL group,LDL+IL-1β group.The level of NPC1 protein was detected by western blot analysis.The total cholesterol and endoplasmic reticulum cholesterol were detected by a cholesterol detection kit.Cell proliferation was measured using a cell count kit.Cell cycle was detected by flow cytometry PI/RN staining.The mRNA levels of NPC1,glucose-regulated protein78(GRP78),protein kinase R-like endoplasmic reticulum kinase(PERK),activating transcription factor 6(ATF6),fibronectin(FN),type IV collagen secretion(Col IV)were measured by real-time quantitativePCR.NPC1,GRP78,FN protein and Colocalization of NPC1/ LAMP-1were detected using immunofluorescent staining.After intervention of u18666 a,a blocker of the NPC1,the level of endoplasmic reticulum cholesterol,cell proliferation,cell cycle,GRP78,FN and Col IV mRNA was measured.Results: We found that LDL loading alone significantly increased the expression of NPC1,promoted intracellular and endoplasmic reticulum cholesterol accumulation,accelerated the proliferation of HMCs,increased the ratio of S phase in cell cycle,and promoted the expression of endoplasmic reticulum stress marker(GRP78,PERK,ATF6)and mesangial matrix marker(FN,Col IV),increased the mean fluorescence intensity of GRP78 and FN.Inflammation further aggravated the level of the above indicators in HMCs loaded by LDL.Compared with LDL+IL-1β group,co-treatment with u18666 a significantly decreased the level of endoplasmic reticulum cholesterol,cell proliferation,ratio of S phase in cell cycle and GRP78,FN,Col IV mRNA.Conclusion: These results suggested that inflammation aggravates intracellular and endoplasmic reticulum cholesterol accumulation in HMCs via up-regulating NPC1 expression and causes endoplasmic reticulum stress(ERS)and cell injury.