肿瘤坏死因子放射免疫分析的建立及初步临床应用

用人重组肿瘤坏死因子（ＴＮＦ－α）和特异性兔抗ＴＮＦ血清建立了液相竞争放射免疫分析，最小检出值＞３０ｐｓ／管，标准曲线范围０．３～２４．３ｎｇ／ｍｌ，批内和批间变异系数平均为７．８％和８．４％。本法检测３７名健康献血员血清ＴＮＦ为０．８３±０．２６ｎｇ／ｍｌ，２３例急性黄疸性肝炎、１３例慢性迁延性肝炎、１９例慢性活动性肝炎、２２例肝硬化、２５例肝癌血清ＴＮＦ分别为１．９５±０．５９ｎｇ／ｍｌ，２，０６±０．５１ｎｇ／ｍｌ，１．５８±０．５４ｎｇ／ｍｌ，１．７５±０．６５ｎｇ／ｍｌ和１．５７±０．５３ｎｇ／ｍｌ。２８例肺癌术前血清ＴＮＦ０．９２±０．４２ｎｇ／ｍｌ，２２例肺癌术后血清ＴＮＦ为１．０８±０．２９ｎｇ／ｍｌ。     

病毒性肝炎患者白细胞介素－6的检测及其临床意义

应用夹心ＥＬＩＳＡ检测了５６例乙型病毒性肝炎患者外周血单个核细胞（ＰＢＭＣｓ）白细胞介素－６（ＩＬ－６）的诱生水平。乙型慢性活动性肝炎（ＣＡＨ）、乙型肝炎后肝硬化（ＨＣ）和乙型重型肝炎（ＳＨ）ＰＢＭＣｓ经脂多糖诱导后，ＩＬ－６的诱生水平分别为１２．８４±３．２６ｎｇ／ｍｌ、１１．７９±３．０１ｎｇ／ｍｌ和３８．４２±８．３７ｎｇ／ｍｌ，明显高于正常对照组（Ｐ＜０．０５～０．０１）。ＣＡＨ和ＳＨ患者的ＩＬ－６水平与肝细胞变性坏死，蛋白质合成障碍和患者的预后也有一定的关系。上述结果提示，机体内ＩＬ－６的调节紊乱可能与病毒性肝炎的发病机理有关。     

肾小管损伤时内皮素1、肿瘤坏死因子α的表达及其对离体肾间质成纤维细胞的影响

目的　研究内皮素１（ＥＴ１）、肿瘤坏死因子（ＴＮＦα） 在肾小管上皮细胞的表达、肾小管的损伤及它们对肾间质成纤维细胞的影响。方法　应用肾小管上皮细胞及肾间质成纤维细胞的体外培养;建立肾小管损伤动物模型;应用逆转录聚合酶链反应（ＲＴＰＣＲ） 、免疫组化ＳＰ法染色、放射免疫测定（ＲＩＡ） 及双重免疫组织化学染色技术,并测定３ＨＴＤＲ 掺入率。结果　肾小管上皮细胞既有ＥＴ１ｍＲＮＡ和ＴＮＦαｍＲＮＡ 的表达（ＲＴＰＣＲ的扩增片段分别为５４６ ｂｐ 和４１５ ｂｐ） ,还有ＥＴ１ 及ＴＮＦα的蛋白合成及分泌;细胞培养上清中ＥＴ１ 和ＴＮＦα含量分别为１．４２ ｐｇ／ｍｌ 和０ ．５８ ｎｇ／ｍｌ;且肾小管损伤及再生过程中,ＥＴ１ 及ＴＮＦα表达增加。对体外培养的大鼠肾间质成纤维细胞分别加入ＥＴ１ 、ＴＮＦα,３ＨＴＤＲ掺入率较对照组明显增高（ Ｐ＜ ０．０５）;并证明肾间质成纤维细胞有ＥＴ受体（ＥＴＲ） ｍＲＮＡ及ＴＮＦ受体（ＴＮＦＲ１） ｍＲＮＡ的表达（ＲＴＰＣＲ 扩增片段分别为５４４ ｂｐ 和３４７ ｂｐ）。结论　损伤及再生的肾小管上皮细胞较正常时合成和分泌更多的ＥＴ１ 和ＴＮＦα,ＥＴ１ 及ＴＮＦα分别     

慢性肾炎患者sIL－2R水平、IL－2活性及mIL－2R表达的观察

本文应用ＥＬＩＳＡ法检测了４０例慢性肾小球肾炎血清可溶性白细胞介素２受体水平，同时对患者外周血单个核细胞膜白细胞介素２受体表达及白细胞介素２活性进行观察。结果患者ｓＩＬ－２Ｒ水平为６３４．８±１４２．９ｕ／ｍｌ，高于正常人２９５．０±１６５．７ｕ／ｍｌ，Ｐ＜０．００１；ｍＩＬ－２Ｒ阳性率为２５．６±４．３％，低于正常人４５．５±５．２％，Ｐ＜０．００１；ＩＬ－２活性为２．８５±１．６１ｕ／ｍｌ，低于正常人７．０６±４．５３ｕ／ｍｌ，Ｐ＜０．００１。并且ｓＩＬ－２Ｒ与ＢＵＮ呈正相关，ｒ＝０．４７０，Ｐ＜０．０２。提示慢性肾小球肾炎患者细胞免疫功能降低，且与肾功能损伤程度有关。     

乙型肝炎患者红细胞CR1密度相关基因多态性与CR1活性的相关性

为研究乙型肝炎患者红细胞ＣＲ１活性与ＣＲ１密度相关基因多态型别的关系,采用ＰＣＲ加ＨｉｎｄⅢ酶切技术测定红细胞ＣＲ１密度相关基因多态型别（ＨＨ、ＨＬ、ＬＬ型）,采用红细胞ＣＲ１（ＥＣＲ１）免疫粘附活性（ＲＣＩＡ）和粘附ＩＣ量免疫酶联（ＥＬＩＳＡ）法测定红细胞ＣＲ１活性。乙型肝炎患者ＨＨ型比率是７３９％（３４／４６）,正常人ＨＨ型比率是８０％（６４／８０）,ＨＨ型乙型肝炎患者ＥＣＲ１ＲＣＩＡ（３７８±０２９ｎｇＡＨＧ／１０７ＲＢＣ）明显高于ＨＬ型乙型肝炎患者（１８８±０２６ｎｇＡＨＧ／１０７ＲＢＣ）和ＨＨ型正常人（２６０±０３０ｎｇＡＨＧ／１０７ＲＢＣ）。提示乙型肝炎患者红细胞免疫功能低下是获得性的。     

反复呼吸道感染儿sIL－2R和T细胞亚群的测定

本文采用双抗体夹心ＥＬＩＳＡ法分别测定了２１例反复呼吸道感染儿，３０例正常儿童，１０例新生儿脐血的血清可溶性白细胞介素２受体（ｓＩＬ＝２Ｒ）水平。结果患儿组ｓＩＬ－２Ｒ为７１６．６０±３０．１０Ｕ／ｍｌ；正常儿童组为３８４．４７±８８．０３Ｕ／ｍｌ（ｐ＜０．０１）；新生儿脐血为４４６．２０±５５．６８Ｕ／ｍｌ，与正常儿童比较Ｐ＞０．０５。同时采用间接免疫荧光技术测定了患儿Ｔ细胞亚群水平，结果ＣＤ８细胞数升高，ＣＤ３细胞和ＣＤ４细胞数、ＣＤ４／ＣＤ８比值下降，与正常儿童比较有显著性差别。提示反复呼吸道感染儿有细胞免疫功能降低及免疫调节紊乱。     

人甲胎蛋白时间分辨免疫荧光分析法的建立

本文应用兔抗ｈＡＦＰ ＩｇＧ包被板条，以异硫氰酸苯基－ＥＤＴＡ－Ｅｕ＾３＋标记抗ｈＡＦＰ ＭｃＡｂ，建立了固相“夹心”二步法分析ｈＡＦＰ技术，结果显示，分析范围是６．２５ｎｇ／ｍｌ－１００ｎｇ／ｍｌ（ｒ＝０．９９），变异系数在１．６７％－１７．７４％之间，ｈＡＦＰ最小可测值为０．３５ｎｇ／ｍｌ。测定７６例孕妇血清样品，同所购ＤＥＬＦＩＡ ｈＡＦＰ最小可测值为０．３５ｎｇ／ｍｌ。测定７６例孕妇血清样     

IgG-RF ELISA定量检测法的建立和应用

本工作以 Ｉｇ Ｇ Ｆｃ 片断为靶抗原, 用自制抗人 Ｉｇ Ｇ Ｆａｂ Ｈ Ｒ Ｐ 标记物为第二抗体, 建立了定量测定血中 Ｉｇ Ｇ Ｒ Ｆ 的 Ｅ Ｌ Ｉ Ｓ Ａ。经验证, 本法可测范围为１０ ～２５６０ Ｉ Ｕ／ｍｌ, 批内 Ｃ Ｖ４８ ％ , 批间 Ｃ Ｖ１１７ ％ 。１００ 例健康人血清 Ｉｇ Ｇ Ｒ Ｆ 值为（４１０ ±３４３ ） Ｉ Ｕ／ｍｌ, 以高于珋ｘ ±ｓ 值的上限１１０ Ｉ Ｕ／ｍｌ 为阳性。９９ 例类风湿关节炎（ Ｒ Ａ） 患者血清 Ｉｇ Ｇ Ｒ Ｆ 值为（４３０ ±６０４） Ｉ Ｕ／ｍｌ,阳性率为６１５ ％ 。临床应用提示 Ｉｇ Ｇ Ｒ Ｆ 是判断 Ｒ Ａ 活动性的良好指标之一。     

糖尿病患者红细胞钙含量与红细胞变形性

用火焰原子吸收法测定Ⅱ型糖尿病（ＮＩＤＤＭ）２４例红细胞钙总量及用激光衍射法测定其红细胞变形性（ＲＣＤ）。测定５０例ＮＩＤＤＭ之红细胞在１５０、２００、２５０、３００及３５０ｍｍｏｌ／Ｌ５种渗透压悬液中的变形性。结果表明，正常成人红细胞钙总量为３３．６２±１２．８０μｍｏｌ／Ｌ，ＲＣＤ降低的ＮＩＤＤＭ患者，其红细胞钙总量明显升高（４６．３７±２３．３７μｍｏｌ／Ｌ，Ｐ＜０．０５），而ＲＣＤ正常之患者则接近正常水平。不同渗透压悬液中的最大ＤＩ值（ＤＩｍａｘ）显示，ＲＣＤ降低之ＮＩＤＤＭ红细胞在２５０及３００ｍｍｏｌ／Ｌ的ＤＩｍａｘ显著减小（Ｐ＜０．００１）；而在１５０及２００ｍｍｏｌ／Ｌ的低渗范围，其ＤＩｍａｘ接近正常，符合内粘度加大的ＤＩｍａｘ渗透压曲线特征。     

病毒性肝炎患者IL—1,IL—6和INFa活性的检测

检测了甲，乙型病毒性肝炎患者外周血单个核细胞ＩＬ－１、ＩＬ－６和ＴＮＦａ的诱生活性及其血清中活性。结果表明，乙型慢性活动性肝炎、乙型肝炎后肝硬化和乙型生型肝炎ＰＢＭＣｓ经脂多糖诱导后，ＩＬ－１活性分别为３５３１．１±８８２．７Ｕ／ｍｌ，２７６９．７±７３０．４±Ｕ／ｍｌ和５３２９．３±１０８９．３Ｕ／ｍｌ，高于正常对照组（Ｐ＜０．０５）或＜０．０１）；ＩＬ－６诱生活性分别为３８．９０±１４．７５Ｕ     

肾活检组织中NEP的表达及其与尿NEP含量之间的相关性研究

目的 观察肾活检组织中NEP的表达,研究它与尿NEP含量之间的关系,以期探讨检测尿NEP的临床意义。方法 将研究组分为对照组(n=100),肾小球疾病组(n=31)、急性肾小管损伤组(n=44)、慢性肾小管损伤组(n=61)、慢性肾功能衰竭组(n=13)。收集各组病人晨尿,然后通过荧光光谱分析,得到尿中NEP的量,并用相应尿肌酐值予以标化。同时对病人组中的68例患者的肾组织切片用免疫组化的方法进行染色,直接观察NEP的在肾组织中的表达情况,且进行其表达量与尿NEP的相关性研究。结果 正常对照组尿NEP为68.41ug/mmol Cr,急性肾小管损伤组尿NEP196.36ug/mmol Cr,明显高于正常对照组(P=0.0001),慢性肾小管损伤组、肾小球疾病所致的CRF组尿NEP分别为31.98、19.40ug/mmol Cr,均明显低于正常对照组(P均<0.01),而单纯肾小球疾病组尿NEP为75.49ug/mmol Cr,与正常对照组无差异(P=0.1425)。在急性肾小管损伤组和慢性肾小管损伤组,尿NEP与肾组织中NEP的表达均呈正相关,而肾小球疾病组内尿NEP与肾组织中NEP的表达无明显相关性。结论 肾小管刷状缘上NEP的表达量与近端小管损伤有良好相关性,尿NEP量实际反映近端小管刷状缘损伤情况。本研究在国内率先建立了尿NEP的检测方法,并用于临床研究。对尿中NEP的检测,提供了一种快速、非损伤性测量手     

Beta 2-glycoprotein-1 (apolipoprotein H) excretion and renal tubular malfunction in diabetic patients without clinical proteinuria.

AIM--To compare the urinary excretion of beta 2-glycoprotein-1 with that of two other markers of early tubular disorder in diabetic patients without clinical proteinuria. METHODS--The urinary excretion of retinol binding protein, beta 2-glycoprotein-1, and N-acetyl-beta-D-glucosaminidase was measured in 90 known diabetic patients who had a negative reagent strip test for proteinuria. RESULTS--Among 43 patients with urinary albumin excretion within the reference range, 23 (53%) had raised urinary N-acetyl-beta-D-glucosaminidase activity, five (12%) increased excretion of beta 2-glycoprotein-1, and five (12%) increased loss of retinol binding protein. Among 47 patients with an albumin excretion of 0.9-7.9 mg/mmol creatinine, 42 (89%) had increased urinary N-acetyl-beta-D-glucosaminidase, 23 (49%) an increased output of beta 2-glycoprotein-1, and 16 (34%) a raised excretion of retinol binding protein. The excretion of these markers of tubular defects seldom exceeded two and a half times the upper reference limit and the differences between the findings in the insulin dependent and non-insulin dependent patients with similar albumin excretion were small and insignificant. CONCLUSIONS--In diabetic patients with a negative dipstick test for proteinuria: (a) assay of urinary beta 2-glycoprotein-1 may be a more sensitive test for the detection of impaired tubular reabsorption of protein than measurement of retinol-binding protein; (b) assay of N-acetyl-beta-D-glucosaminidase can detect tubular injury at a time when protein reabsorption remains normal; and (c) impaired renal tubular function may be present in the absence of evidence of glomerular malfunction.     

Renal tubular impairment in children with idiopathic hypercalciuria

Idiopathic hypercalciuria (IH) is defined as hypercalciuria that persists after correction of dietary inbalances and has no detectable cause. The excretion of urinary N-acetyl-beta-D-glucosaminidase (U-NAG), a marker of proximal tubular damage, has been previously reported as either increased or normal in children with IH. We evaluated U-NAG in 20 children (13 boys and 7 girls, mean age 10.3 years +/- 5.7 SD) with IH (urinary calcium excretion above 0.1 mmol/kg/24 hours, with no detectable cause) and with otherwise normal renal function tests. Ultrasound examination revealed urolithiasis (n=4) and nephrocalcinosis (n=1). The U-NAG values were evaluated in the spot urine collected from the second morning void and calculated as the urinary NAG/creatinine ratio (U-NAG/Cr) and expressed in nkat/mmol. The 24-hour urinary calcium excretion (U-Ca/24h) was assessed in a urinary sample from 24-hour collected urine and calculated in mmol/kg. The obtained results of U-Ca/24h and U-NAG/Cr were expressed as Z-scores. When compared to the reference data, the U-Ca/24h and U-NAG/Cr were significantly higher (p = 0.0004 and p = 0.006, respectively). There was no correlation between the U-NAG/Cr and U-Ca/24h (r = 0.18, p = 0.20). The U-NAG/Cr values were significantly higher in the 5 patients with urolithiasis/nephrocalcinosis, whether compared to the rest of the group (p = 0.02), or to the reference data (p = 0.01). The U-NAG/Cr activity was higher in 15 children without urolithiasis/nephrocalcinosis when compared to reference data (p < 0.01). There was no difference in U-Ca/24h between the children with and without urolithiasis/nephrocalcinosis (p = 0.58). These findings suggest that tubular impairment, as reflected by U-NAG/Cr, might occur in children with IH, especially in patients with urolithiasis/nephrocalcinosis. There doesn't seem to be a direct relationship between the U-NAG/Cr activity and the degree of calcium leakage.     

Urine laminin and kallikrein, markers of tubulointerstitial damage in experimental protein overload on pre-existing renal damage

We studied the response of urinary protein overload on preexisting tubulointerstitial nephritis (TIN), which was induced in male Sprague Dawley rats by hexachloro-1,3-butadiene (HCBD). Five days after the development of TIN, puromycin aminonucleoside (PAN) was administered to induce urinary protein overload. Urinary laminin and kallikrein were measured. Urine specimens were collected daily for 14 days and on day 21; and tissue specimens were collected on days 1, 4, 7, 10, 14 and 21. Urinalysis was correlated with the renal pathology at the light microscopic level. Laminin excretion was increased on day 4; one day before total protein, indicating damage to the basement membrane. Kallikrein levels also fell early indicating distal tubular damage. There is clear evidence that urine protein overload in a previously damaged kidney with tubulointerstitial injury leads to accelerated and more severe renal damage. Laminin and kallikrein are early and sensitive markers of renal injury.     

肾脏病患者尿液中SIgA的含量的观察

用放射免疫(双抗体-PEG法)测定109例正常人和85例各种类型肾脏病患者尿液中SIgA含量。表明慢性肾炎普通型、慢性肾炎肾病型活动期和慢性肾炎所引起的慢性肾功能不全均较正常对照组明显增高。慢性肾炎肾病型好转期尿 SIgA与正常对照组无显著差异,但较活动期明显减少。高分子和混合蛋白尿患者尿 SIgA较中分子和低分子蛋白尿患者明显增加。肾病患者24小时尿蛋白排出量与尿 SIgA呈正相关,而与血清 IgA无相关性。文中讨论了非感染性肾脏疾病时尿 SIgA增高的可能原因。     

A Case Report of a Middle East Respiratory Syndrome Survivor with Kidney Biopsy Results

A 68-year old man diagnosed with Middle East Respiratory Syndrome-Coronavirus (MERS-CoV) presented with multiple pneumonic infiltrations on his chest X-ray, and the patient was placed on a mechanical ventilator because of progressive respiratory failure. Urinary protein excretion steadily increased for a microalbumin to creatinine ratio of 538.4 mg/g Cr and a protein to creatinine ratio of 3,025.8 mg/g Cr. The isotope dilution mass spectrometry traceable serum creatinine level increased to 3.0 mg/dL. We performed a kidney biopsy 8 weeks after the onset of symptoms. Acute tubular necrosis was the main finding, and proteinaceous cast formation and acute tubulointerstitial nephritis were found. There were no electron dense deposits observed with electron microscopy. We could not verify the virus itself by in situ hybridization and confocal microscopy (MERS-CoV co-stained with dipeptidyl peptidase 4). The viremic status, urinary virus excretion, and timely kidney biopsy results should be investigated with thorough precautions to reveal the direct effects of MERS-CoV with respect to renal complications.     

111例肾脏及泌尿道疾病尿β_2微球蛋白测定的初步观察

本文报道应用酶联免疫抑制法对111例肾脏及泌尿道疾病进行尿β_2微球蛋白测定,并与常规肾功能试验对照比较,并初步探讨了该项检查的临床意义。     

Induction of interstitial nephritis in rats by basement membrane of human origin

A study was conducted to investigate nephritogenic tubular basement membrane antigens common to human and rat kidneys. Brown Norway (BN) rats were immunized with human renal basement membrane in complete Freund's adjuvant simultaneously with Bordetella pertussis vaccine. The immunized rats developed polyuria and increased levels of serum creatinine one week after the second immunization. Renal histology at this time revealed marked, acute tubulointerstitial nephritis with linear deposition of IgG and C3 along the tubular basement membrane and Bowman's capsule, but not along the glomerular basement membrane. Rats with this tubulointerstitial nephritis rapidly developed antibodies against renal antigens from normal BN rats such as tubular basement membrane and proximal tubule brush border, however antibodies to glomerular basement membrane appeared later. Western blotting using the same rat sera detected a 145-kDa antigen from 8 M urea-solubilized human renal basement membrane and 120-kDa, 135-kDa and 145-kDa antigens from 8 M urea-solubilized BN rat renal basement membrane. This suggests that renal basement membranes of human and rat origin have common antigens involved in the pathogenesis of tubulointerstitial nephritis.     

尿微量白蛋白ELISA测定法及其临床应用

定期监测糖尿病患者的尿微量白蛋白,对尚处于可逆性阶段糖尿病性肾病的早期诊断具有重要价值。尿白蛋白也是高血压病引起血管损伤及肾血管早期病变的敏感指标。我们自制高效价抗人Alb血清、抗(人Alb)IgG-HRP酶标记结合物和抗(人Alb)IgG包被微孔滴定板,建立了一种灵敏、简便的尿微量白蛋白酶联免疫测定法,测定范围为5～500ng/ml;灵敏度0.76ng;批内CV％ 6.4％;批间CV％ 16.4％。24小时尿白蛋白排泌量,正常人为8.86(SD 9.23)mg/24h(n=80);糖尿病患者为40.4(SD 40.4)mg/24h(n=16);Ⅰ、Ⅱ、Ⅲ期高血压病患者分别为15.17(SD 14.42)mg/24h、21.46(SD 24.34)mg/24h和24.6(SD 18.9)mg/24h。     

Urinary dopamine excretion in healthy volunteers: effect of sodium diet and acute water load

The present study was designed to investigate, in human subjects, urinary dopamine excretion under different conditions of sodium and water homeostasis. In a cross-over trial, ten healthy volunteers were subjected to low-salt (LS; dietary salt restriction, sodium chloride (NaCl) intake <5 g per day), normal-salt (NS; normal food ad libitum), and high-salt (HS; normal food plus NaCl 100 mg/kg per day) regimens for 8 days in a randomized order. On day 7, urine was collected for 24 h. The variations in urinary sodium excretion reflected the dietary salt intake (LS: 16.3+/-4.7; NS: 144.1+/-18.2; HS: 221.9+/-12.9 mmol 24 h(-1) 1.73 m(-2)), but were not accompanied by significant changes in urinary dopamine excretion. On day 8, clearance studies showed that an acute oral water load of 1500 ml did not alter glomerular filtration rate or renal plasma flow but significantly increased urinary flow rate without affecting dopamine excretion. Assuming that excreted dopamine is not metabolized or reabsorbed during the tubular passage, both the unchanged urinary dopamine output in spite of 14-fold variations in sodium excretion and its independence of an acute water load argue against the hypothesis that dopamine in the tubular lumen acts as a natriuretic and/or diuretic factor in humans.