人工合成高密度脂蛋白样颗粒测定血清胆固醇酯转运蛋白活性

目的建立血清胆固醇酯转运蛋白活性测定方法。方法用１４Ｃ标记的胆固醇酯合成高密度脂蛋白（ＨＤＬ）样颗粒作为胆固醇酯的供体，以低密度脂蛋白为受体测定血清中胆固醇酯转运蛋白（ＣＥＴＰ）活性，并对４０例健康人、４５例冠心病（ＣＨＤ）、２６例脑卒中患者血清ＣＥＴＰ活性进行分析。结果　该方法线性范围为０～３０％；高（２６％）、低（６．８％）活性批内变异系数（ＣＶ）分别为６．０％、７．５％，批间ＣＶ分别为１１．５％和１２．３％；４５例ＣＨＤ患者血清ＣＥＴＰ活性（ｘ±ｓ）为１７．６％±５．４％，２６例脑卒中患者为１５．２％±３．８％，均明显高于健康对照（１２．７％±３．９％）（Ｐ＜０．０１）。结论该法简便快速、精密度较高，为临床及科研工作中测定ＣＥＴＰ活性提供了一种有效的方法。     

3－磷酸甘油脱氢酶偶联法测定血清醛缩酶活性

探讨用３－磷酸甘油脱氢酶偶联法测定血清中醛缩酶（ＡＬＤ）活性的最佳反应条件。反应体系的终末浓度：ＴＥＡ１００ｍｍｏｌ／Ｌ，ＦＤＰ４ｍｍｏｌ／Ｌ，碘乙酸０．２２ｍｍｏｌ／Ｌ，ＮＡＤＨ０．２６ｍｍｏｌ／Ｌ，ＧＤＨ≥１０００Ｕ／Ｌ，ＴＰＩ≥１５００Ｕ／Ｌ，ＬＤ≥１０００Ｕ／Ｌ。最适ｐＨ在７．８～８．０，Ｋｍ为７．２×１０－３ｍｍｏｌ／Ｌ。批内ＣＶ：酶活性在７．３４Ｕ／Ｌ和６５．０６Ｕ／Ｌ时，ＣＶ分别为５．７％和１．４％；批间ＣＶ：酶活性在１１．８９Ｕ／Ｌ和１００．０８Ｕ／Ｌ时，ＣＶ分别为６．０％和３．３％，酶活性线性范围至少可达１８０Ｕ／Ｌ。健康人６０名，ＡＬＤ活性为４．５３±１．１７（ｘ±ｓ）Ｕ／Ｌ，男女两组均值无显著性差异。本文对ＴＥＡ－ＨＣｌ（ｐＨ８．０）、Ｔｒｉｓ－ＨＣｌ（ｐＨ８．０）和Ｃｏｌｉｄｉｎｅ－ＨＣｌ（ｐＨ７．５）三种缓冲液用于ＡＬＤ活性测定效果进行了评价，结果表明在ＴＥＡ缓冲液中所测ＡＬＤ活性最高；ＴＥＡ和Ｃｏｌｉｄｉｎｅ两种缓冲液的浓度在２５～１５０ｍｍｏｌ／Ｌ范围内对ＡＬＤ活性无影响，Ｔｒｉｓ缓冲液在５０ｍｍｏｌ／Ｌ时测得酶活性较高，缓冲液浓度过高或过低，酶活性均有所下降     

HCV在肝病患者中的感染模式及其免疫状况研究

对４１６例病毒性肝炎患者进行了抗－ＨＣＶ及其它血清标志物检测，结果共发现５２例抗－ＨＣＶ阳性患者，其中单独抗－ＨＣＶ阳性占２３．０％，与ＨＢＶ感染占４０．４％，与ＨＢＶ、ＨＤＶ感染占９．６％。抗－ＨＣＶ在慢性活动型肝炎、肝硬化中的阳性率分别为３０．４％和２２．７％，明显高于急性肝炎和非甲非乙型肝炎中的阳性率（２．８％、１３．０％），提示ＨＣＶ在促使肝病慢性化及加重病情方面有重要作用。对抗－ＨＣＶ阳性患者外周血Ｔ淋巴细胞分型结果显示：各型丙肝息者ＣＤ＿３、ＣＤ＿４比例均下降，而ＣＤ＿８比例上升，表现为Ｔｈ细胞功能下降，Ｔｓ细胞功能增强，提示细胞免疫功能紊乱是丙肝患者高慢性化率的免疫学基础。     

血清和尿镁的酶法测定

作者报告了一种测定血清和尿镁的方法，该方法只用一种酶－异柠檬酸脱氢酶（ＮＡＤＰ＾＋），该反应需镁离子作激动剂，在酶的作用下，异柠檬酸使ＮＡＤＰ＾＋减少，增加了反应速率，在有适当浓度的金属螯合物ＥＤＴＡ和ＧＥＤＴＡ存在下，镁浓度在２０ｍｍｏｌ／Ｌ内呈线性，批内ＣＶ（ｎ＝２０）和批间ＣＶ（ｎ＝１０）分别为≤１．５％和≤２．６％，回收率为９６％￣１００％，该法不受胆红素、血红蛋白和脂血的干扰。该法（Ｙ）     

输血后丙型肝炎患者不同功能区抗体的研究

为了了解丙型肝炎病毒（ＨＣＶ）不同功能区抗体的诊断价值，利用体外表达的ＨＣＶＣ、Ｅ１、Ｅ２／ＮＳ１、ＮＳ３、ＮＳ５蛋白作为重组抗原，按ＥＬＩＳＡ方法检测了已确诊为输血后丙型肝炎患者的血清１１２份。结果，检出率最高的是抗ＨＣＶ－Ｃ（８１．３％），最低的是抗ＨＣＶ－Ｅ１（１０．７％），输血后１个月内检出率最高的也是抗ＨＣＶ－Ｃ（８０％）。表明Ｃ蛋白是进行ＨＣＶ诊断的最重要抗原。另外，动态观察１０例输血     

抗HCV ELISA精密度问题的探讨

采用三种不同方法：（１）国产试剂微板ＥＬＩＳＡ,（２）进口试剂微板ＥＬＩＳＡ,（３）全自动ＥＬＩＳＡ分析法同时检测人血清中抗ＨＣＶ抗体,结果批内变异系数ＣＶ％分别为１０．３％,５．９％和２．６％。自动分析法由于减少了手工操作的误差,精密度明显高于另外二法。用（１）法测定了三种不同稀释度的抗ＨＣＶ阳性血清,结果Ａ值越小,ＣＶ越大,Ａ值在阳性判断值附近同一标本可得到阳性或阴性结果,在实际检测中应予注意     

脂蛋白电泳结合酶显色测定血清各脂蛋白中甘油三酯及临床应用

采用琼脂糖脂蛋白电泳分离脂蛋白结合甘油三酯（Ｔｇ）酶试剂呈色，扫描出各种脂蛋白区带中Ｔｇ所占比例，同血清总Ｔｇ浓度相乘计算各脂蛋白组份中Ｔｇ的含量。结果示本法与参考法比较相关系数：ＨＤＬ－Ｔｇ、ＶＬＤＬ－Ｔｇ、ＬＤＬ－Ｔｇ分别为０．８７３、０．９５５、０．９１２。方法的批内ＣＶ：ＨＤＬ－Ｔｇ、ＶＬＤＬ－Ｔｇ、ＬＤＬ－Ｔｇ分别为８．０％～１０．２％、３．８％～６．５％、４．３％～７．０％；批间ＣＶ分     

戊型肝炎患者血清IgG,IgM抗HEV动态变化

目的：了解ＩｇＧ、ＩｇＭ抗ＨＥＶ在戊型肝炎患者的动态变化规律，探讨其临床意义。方法：采用酶联免疫吸附试验（ＥＬＩＳＡ）检测５２例散发性戊型肝炎患者的ＩｇＧ、ＩｇＭ抗ＨＥＶ系列血清。结果：ＩｇＧ抗ＨＥＶ于发病后第５天即可阳转，于发病后第１５￣２９天和６１￣１２０天累积阳转率分别为８２％和１００％，多数病人ＩｇＧ抗ＨＥＶ持续时间较短，发病后１￣３０天和１２１￣１８０天分别有７．７％和７０．０％的病人阴     

急性出血性脑血管病并发多脏器功能失常综合征患者血浆内皮素-1及降钙素基因相关肽水平的观察

目的：探讨血浆内皮素１（ＥＴ１）和降钙素基因相关肽（ＣＧＲＰ）在急性出血性脑血管病（ＡＨＣＶＤ）并发多脏器功能失常综合征（ＭＯＤＳ）发病中的作用。方法：采用放射免疫法分别测定２１例ＡＨＣＶＤ合并ＭＯＤＳ患者（ＭＯＤＳ组）、２０例ＡＨＣＶＤ患者（ＡＨＣＶＤ组）及３０例正常人（正常对照组）血浆中ＥＴ１和ＣＧＲＰ水平。结果：ＭＯＤＳ组及ＡＨＣＶＤ组血浆ＥＴ１水平明显高于正常对照组（Ｐ均＜０．０１），ＭＯＤＳ组ＥＴ１水平又明显高于ＡＨＣＶＤ组（Ｐ＜０．０１）。ＡＨＣＶＤ组血浆ＣＧＲＰ水平高于正常对照组，但无显著性差异（Ｐ＞０．０５）。而ＭＯＤＳ组血浆ＣＧＲＰ水平明显低于正常对照组，ＥＴ１／ＣＧＲＰ（Ｅ／Ｃ）比值明显高于ＡＨＣＶＤ组及正常对照组（Ｐ均＜０．０１）。结论：血浆ＥＴ１水平升高、ＣＧＲＰ水平降低、Ｅ／Ｃ比值严重失衡与ＭＯＤＳ的发生相关；检测血浆ＥＴ１和ＣＧＲＰ水平对评估ＡＨＣＶＤ患者预后有一定意义     

高HDL—C个体CETP基因14内含子＋1位G：A突变的检测及其临床意义

用聚合酶链反应-限制性内切酶酶切片断长度多态性分析的方法检测高ＨＤＬ－Ｃ（ＨＤＬ－Ｃ≥１．７ｍｍｏｌ／Ｌ）个体胆固醇酯转移蛋白（ＣＥＴＰ）基因１４内含子＋１位Ｇ：Ａ突变的阳性率。共检测５０例高ＨＤＬ－Ｃ个体,发现杂合子１２例,纯合子１例,而对照组７０例中无一例发生突变、据此认为ＣＥＴＰ基因１４内含子＋１位Ｇ：Ａ突变是导致高ＨＤＬ－Ｃ的重要因素。     

Contribution of glycaemic control, endogenous lipoproteins and cholesteryl ester transfer protein to accelerated cholesteryl ester transfer in IDDM

Abstract In an earlier study we demonstrated that the transfer of cholesteryl ester (CET) estimated as the net mass of CE lost from HDL to the apoB-containing lipoproteins (VLDL + LDL) during incubation of plasma is accelerated in normolipidaemic patients with insulin-dependent diabetes mellitus (IDDM). Recombination experiments with isolated lipoprotein fractions employing this same mass transfer assay indicated that this disturbance resulted from dysfunction of VLDL and not from changes in the activity of CE transfer protein (CETP). In this study, we sought first to determine whether CET estimated with an isotopic method that measures the transfer of radiolabelled CE from exogenous HDL from non-diabetic controls to endogenous VLDL + LDL was also increased in IDDM and, if so, the extent to which this disturbance was affected by glycaemic control, VLDL and CETP. As observed with the mass transfer assay, the rate of transfer of the HDL-CE label to VLDL + LDL was also significantly accelerated in IDDM plasma (IDDM: k = 0·256±0·07; control: k = 0·092±0·05; mean±SD; P < 0·001). Fasting glucose and fructosamine correlated with both isotopic transfer (k) (r= 0·54, P= 0·009; r= 0·57, P= 0·005, respectively) and the mass of CE transferred at 2 h (r= 0·55, P= 0·006; r= 0·59, P= 0·004, respectively). Recombination experiments revealed that isotopic CET was accelerated when: (a) IDDM VLDL were combined with controls HDL and d > 1·21 fractions; and (b) IDDM d > 1·21 plasma fractions containing CETP were combined with controls VLDL + LDL and HDL. While CETP concentrations in a subset of the study group were higher in the diabetic than in the non-diabetic controls, the difference was not statistically significant (IDDM 2·25±0·97 vs. control 1·58±0·58 μg ml^-1; mean±SD; P<0·1). These findings indicate that dysfunction of VLDL and increased CETP concentrations both contribute to the pathological acceleration of isotopic transfer in IDDM plasma and that the magnitude of this proatherogenic defect correlates closely with glycaemic control.     

Accelerated cholesteryl ester transfer in patients with insulin-dependent diabetes mellitus

Abnormalities in cholesteryl ester transfer (CET) may play a role in the development of diabetic arterial vascular complications. To assess this important step systematically in reverse cholesterol transport, we have studied 20 treated, clinically stable, normolipidaemic patients. Contrary to the impairment in CET described previously in NIDDM, the mass of CE transferred from HDL to VLDL + LDL was significantly greater in IDDM patients than in controls at 1,2, and 4 h (P less than 0.001). When the d less than 1.063 plasma fractions from IDDM subjects were combined with controls d less than 1.063 fractions, an accelerated CET response was observed which was identical to that found in intact IDDM plasma. This finding, which indicates that this disturbance in CET was associated with the acceptor lipoproteins, was confirmed when we found that it was reproduced by the addition of IDDM VLDL and not LDL to control d greater than 1.063 fractions. Changes observed in lipoprotein core lipid composition were consistent with accelerated CET occurring in IDDM in vivo: the TG/CE core lipid ratio was decreased in VLDL from six subjects (diabetic 9.5 +/- 0.8 vs control 12.9 +/- 3.4; P less than 0.1) and increased in their HDL (diabetic 0.55 +/- 0.11 vs control 0.42 +/- 0.04; P less than 0.025). No correlation was demonstrable between estimates of diabetic control (glycoalbumin, fasting glucose) and CET. These data indicate that CET may be abnormally increased in normolipidaemic IDDM patients. A defect of this type may be atherogenic because it increases the number of lipoprotein particles in plasma which resemble cholesteryl ester-enriched chylomicron and VLDL remnants but whose normal receptor-mediated catabolism may be altered.     

新生儿血清胆固醇酯转运蛋白浓度及其与LDL组成的关系

目的分析新生儿血清胆固醇酯转运蛋白(CETP)浓度与LDL组成的关系,以及对血脂、脂蛋白水平的影响。方法采用ELISA法测定40例脐带血CETP浓度,并与80例健康成人比较;用序列超速离心法分离LDL后再用常规方法分析其中TG、CE含量。结果新生儿CETP呈偏态分布、中位数0.20 mg/L,范围(0.11~0.49)mg/L,均值(0.25±0.11)mg/L,明显低于成人(2.19±2.04)mg/L(P<0.0001);男女间无显著差异[(0.26±0.10)mg/L vs(0.22±0.16)mg/L,P>0.05]。新生儿血脂、脂蛋白浓度均低于成人,LDL-CE/LDL-TG比值也明显低于成人(1.5 vs 4.7)。结论新生儿血清CETP浓度仅是成人的九分之一,低CETP水平可能是新生儿LDL-CE/LDL-TG比值低的主要原因。     

慢性肾衰透析患者胆固醇酯转运蛋白及血脂变化

本文检测 77例慢性肾衰血透患者 (HD)的胆固醇酯转运蛋白 (CETP)和血脂 ,结果显示 HD患者较对照组的 CETP、TC、HDL - C、apo AI明显降低 ,TG明显升高。按 TG分组 ,高 TG组和正常 TG组 HD患者 CETP仍显著低于对照组 ,但高 TG组 HD较正常 TG组 HD患者 TC、apo B明显升高 ,apo AI、HDL - C下降。CETP和血脂异常与 HD患者高发心、脑血管疾病密切相关 ,其发病机理需进一步探讨。     

胆固醇酯转运蛋白的提纯和鉴定

目的建立一种新的分离纯化人血清胆固醇酯转运蛋白（ＣＥＴＰ）的方法,为制备ＣＥＴＰ单克隆抗体和ＣＥＴＰ浓度测定提供物质基础。方法制备无脂血清,用ＢｕｔｙｌＳｅｐｈａｒｏｓｅ４ＦＦ、ＣＭＳｅｐｈａｒｏｓｅＦＦ和ＳｅｐｈａｃｒｙｌＳ２００三步柱层析分离、纯化ＣＥＴＰ。结果纯化ＣＥＴＰ经ＳＤＳＰＡＧＥ电泳鉴定呈单一区带,分子量６５０００,蛋白质含量０３２５ｍｇ／ｍｌ,与国外和作者研制ＣＥＴＰ单抗的免疫斑点试验呈特异性反应,与纯化脂蛋白（ａ）、载脂蛋白（Ａｐｏ）Ａ１、ＡｐｏＢ、ＩｇＧ、ＩｇＡ、ＩｇＥ、Ｃ３、Ｃ４抗体无免疫反应。结论经三步柱层析获得了纯化ＣＥＴＰ,得率约２５％,活性每小时３０ｎｍｏｌ／μｇ。     

Effect of hypoalbuminemia on the increased serum cholesteryl ester transfer protein concentration in children with idiopathic nephrotic syndrome

OBJECTIVES: To examine the alteration of cholesteryl ester transfer protein (CETP) mass with the regression of albumin level in childhood nephrotic syndrome (NS) in order to clarify the effect of albumin on CETP in NS. DESIGN AND METHODS: Serum concentrations of CETP, kidney parameters and lipid traits were determined in 110 children with idiopathic NS and 150 control subjects. Of the NS patients, 69 children with an active phase formed group 1, and 41 in remission formed group 2. RESULTS: Group 1 presented severe hypoalbuminemia and hyperlipidemia, while group 2 exhibited marked recovery in both serum albumin level and lipid/lipoprotein profile. CETP concentration was significantly higher in group 1 (7.36+/-2.43 mg/L, compared with controls 3.38+/-1.83 mg/L, P<0.0001), and declined to within normal range in group 2 (2.91+/-1.77 mg/L). CETP concentration had a strong inverse correlation with serum albumin level (r=-0.688, P<0.0001) in NS patients. Furthermore, when multiple linear regression analysis was performed, in which albumin, proteinuria, lipid traits, and prednisone dose were treated as independent variables, albumin was the only variable showing a significant correlation with CETP in the NS patients (R(2)=0.587, beta=-0.475, P<0.0001). CONCLUSIONS: The results demonstrate that the decreased serum albumin level might be a main determinant of the increased CETP concentration in pediatric NS.