电镜技术在测定消毒剂对乙型肝炎病毒灭活效果中的应用

在电镜下观察了消毒剂作用后乙型肝炎病毒颗粒破坏情况，并与用ＳＰＲＩＡ法检测ＨＢｓＡｇ抗原性破坏结果作比较。结果，含二氯异氰尿酸钠、碘伏与戊二醛的３种消毒液，破坏ＨＢＶ颗粒结构的最低有效浓度均高于破坏ＨＢｓＡｇ抗原性者。     

电镜技术在测定消毒剂对乙型肝炎病毒灭活效果中中的应用

在电镜下观察了消毒剂作用后乙型肝炎病毒颗粒破坏情况，并与用ＳＰＲＩＡ法检测ＨＢｓＡｇ抗原性破坏结果作比较，结果，含二氯异氰尿酸钠，碘伏与戊二醛的３种消毒液，破坏ＨＢＶ颗粒结构的最低有效浓度均高于破坏ＨＢｓＡｇ抗原性者。     

肝组织HCV—NS3抗原与HCV感染血清不同模式的关系及临床意义

采用逆转录套式聚合酶链反应（ＲＴ－ｎｅｓｔｅｄ ＰＣＲ）和ＥＬＩＳＡ方法检测血清ＨＣＶ ＲＮＡ、抗－ＨＣＶ，采用鼠单克隆抗体ＰＡＰ方法检测肝组织内ＨＣＶ－ＮＳ３抗原。结果发现被检测的４５例丙型肝炎患者随着病情加重和病程延长，血清抗－ＨＣＶ和ＨＣＶ ＲＮＡ及肝组织ＨＣＶ－ＮＳ３抗原阳性率逐步增高；某些患者抗－ＨＣＶ和ＨＣＶ ＲＮＡ阳性结果不一致；血清ＨＣＶ ＲＮＡ阳性肝组织ＨＣＶ－ＮＳ３阳性率明显同     

静注免疫球蛋白与丙型肝炎病毒传播的安全性研究

笔者就输注国产ＩＶＩＧ后传播ＨＣＶ的危险性作了研究。２９名有ＩＶＩＧ应用指针的病儿，输注经加热法灭活病毒的ＩＶＩＧ后１周、２周、１月，抗－ＨＣＶ阳性率分别为８３．３％、４５．５％、４．６％，３个月后全部转阴，显示抗－ＨＣＶ随输注ＩＶＩＧ后时间延长而转阴。随机抽取１１名输注ＩＶＩＧ后抗－ＨＣＶ阳性病儿的２４份系列血标本，用ｎｅｓｔ－ＰＣＲ检测ＨＣＶ－ＲＮＡ，无阳性发现。检则１０份ＩＶＩＧ中抗－ＨＣＶ均阳性，抗－ＨＣＶ滴度１∶１６，抽取６份ＩＶＩＧ，ＨＣＶ－ＲＮＡ阴性。经３个月～１年观察，所有病例无肝炎的临床症状和体征，结果支持：用于本研究的国产ＩＶＩＧ输注可被动输入抗－ＨＣＶ，但不传播ＨＣＶ。     

干扰素治疗丙型肝炎的临床及抗病毒效应研究

１５例急性、５２例慢性丙肝在干扰素治疗２４周后，血清ＡＬＴ复常率为８０％与７５％，ＨＣＶ ＲＮＡ阴转率为８０％与６９．２％，停药后追踪２４周，持续ＡＬＴ正常并ＨＣＶ－ＲＮＡ阴转（称为完全反应、ＣＲ）者有５３．３％与４２．３％，均明显高于急、慢性对照组的疗效（治疗结束时ＡＬＴ复常率急性对照组为３６．４％，慢性对照组为０。ＨＣＶ－ＲＮＡ阴转为２／１１及３／２０，停药后全部对照组均无１例呈ＣＲ者），统计     

聚合酶链反应检出石蜡包埋肝癌组织的乙型肝炎病毒DNA

传统免疫组织化学方法及分子杂交检测石蜡包埋肝组织乙型肝炎病毒（ＨＢＶ）ＤＮＡ灵敏度低，聚合酶链反应（ＰＣＲ）方法能否检测到石蜡肝癌组织中的ＨＢＶＤＮＡ报道尚不多见。为此，采用ＨＢＶ与丙型肝炎病毒二合一ＰＣＲ试剂盒对１７例肝癌患者的石蜡包埋癌组织进行了ＨＢＶＤＮＡ检测，结果７例阳性，阳性率为４１％，而８例非肝癌对照患者则均阴性。组织ＨＢＶＤＮＡ结果与血清ＨＢｓＡｇ比较显示，该试剂盒有较高的检出率及符合率。这为今后进一步研究大量保存的肝病石蜡包埋组织的ＨＢＶ感染提供了一个可靠灵敏的方法，也为研究其他病毒感染的石蜡包埋材料提供了参考。     

庚型肝炎病毒不同基因区的聚合酶链反应扩增

分别在庚型肝炎病毒（ＨＧＶ）基因５′端非编码区（５′－ＵＴＲ）、非结构（ＮＳ）３及区非结构（ＮＳ）５区设计套式引物，建立逆转录套式聚合酶链反应（ＲＴ－ｎｅｓｔｅｄ ＰＣＲ）检测血清ＨＧＶ ＲＮＡ。在２９９份不同血汪标本中ＨＧＶ ＲＮＡ阳性者４１例，基于５′－ＵＴＲ、ＮＳ３区及ＮＳ５区引物ＰＣＲ的阳性率分别为８７．８％、８０．５％和９７．６％。本研究结果表明根据中国株ＨＧＶ基因ＮＳ５区部分序列设计引     

乙型肝炎血清学标志物抗力的比较

经检测，对钴－６０辐照，病人血清中ＨＢｓＡｇ、ｐＨＳＡ－Ｒ与ＨＢＶ－ＤＮＡ抗力较其中ＨＢＶ－ＤＮＡｐ、ＨＢｅＡｇ及纯化ＨＢｓＡｇ者为强．ＨＢＶ颗粒经１７．９０ｋＧｙ剂量辐照后在电镜下仅见碎片．对所试含氯消毒剂、碘酊、高锰酸钾、盐酸与过氧化物消毒剂，病人血清中的ＨＢｓＡｇ较其中ｐＨＳＡ－Ｒ、ＨＢｅＡｇ及纯化ＨＢｓＡｇ的抗力为强．     

HCV感染基因型病毒复制程度与肝病关系的研究

采用限制性内切酶Ｓａｕ３Ａｌ和ＨａｅⅢ对５４例ＲＴ－ＰＣＲ法ＨＣＶＲＮＡ阳性的５＇ＮＣ区扩增产物进行酶切分型；ＰＣＲ滴定法半定量分析ＨＣＶＲＮＡ含量。结果显示４种酶切类型：ＨＣＶⅠ型感染３．７％（２／５４），ＨＣＶⅡ型感染７７．８％（４２／５４），ＨＣＶⅢ、Ⅳ型感染１１．１％（６／５４），ＨＣＶⅠ、Ⅱ／Ⅲ、Ⅳ型混合感染７．４％（４／５４），ＨＣＶⅡ型感染阳性率在ＡＨ、ＣＡＨ、ＬＣ三组间无显著差异（     

复合PCR检测乙型肝炎和丙型肝炎病毒

设计了ＨＢＶ和ＨＣＶ各一对引物，建立了复合ＰＣＲ，并对临床标本进行检测，５例ＨＢｓＡｇ和ＨＣＶ抗体均阳性者，其ＨＢＶ－ＤＮＡ和ＨＣＶ－ＲＮＡ均阳性；２０例单ＨＢｓＡｇ阳性标本，１１例单ＨＢＶ－ＤＮＡ阳性，５例ＨＢＶ－ＤＮＡ和ＨＣＶ－ＲＮＡ均阳性；２０份单ＨＣＶ抗体阳性者，ＨＣＶ－ＲＮＡ均阳性，３例合并ＨＢＶ－ＤＮＡ阳性；１０例慢性ＮＡＮＢ抗ＨＣＶ阴性肝炎标本８份ＨＣＶ－ＲＮＡ，１例ＨＢＶ－ＤＮＡ和     

猪主动脉脱细胞血管基质制备及保存

目的探讨脱细胞血管基质的制备和保存方法。方法采用酶、低渗溶液、化学除垢剂联合法处理猪胸主动脉制备脱细胞血管基质,标本行苏木精-伊红染色,大体、光镜及扫描电镜、透射电镜观察。并将标本放入液氮中保存,于保存1、2、3月分别取标本用20g/L戊二醛固定,扫描电镜观察。结果经该法处理的猪血管细胞全部脱除,胶原纤维、弹性纤维无断裂,细胞外基质保持完好。液氮保存后1、2、3月扫描电镜观察与新鲜标本无明显差别。结论酶、低渗溶液、化学除垢剂联合法是制备脱细胞血管基质的较好方法,制得的脱细胞血管基质可放入液氮中保存。     

无水甘油保存同种异体肌腱的移植

背景：如何处理和保存同种异体肌腱,使之尽可能保留生物活性并降低其免疫原性是近年研究热点。目的：将保存于无水甘油的兔肌腱进行同种异体移植,观察移植后肌腱的组织学变化。方法：将兔肌腱经特殊处理后,常温避光保存在无水甘油中,保存7,12个月后取出肌腱,移植到兔后腿屈肌腱中。饲养3个月后处死并取出移植的肌腱,进行苏木精-伊红染色石蜡切片及透射电镜观察。结果与结论：苏木精-伊红石蜡切片示肌腱纤维排列尚整齐,细胞形态正常,切片某些区域细胞密度较正常组低,未见明显的炎性细胞浸润。透射电镜示肌腱纤维排列整齐,细胞形态正常。结果提示经无水甘油保存的肌腱进行同种异体移植后,在组织学上与正常肌腱相似,无明显的免疫排斥反应。     

甘油常温保存肌腱的两种方法比较

背景：最大程度保留肌腱的生物活性是进行同种异体肌腱移植的条件。目的：通过比较选择出最佳的肌腱常温保存方法。方法：使用无菌操作的方法将兔肌腱随机分为4组：新鲜肌腱对照组、生理盐水组、无水甘油Ⅰ组（肌腱梯度脱水后在无水甘油中保存）、无水甘油Ⅱ组（肌腱直接在无水甘油中保存）,分别于保存的第2,4,7个月进行检测。结果与结论：苏木精-伊红染色显示：无水甘油Ⅰ组肌腱细胞完整率明显高于无水甘油Ⅱ组。电镜观察发现,无水甘油Ⅰ组大部分肌腱细胞形态正常,肌腱组织结构完整;无水甘油Ⅱ组保存4,7个月后细胞核凝固、固缩、崩解状。无水甘油Ⅰ组的超氧化物歧化酶活性也明显高于无水甘油Ⅱ组。说明肌腱经梯度脱水后在无水甘油中常温保存效果更佳。     

甘油常温保存肌腱的两种方法比较

背景:最大程度保留肌腱的生物活性是进行同种异体肌腱移植的条件。目的:通过比较选择出最佳的肌腱常温保存方法。方法:使用无菌操作的方法将兔肌腱随机分为4组:新鲜肌腱对照组、生理盐水组、无水甘油Ⅰ组(肌腱梯度脱水后在无水甘油中保存)、无水甘油Ⅱ组(肌腱直接在无水甘油中保存),分别于保存的第2,4,7个月进行检测。结果与结论:苏木精-伊红染色显示:无水甘油Ⅰ组肌腱细胞完整率明显高于无水甘油Ⅱ组。电镜观察发现,无水甘油Ⅰ组大部分肌腱细胞形态正常,肌腱组织结构完整;无水甘油Ⅱ组保存4,7个月后细胞核凝固、固缩、崩解状。无水甘油Ⅰ组的超氧化物歧化酶活性也明显高于无水甘油Ⅱ组。说明肌腱经梯度脱水后在无水甘油中常温保存效果更佳。     

Observations on Autologous, Previously Frozen, Deglycerolized, Agglomerated, Resuspended Red Cells

Forty-four units of human erythrocytes preserved with glycerol, the slow-freeze technic, and agglomeration were evaluated by autotransfusions of chromium-labeled 10 cc aliquots. The shipment of frozen glycerolized red cells in dry ice and of thawed, deglycerolized, resuspended red cells in wet ice did not adversely affect the in vivo or in vitro measurements. Excessive in vitro loss of cellular hemoglobin and unacceptable posttransfusion survival were observed when preserved erythrocytes were stored at + 4 C for longer than 24 hours, at —20 C for longer than three days, and at — 30 C for longer than seven days between periods of storage at — 80 C. A storage period at —20 C alone for 50 days resulted in poor preservation.
Adenine supplementation of the glycerolized red cells (0.6 mM per liter) prior to freezing did not change significantly the in vivo or in vitro characteristics of red cells stored at the warmer temperatures.
Highly significant correlations were noted between the 24-hour posttransfusion survival and MCV, MCHC, osmotic fragility, and density distribution of the preserved red cells.     

Stability of Lipids of Erythrocytes Preserved in Liquid Nitrogen

Human erythrocytes preserved by a low-glycerol, rapid-freeze method maintained normal lipid concentrations after storage at -196 C for three to 146 weeks. In contrast to erythrocytes stored in ACD at 4 C, the frozen cells exhibited no tendency to develop progressive losses of total lipid weight, cholesterol, or lipid phosphorus. Relative distribution of individual phospholipids was similar to that of fresh control specimens. No streaking or other evidence of products of oxidative degradation was observed in thin-layer chromatograms, and assays for peroxide formation in lipid extracts were unaffected by prolonged storage of the frozen cells. These findings indicate that the structural integrity of erythrocyte membrane lipoproteins remained intact for up to three years after preservation by a low-glycerol, rapid-freeze technic and storage in liquid nitrogen at -196 C.     

Whole blood storage solution for erythrocyte sodium and sodium-lithium countertransport rate determination

A solution consisting of heparinized and buffered isosmolar magnesium chloride is described in which whole blood may be stored for preservation of erythrocyte sodium (Na) and membrane sodium-lithium countertransport (Na-Li CT). Correlation (r) of fresh versus 24-h stored erythrocytes for Na was 0.990 (y = 0.30 + 0.956x) and for Na-Li CT was 0.995 (y = -0.014 + 1.022x). Na-Li CT rate was preserved in the storage solution for up to five days and erythrocyte Na concentration for at least 24 h. The solution should find application in epidemiological studies of erythrocyte Na and Na-Li CT rate in human essential hypertension as the blood specimens require no centrifugation or erythrocyte washing procedures prior to laboratory analysis.     

The effect of storage time of human red cells on intestinal microcirculatory oxygenation in a rat isovolemic exchange model

OBJECTIVE: To determine whether the storage time of human leukodepleted red blood cell concentrates compromises intestinal microvascular oxygen concentration oxygen (muPo(2)) during isovolemic exchange transfusion at low hematocrit. DESIGN: Prospective, randomized, controlled study. SETTING: University research institute laboratory. SUBJECTS: Male Wistar rats. INTERVENTIONS: Intestinal muPo(2) was determined by Pd-porphyrin phosphorescence life-time measurements. MEASUREMENTS AND MAIN RESULTS: Rats were brought near to a state of oxygen supply dependency by hemodilution with a pasteurized plasma protein solution to a hematocrit of 14.3 +/- 1.1% (n = 24). Subsequently, an isovolemic exchange transfusion with human leukodepleted red blood cells, stored for 2-6 days (fresh, n = 8), 2-3 wks (intermediate, n = 8), or 5-6 wks (old, n = 8), was performed to determine whether intestinal muPo(2) would be preserved. Immunologic reactions were avoided by washing the red blood cell concentrates three times before use. Isovolemic exchange with fresh and intermediate red blood cells maintained muPo(2) whereas old cells decreased muPo(2) with 26%. Subsequent transfusion with red blood cells (hematocrit approximately 60%) until reaching a hematocrit of 32.4 +/- 2.1 % (n = 24) increased intestinal muPo(2) in all three groups to the same extent between 28% and 32%. No changes in red blood cell deformability, as determined by a Laser-assisted Optical Rotational Cell Analyzer, could be demonstrated during 5 wks of storage. CONCLUSION: This study shows that at low hematocrit, the oxygen-delivering capacity of human red blood cells stored 5-6 wks is reduced compared with fresh cells and red blood cells stored for an intermediate period. Although red blood cells stored for 2-3 wks are completely devoid of 2,3-diphosphoglycerate, their oxygen-delivering capacity to the intestines was the same as fresh red blood cells. Our study showed that red blood cell deformability was preserved during storage, suggesting that other mechanisms may account for the observed decrease in oxygen delivery by red blood cells stored 2-3 wks.     

不同年龄大鼠大脑中动脉弹性纤维的变化

背景大脑中动脉弹性纤维的变化与老年性脑血管疾病密切相关.目的观察不同年龄大鼠大脑中动脉弹性纤维的变化.设计以实验动物为观察对象的对比实验.单位解放军第三军医大学解剖教研室和中心实验室.材料健康Wistar大鼠36只,雌雄不拘,体质量200～280 g,由重庆第三军医大学动物所提供[生产合格证号SCXX(军)2002-007].干预运用光镜、电镜和图像分析仪对不同年龄的大鼠大脑中动脉弹性纤维进行系统研究.主要观察指标①主要结局各年龄大鼠大脑中动脉内弹性膜的变化.②次要结局透射电镜下观察内弹性膜超微结构变化.结果随着年龄增加,内弹性膜折叠的幅度和数量均减小,弹性纤维含量显著减少(P＜0.01),胶原纤维与弹性纤维的比值显著增加(P＜0.01);＞24月龄组内弹性膜变薄,不均质,内皮细胞和平滑肌细胞向内弹性膜穿过,内弹性膜内出现脂质,并有分层、断裂现象.结论弹性纤维的变化可能与年老后易发生脑血管疾病有关.     

Mitochondrial ischemia-reperfusion injury of the transplanted rat heart: improved protection by preservation versus cardioplegic solutions

Cold ischemia time and preservation of organs are limited by I/R injury leading to primary nonfunction of the graft. In a rat heart transplant model, we compared cardioplegic St Thomas (ST) to histidine-tryptophan-ketoglutarate (HTK) and University of Wisconsin preservation solutions in terms of contractile function, and mitochondrial respiratory and enzymatic defects after prolonged cold ischemia and reperfusion. Contractile function was scored after transplantation and 24 h of reperfusion. Mitochondrial function was investigated by high-resolution respirometry of permeabilized myocardial fibers. Graft performance in terms of contractile function declined with the duration of cold storage. Recovery was significantly improved after 10 h of cold storage in HTK compared with ST (cardiac scores, 3.3+/-0.5 and 1.8+/-0.8, respectively). Tissue lactate dehydrogenase was better preserved in HTK than ST. Increase of tissue water content (edema) was less pronounced in HTK than ST (3.33+/-0.14 and 3.73+/-0.21 mg/mg dry weight, respectively). Similar cardiac scores (2.6+/-0.9 and 2.9+/-1.2, respectively) and mitochondrial respiratory parameters were obtained after preservation in HTK and University of Wisconsin. Decline in contractile function of individual grafts correlated well with loss of mitochondrial respiratory capacity, whereas citrate synthase activity remained largely preserved, indicating specific damage of respiratory complexes. Our data provide evidence for the superiority of preservation solutions versus a cardioplegic solution for prolonged cold storage of the heart. The correlation of graft performance and mitochondrial function indicates the potential of high-resolution respirometry for quantitative assessment of myocardial injury upon cold I/R, providing a basis for diagnostic approaches and evaluation of improved preservation solutions for heart transplantation.