The Role And Mechanism Of High-density Lipoprotein Regulating CD4+T Cell Subsets In Rheumatoid Arthritis Mice

Objective: To investigate the possible mechanism of high-density lipoprotein in the treatment of rheumatoid arthritis.Methods: DBA/1 mice were divided into three groups: control group,CIA group and CIA+HDL group.Mice in the CIA and CIA+HDL groups were immunized with an emulsion of Freund's complete adjuvant and bovine type ? collagen by intradermal injection at the root of the tail for the first time on day 0.The control group was injected with saline at the same site.On day 21,the CIA group and CIA+HDL group were injected with an emulsion of Freund's incomplete adjuvant and bovine type ? collagen to enhance immunity effect,and the control group was injected with saline.The CIA+HDL group was injected with high-density lipoprotein(10 mg/kg)through the tail vein on day 40 followed by HDL injections five times every 3 days.The control group and the CIA group were injected with the same volume of saline.Changes in mouse body weight and clinical scores of arthritis were recorded.The mice were sacrificed on day 75,and the serum levels of tumor necrosis factor-?,anti-collagen-II antibody Ig G2 a of the mice were measured using an ELISA kit.Then the tissue sections of ankle joints and major organs of mice were stained with hematoxylin and eosin to analyze the severity of rheumatoid arthritis in mice and the pathological changes of heart,liver,kidney and colon in each group of mice to evaluate the effect of high-density lipoprotein in the treatment of rheumatoid arthritis.Compared the changes of the proportion of Th1,Th2,Th17 and Treg in the spleen CD4 +T cells and the changes of Th1/Th2,Th1/Th17 ratios of the three groups of DBA/1 mice.In vitro,we extracted the bone marrow of tibia and femur of 8-week-old DBA/1 mice and red blood cells were lysed.Then,the cells were cultured in IMDM medium which containing 20 ng/ml GM-CSF,10ng/ml IL-4,and 10% FBS.After three days,a half-volume exchange was performed and collected the suspend cells on day 6.At this time,immature dendritic cells were collected.The immature dendritic cells were divided into a control group,a positive control group,and an experimental group.The positive control group was added with LPS,the control group was added with the same amount of IMDM medium,and the experimental group was added with LPS and HDL at a dose of 1?g/m L,10 ?g/m L,100 ?g/m L.The cells of the above group cultures 48 hours and harvested to analyze the expressions of surface costimulatory molecules of MHC-II,CD86,CD40,and CD80 by flow cytometry.Results: Compared with the CIA group,intravenous injection of HDL at a dose of 10mg/kg can significantly improve the symptoms of rheumatoid arthritis in mice.The results showed that HDL can reduce the clinical score of arthritis and the levels of serum pro-inflammatory factor TNF-? and specific antibody Ig G2?(P<0.05).Histopathological examination showed that HDL can smoothen the ankle articular surface,significantly decrease synovial hyperplasia,inflammatory cell infiltration and bone erosion and other pathological changes.But HDL has no significant effect on liver,heart,kidney,colon and other organs.The results of flow cytometry showed that,HDL could significantly reduce the ratio of Th1/Th2(P<0.05)and significantly increase the ratio of Th1/Th17(P<0.05)compared with the CIA group.After treatment with HDL,the content of Treg cells increased,but the results were not significantly different compared with the CIA group(P>0.05).In vitro,we found that compared with the control group,the expression of surface costimulatory molecules such as MHC-?,CD86,CD40,and CD80 in the positive control group was significantly increased(P<0.05).The expressions of MHC-?,CD86,and CD40 were inhibited in experimental group and the effect was most significant at high-dose(100 ?g/m L HDL)(P <0.05).But HDL at different dose concentrations had no significant inhibited effect on the expression level of CD80.Conclusion: HDL can significantly inhibit collagen-induced rheumatoid arthritis,and its mechanism may be related to the regulation of dendritic cell maturation and the balance of Th1/Th2 and Th1/Th17.This study provides new ideas and directions for the clinical treatment of rheumatoid arthritis and the development of new drugs.