Study On Hyperhomocysteine's Mechanism At Chronic Kidney Disease Accomopanied Cardiac-Cerebrovascular Disease

Homocysteinemia(Hcy) is the intermediate product of methionine.It would cause hyperhomocysteinemia(HHC) when the concentration of total Hcy(tHcy) increases pathologically.Elevated homocysteine (HCY) levels can be caused by a number of factors, including folate and B-vitamin deficiency, pre-existing atherosclerotic disease, diabetes and various drugs. Epidemiological evidence suggests that hyperhomocysteinemia (HHC) is an independent risk factor for cardiovascular and cerebrovascular disease. Chronic kidney disease (CKD) is one of the most frequent causes of HHC and when the renal insufficiency reaches the stage when dialysis treatment is initiated, more than 90% have a moderate degree of HHC (>15 lmol/l). Patients with chronic kidney disease (CKD) have high prevalence of cardiovascular disease (CVD). Study by Moustapha showed that a lμmol/l increase in tHcy was associated with a 1% higher risk of cardiovascular events. As a meta-analysis by Wald et al. showed that a 5 lmol/l increase in tHcy was associated with a 32%higher risk of cardiovascular events and a 59%higher risk of stroke, the increased Hcy levels observed in CKD patients could be considered clinically significant.But it's not the fact. So someone suggestted that HCY is a marker, rather than a cause of CVD.It has been indicated that the expression of tissue factor (TF) in blood cells especially mononcytes plays an important role in the pathogenesis of AS and thrombosis. It was found recently that there were higher Mo TF expression in plsma of ACS patients. And it is also important in AIS patients.Many factors can induce MoTF expression. It has been reported that through the ways such as induction of the TF expression in endothelial cells (ECs), homocysteine (Hcy) could increase the ability of antithrombosis and promote thrombosis. So if can Hcy increase AMI and AIS by inducing MoTF expression? And how to induce MoTF expression? There are no reports.We carried out a series of studies to clarify this issue. Part1Objective To determine the relationship between hyperhomocysteinemia(HHC) and atherosclerosis and cardiovascular and cerebrovascular disease in patients with CKD.Methods Plasma total homocysteine (tHcy) concentrations were measured by using high efficiency liquid chromatography. Carotid initial-medial thickness (IMT) the prevalence of atherosclerosis plaques were measured by echocardiography. Correlation analysis and logistic regression analysis were used to analyse the results.Results The prevalence of HHC was 84.4% in the CKD patients. The tHcy concentration was higher in patients than those in controls (P<0.05). Cartid IMT and the prevalence of atherosclerosis plaques were positively correlated with tHcy (P<0.05). logistic regression analysis indicated that there were positively correlations a independently between cardiovascular and cerebrovascular disease in CKD patients and tHcy and other conventional risk factors such as TG,Hb.Conclusion The results indicate that hyperhomocyseinemiamay be an independent risk factor for atherosclerosis found and cardiovascular and cerebrovascular disease in CKD patients. Part 2Objective This research systematicly observed blood tatal homocysteine(tHcy)level and tissue factor(TF) level produced by monocytes(Mo) in patients of CKD to determine the mechanism of Hcy in acute thrombo-cardiac-cerebro-vascular disease in patients of CKD.Methods 10 patients with CKD who suffered AMI and 9 patients with CKD suffered ASI and 24 patients with CKD were observed. Efficiently liquid color fluorescence method measured blood tHcy level; Sandwich ELISA was performed to detect the TFAg. Chromo-substrate method was used to measure TFact on the suface of Mo,Correlation analysis and Logistic regression analysis were carried out to satistic the results.Results The incidence rates of smoking,drinking,hypertension diabetes and CVD were higher in AMI group and AIS than that in control group (P<0.05).The levels of creatinine and TG were higher in testing group than control. The tHcy and TFAg in AMI group and AIS is obviously higher than that in control group (P<0.05).The level of TFact in AIS were higher significantly than that in AMIgroup and control group (P<0.05).There were positive relationships between tHcy and TFAg and TFact (P<0.05).tHcy and LDL-C were both independent factors of acute thrombo-cardiac-cerebro-vascular disease in patients of CKD(r=0.194,0.114).Conclusion The levels of tHcy and TF increased in acute thrombo-cardiac-cerebro-vascular disease in patients of CKD.So tHcy may cause acute thrombo-cardiac-cerebro-vascular disease in patients of CKD by inducing the production of TF. Part 3Objective The Hcy-induced tissue factor (TF) expression in human monocytes(Mo) and the effect of Hey on the activity of nuclear factor-kappaB (NF-κB) were investigated to determine the action mechanism of Hey.Methods Monocytes of ten patients of CKD were isolated and cultured, and incubated with different concentrations of Hcy/PTDC (NF-κB inhibitor).Sandwich ELISA was perfoumed to detect the TFAg.One-stage-coag method was used to measure PCA, Semi-quantitative RT-PCR was performed to detect the expression of TF mRNA in Mo.Western blot was carried out to detect the expression of NF-κB protein in nuclei.Results There was low expression level of TF protein in the resting Mo.Hey could induce Mo expressing TF mRNA,TFAg,PCA dose-dependently after the Mo were incubated with Hey at concentrations of 10,50,100,500μmol/L (P<0.05).The concentration of TFAg were 92.6±18.7,109.1±27.3,110.3±20.7,290.0±175,341.8±35.8pg/ml after0.5h,1h,2h,3h,4h the Mo were incubated with 100μmol/L Hey (P< 0.05).Also Hey could induce TF mRNA and PCA time-dependently. Additionally, Hey could rapidly induce the activation of NF-κB and this effect could be significantly inhibited by PDTC. It was concluded that Hey could significantly induce the expression of TF in Mo and enhance the activation of NF-KB, subsequently mediate TF gene expression and protein synthesis.Conclusion NF-KB-mediated expression of TF in Mo might be the important mechanism of atherosclerosis and thrombosis induced by Hey.