Oxidized Lipoprotein (a) Detection And Its Clinical Application

Elevated lipoprotein(a)is an independent risk factor for cardiovascular disease.Oxidized Lp(a)[ox-Lp(a)]can lead to macrophage change into foam cell,promote to release cytokine ,accelerate Atherosclerosis.Objective:(1)To isolate and identify human autoantibodies against ox-Lp(a).(2)To develop new ELISA for ox-Lp(a)by using isolated human autoantibodies and multiclone antibodies against ox-Lp(a).(3)To study the significance of Lp(a)in the pathogenesis of coronary lesion.(4)To oxidize Lp(a)for different level and develop standardization of ox-Lp(a)measurements.Methords:(1)We isolated and identified ox-Lp(a)autoantibodies from healthy subjects by affinity chromatography,with the use of ox-Lp(a)cross-linked to Sepharose 4B.(2)We developed two "sandwich" ELISAs for measuring plasma ox-Lp(a)level,using human autoantibodies against ox-Lp(a)or specific rabbit antiserum against human ox-LDL as the capture antibody and quantitating with monoclonal anti-apo(a)enzyme conjugate, respectively.And ox-Lp(a)levels were studied in 100 patients with coronary heart disease (CHD)and 100 control subjects.(3)ELISA was used to measure the Lp(a)in 139 patients with coronary heart disease.The patients were divided by coronary angiography into a multi-vessel diseased group,double-vessel diseased group and single-vessel diseased group. (4)To oxidize Lp(a)for different level with different concentration of Cu²⁺and malondialdehyde(MDA),to observe the changed character of thiobarbituric acid-reactive substances(TBARS),conjugated dienes(CD)and oxidized site.Results:(1)Ox-Lp(a)autoantibodies were isolated from all the selected 8 healthy subjects. Two of them had both specific autoantibodies against ox-LDL and ox-apo(a).(2)The two ELISA methods we setted up have good character of accuracy,reliability,and specificity.(3) The Lp(a)levels in the patients with CHD were significantly different from those of control (279.6±162.7mg/l vs.206.3±126.4mg/l,P＜0.0001).Plasma ox-Lp(a)levels in patients with CHD detected by two ELISAs were both significantly higher than those of control(ELISA using human antibodies against ox-Lp(a):24.3±33.4 vs.8.4±9.3μg/ml,P＜0.0001;ELISA using antibodies against ox-LDL:13.0±13.8 vs.7.3±9.7μg/ml,P＜0.0001,respectively). Furthermore,a significantly positive relation between ox-Lp(a)levels detected by two ELISAs was also found(r=0.78,P＜0.0001).The TC level was higher in the double-vessel diseased group than in the single-vessel diseased group(p＜0.01).The Ox-Lp(a)level was significantly higher in the multi-vessel diseased group than in the double-vessel and single-vessel diseased group.(4)After oxidization,the content of TBARS and CD increase first,with time extended,the content decrease.The oxidized site is also changed.There is a negative relationship between the Lp(a)levels and oxidative time(r=-0.437,p=0.002).Conclusion:Ox-Lp(a)autoantibodies against ox-Lp(a)are isolated from healthy subjects. We develope a new ELISA for ox-Lp(a)by using human antibodies.And ox-Lp(a)level increased in the CHD patients.Ox-Lp(a)might play some roles in the pathogenesis of coronary lesion.Oxidative modification of Lp(a)causes changes in structure and biological properties of Lp(a),enhances conjugation of macrophageand scavenger receptor,accelerates Atherosclerosis.