Autoantibodies Against Oxidized Low Density Lipoprotein Correlated With Coronary Heart Disease

Atherosclerosis is a chronic inflammation in the vascular wall,where activated T cells,monocytes and macrophages are present in significant amounts.Autoimmune factors have been shown to play a dominant role in the progression of the atherosclerotic plaque Accrodingly,a humoral and cellular immune responses enhance foam cell formation and inflammatory cell recruitment at sites of endothelial injury prone to atherosclerosis.Candiate autoantigens suggested to play a role in the autoimmune process mclude:oxidized low density lipoprotem(OXLDL) and heart shock protein 65.Antibodies to OXLDL(OXLDL-Ab) is present both in the atherosclerotic lesions and in human serum.Case-control studies have shown that serun OXLDL-Ab titer was elevated in patients with coronary heart diease,early-onset peripheral vascular disease and severe carotid atherosclerosis.However,immunization of experimental animals with OXLDLsuch that high OXLDL-Abtiters were generated decreased atherosclerotic lesions.So far,it is not established whether the immune responses to OXLDL are proatherogenic or antiatherogenic in vivo2002ObjectivesIn the present study, to evaluate the possible role of autoantibodies against oxidized low density lipoprotein(OXLDL-Ab) in the development of atherosclerosis and the relationship between OXLDL-Ab and coronary heart disease(CHD),we used an enzyme-linked immunosorbent assay(ELISA) technique to measure OXLDL,OXLDL-Ab(IgG and IgM) and circulation immune complex in 61 patients with CHD underwent coronary angiography and 116 patients with essential hypertension compared with 123 age-matched normal control subjects.Objects and Methods1. Patients and ControlsCoronary heart disease(CHD) group: There are 61 patients with CHD in our study (45 males, 16 females,mean age=65.31 ?.38 years),who presented to 2nd affiliated hospital of mwdical college of zhejiang university.The diagnosis of CHD was verfied by coronary artery angiography. The subjects accompanying diabetes,tumor,cerebrovascular diseases,penpheral vascular disaeses and autoimmune diseases are excludedEssential hypertension(EH) group: There are 116 patients with EH(81 males,35 females, mean age=62.79?10.41 years) in our study ,defined according to the 1998 WHO/ISH criteria and absent of secondary hypertension.Their mean blood pressure is (151.04 ?2750/89.38 ?14 68)mmHg.The subjects accompanying CHD,diabetes ,tumor ,cerebrovascular diseases,peripheral vascular disaeses and72002autoimmune diseases are excluded.Normal control group : The normal control group consists of 123 subjects (82 males,41 females,mean age=62.75?.11 years) draw from the population of clinical healthy examination randomly.2. Methods2.1 Separation OXLDLThe technique of affinity chromatography is used to separate OXLDL from human plasma.Sepharose 4B and mouse anti-OXLDL monoclonal antibody were linked in the column The PH 2.8 O.lmmol/L aminoacetic acid Hcl buffer solution was used to elution OXLDL from the antibody. SDS-PAGE and Western-blot are respectively used to analyze the character of purity and immune of OXLDL preparation.2.2 Measurement of OXLDL-IgG/IgM l)coated with 96ug/L OXLDL lOOul/well; 2)blocked with 10% bovine serum albumin 200ul/well; 3)add object serum 1:20/1:10 lOOul/well;4)add horseradish peroxidase conjugated goat antihuman immunoglobulin G/M1:1000 lOOul/well;5)substrate solution of orthophenylenediamine(OPD) lOOul/well, 6)stop reaction with 2mol/L of sulfunc acid 50ul/well; 7)absorbance measured at 490nm in a micro-ELISA plate reader.2.3 Measurement of OXLDL-CICl)coated with mouse anti-OXLDL monoclonal antibody lOOul/well; 2)blocked with 10% bovine serum albumin 200ul/well, 3)add object serum 1 :2 lOOul/well,820024)add horseradish peroxidase conjugated goat antihuman immunoglobulin 1:2000lOOul/well; 5)substrate solution of orthophenylenediamine(OPD) lOOul/well;6)stop reaction with 2mol/L of sulfuric acid 50ul/well;7)absorbance measured at 490nm in a micro-ELISA plate reader.