Molecular Weight of Cytoplasmic Malate Dehydrogenase,Mitochondrial Malate Dehydrogenase and Lactate Dehydrogenase of a Freshwater Catfish

The multiple molecular forms of cytoplasmic malate dehydrogenase(cMDH),mitochondrial malate dehydrogenase(mMDH)and lactate dehydrogenase(LDH)were studid in the liver and skeletal muscle of the freshwater catfish,Clarias batrachus.There were two electrophoretically distinguishable bands(AAand BB)of cMDH and mMDH which suggests that they are apparently encoded at two gene loci(A and B)in both the tissues.However,the presence of a single band(LDH-1)of LDH in liver and double bands(LDH-1and LDH-2)in skeletal muscle in which LDH-2 was predominant reflects the differentialexpression of LDH genes in different metabolic tissues to meet the requirement of energy production.The AA isoform(74kd)of liver cMDH was smaller than those o the AA form(110kd)of skeletal muscle.In contrast,the BB isoform of liver(42kd)and skeletal muscle(54kd)were more or less similar in size.Unlike the case of cMDH,the molecular weight of AA isoform(115kd)of liver mMDH was higher than those of the AA form(87kd)of skeletal muscle.Whereas the molecular weight of BB isoform(58kd)of liver was in proximity to the weight of BB form(44kd)of skeletal muscle mMDH.The size of AA muscle was larger as compared to AA form of mMDH in the liver(115kd) and skeletal muscle(87kd).But the size of BB isoform of both the isozyme was almost equal in these metabolic tissues.The molecual weight of liver LDH-1(96kd)was close to the weight of LDH-1(82kd)in skeletal muscle.The molecular weight of skeletal muscle LDH-2 was deduced as 37kd which is much more lower than the weight of LDH-1 in liver and skeletal muscle.The smaller size of LDH-2 in skeletal muscle may be of a physiological significance in this anaerobic tissue.     

A Preliminary Studay of Lactate Dehydrogenase and Its Isoenzymes in Schizophrenics

The Activity of iactate dehydrogenase(LDH)andits isoenzymes may be increased in some physicaldiseases as well as in some mental diseases.ooBck(1974) reported that the mean activity unit of serumLDH and its isoenzymes tended to increase in 39schizophrenics. These results were similar to those     

Isolation and Characterization of Rat Liver Polyribosomes.

As the first step of selecting a specific mRNA,we would have polyribosomcs available.Weused ultracentrifugation method to isolate poly-ribosomes from rat liver.The UV spectrum andA260/A280 ratio of this preparation showed thecharacteristics of polyribosomes,so did the sedi-     

Studies on Active Immunization of Rhesus Monkeys with Mouse Lactate Dehydrogenase C4

Ten fertile female rhesus monkeys were immunized with mouse LDH-C4.Antibody titer and specificity were examined by the methods of double immunodiffusion and Bio-Dot enzyme immunoassay(BDEIA).Our results indicated that the antibody reactivity of immunized monkeys can be detected by BDEIA on day 7 after primary immunization.The highest response of the monkeys to LDH-C4 immunization was observed on day 35 of immunization.Individual variation of the antibody titer from 1:800 to 1:12800 by BDEIA and 1:1 to 1:32 by double immunodiffusion assay were evident.Specific precipitation lines can only be seen between the antisera and LDH-C4 antigen,but not between the antibodies and LDH isozymes from other somatic tissues.Meanwhile,the monkey antisera not only reacted with mouse and monkey LDH-C4,but also with LDH-C4 from human semen and testis.No antibody reaction was detectable in the control animals.Comparison of the pregnancy rate indicated that there was no significant difference between the immunized and control groups,However,a remarkable delay of the course of pregnancy and a decrease in the body weight of the neonatal animals were observed in the immunized monkeys.Our studies provided valuable new information of the active immunization of rhesus monkeys against mouse LDH-C4.     

Changes of Lactate Dehydrogenase Activity in Airways of Guinea-pigs after Diphosgene Poisoning

Lactate dehydrogenase(LDH),as an indicator of cell damageor death,was used to determine the diphosgene damage of respiratorytract cells.Guinea-pigs were exposed to diphosgene at concentrationswithin the LCt_(50) range and above.The content of LDH in thebronchopulmonary washings of guinea-pigs was significantly increa-sed after poisoning.There was no increase of free hemoglobincontent in the washings.It was indicated that lysed erythrocyteswere not the source of LDH in the poisoned group.The LDH iso-enzyme pattern in the poisoned group washings suggests that theincreased LDH comes from the respiratory tract cells.These resultsshow diphosgene can directly damage cells of the respiratory tract.     

Isolation and Purification of Terminal Deoxynucleotidyl Transferase by Using Chromatofocusing and Preparation and Identification of Its Autiserum

Using a combined method of chromatofocus-ing and oligo(dT)-cellulose affinity chromato-graphy, the terminal deoxynucleotidyl transferase(TdT) isolated from the calf thymus was puri-     

The Isolation,Purification and Identification of Fumitremorgin B Produced by Aspergillus fumigatus

Twenty-six strains of Aspergillus fumigatus were screened for toxigenicity for fumitremorgins A and B.Twenty-three of 26 strains can produce fumitremorgin B in rice medium determined by TLC and HPLC,and no fumitremorgin A was detected.The strains of no.C4104 and no.3656 were inoculated onto 5 kg of rice media and incubated in a modified procedure.Finall,4.0g of fumitremorgin B was obtained after extraction and purification by modified methods,and was confirmed by TLC,HPLC,spectral analysis together with other physicochemical analysis.This is the first report of the preparation of fumitremorgin B in China.     

Expression of 11β-Hydroxysteroid Dehydrogenase mRNA in Rat Leydig Cells

11β-hydroxysteroid dehydrogenase (11β-HSD) in Leydig cells regulates sterodogenesis by controlling intracellular glucocorticoid (corticosterone, B,in rat) concentration.Prior to the 26th postnatal day, 11β-HSD is absent from rat immature Leydig cells. As the Leydig cells are matured,     

Analysis of DNA Content of Various Types of Spermatogenic Cells in Rat after Testicular Heating with Flow Cytometry

To measure DNA contents of spermatogenic cells and analyze the efficiency of spermatogenesis after testicular heating in rat Methods Eighty adult male Sprague-Dawley rats were randomly divided into experimental group （43 ℃, 30 min） and control group （22 ℃, 30 min）. According to day 0.5, 1, 3, 6, 10, 25, 35 and 50 after local testicular heating, every group was divided into 8 subgroups： experimental subgroups （n=6） and control subgroups （n=4）. DNA contents of various types of germ cells were observed with flow cytometry （FCM） in all groups. Results Compared with control groups, percentages of 4C cell （primary spermatocyte） in 0.5-35 d groups and percentages of 1C cell （spermatid and sperm） in 6-50 d groups significantly decreased in experimental groups （P〈0.05）, and percentages of 2C cell （spermatogonium and second spermatocyte） in 3 -35 d experimental groups increased significantly after heating （P〈0.05）. 4C：2C in all of 8 experimental groups and 1C：2C in 3-35 d experimental groups were down （P〈0. 05）, and in 1 d experimental group 1C：4C was up after heating （P〈0.05）. Conclusions After being heated, the number of spermatocyte firstly decreased, and then that of spermatid and sperm decreased too. Heat influences several stages in spermatogenesis and results in suppression of spermatogenesis. Flow cytometry is an effective method for researching on the change of spermatogenesis and has significance on mechanism about changing of spermatogenic cells induced by heat.     

Further Analysis of El Tor Vibrio Cholerae Strains 1d： Comparison of Two Typing Methods

ＦｕｒｔｈｅｒＡｎａｌｙｓｉｓｏｆＥｌＴｏｒＶｉｂｒｉｏＣｈｏｌｅｒａｅＳｔｒａｉｎｓ１ｄ：ＣｏｍｐａｒｉｓｏｎｏｆＴｗｏＴｙｐｉｎｇＭｅｔｈｏｄｓＪｉａｎｇＸｉａｏｗａｎ（蒋小皖）；ＣｈｅｎＳｈｕｃｈａｎ（陈树昶）；ＪｉａｎｇＸｉａｏｌｉｎ（蒋小陵...     

大鼠睾丸乳酸脱氢酶同工酶－C_4的分离纯化研究

采用加热、硫酸铵分段盐析和ＤＥＡＥ－ＳｅｐｈａｄｅｘＡ－５０柱层析，从大鼠睾丸中分离纯化了ＬＤＨ－Ｃ４。比活性提高７８４倍，活性回收率为１２．３％，纯化的ＬＤＨ－Ｃ４经聚丙烯酰胺凝胶电泳（ＰＡＧＥ）鉴定均为一条酶带，位于ＬＤＨ３和ＬＤＨ４之间。     

乳酸脱氢酶-C_4在大鼠不同成熟期生精细胞中的定位

用ABC免疫组织化学法观察乳酸脱氢酶C_4(LDH-C_4)在性成熟后的Wistar大鼠生精上皮细胞中的定位。结果表明:LDH-C_4定位于粗线期初级精母细胞以后的各级生精细胞中,精原细胞中缺如。显微分光光度分析表明,随着精子的发生,LDH-C_4的相对含量逐渐增加,精母细胞中为0.55±0.09,精子细胞中为0.86±0.13,精子中为0.94±0.04,三者之间有显著性差异(P<0.01)。ABC免疫电镜显示LDH-C_4在大鼠精子的细胞质、线粒体及部分细胞膜上均有分布。免疫荧光显示LDH-G_4在精子表面主要定位于尾部的主段和末段,少量分布于精子头冠部。作者对LDH-C_4细胞定位的生理意义作了讨论。     

大鼠精子特异的LDH-C_4的提纯及其免疫性质分析

采用硫酸铵盐析、N~6-(6氨己基)-5′-AMP-Sepharose亲和层析和DEAE-Cellulose离子交换层析,从大鼠睾丸中纯化了LDH-C_4。比活性为16.3 IU/mg,纯化1087倍,产率32.5％。纯化的LDH-C_4经聚丙烯酰胺凝胶电泳(PAGE)鉴定为一条酶带。将纯化的LDH-C_4免疫兔制备了效价为1:64的抗血清。BA-ELISA检测抗血清与大鼠心脏、肌肉和人、小鼠睾丸提取液之间的交叉反应,结果表明抗血清与大鼠心脏、肌肉提取液间存在微弱的交叉反应,而双向免疫扩散未形成沉淀线;抗血清与小鼠睾丸提取液之间存在很强的交叉反应,但与人睾丸提取液之间的交叉反应却很弱,双向免疫扩散显示它们的抗原决定簇部分相同,提示不同种属的LDH-C_4之间抗原性存在差异。     

大鼠胰岛细胞的分离纯化及培养

目的　探讨大鼠胰岛细胞分离纯化和培养的方法。方法　采用胰导管逆行灌注Hanks液分离成年大鼠胰岛,Ⅴ型胶原酶消化,不连续密度梯度Ficoll 4 0 0纯化胰岛。用台盼蓝和双硫腙染色检测胰岛活性和纯度,葡萄糖刺激胰岛素释放试验检测胰岛功能。结果　胰岛细胞的收获量为5 4 0±15 0个胰岛 胰腺,活性>90 % ,纯度>90 % ,高糖刺激后胰岛素释放量为低糖刺激的2倍多。结论　胰导管灌注Hanks液,Ⅴ型胶原酶消化和不连续密度梯度葡聚糖纯化胰岛,可以获取数量多,活性和纯度好的胰岛     

两种不同纯化大鼠胰岛细胞方法的比较

  目的 比较分析OptiPrep和Ficoll两种介质纯化大鼠胰岛细胞的效果。方法 将24只SD大鼠随机分成2组,采用明尼苏达大学改良方法提取大鼠胰岛细胞,消化分离胰岛细胞后,分别采用不连续密度梯度Ficoll离心法和OptiPrep离心法纯化胰岛,比较纯化后胰岛细胞数量、纯度、存活率及功能,观察OptiPrep和Ficoll两种介质对大鼠胰岛纯化效果的影响。结果 与 Ficoll组相比,OptiPrep组胰岛细胞的产量（分别为304±13.57和346±9.09）和纯度（分别为88.25%±2.22%和94%±0.82%）均显著增加（P0.05）。 结论 OptiPrep法纯化大鼠胰岛比传统的Ficoll法更有优势,而且OptiPrep价格相对便宜,可以作为纯化大鼠胰岛的常规方法。     

大鼠IgE依赖组胺释放因子重组蛋白纯化效果的优化

目的提高大鼠IgE依赖组胺释放因子(rHRF)的可溶性表达水平,改善其亲和层析纯化效果,为深入研究该重组蛋白的生物学功能奠定基础。方法选择多克隆酶切位点BamHⅠ和XhoⅠ,将rHRF完整编码区基因从原有的pET-30a-rHRF重组质粒亚克隆至pET-28a载体,构建重组质粒pET-28a-rHRF;转化大肠杆菌BL21(DE3)后,通过改变IPTG浓度(0.1～1.0mmol/L),调节诱导表达温度(25～37℃)和时间(1～5h),筛选能够提高rHRF可溶性表达水平的条件;同时改进亲和层析纯化的条件,提高纯化效果。结果成功构建重组质粒pET-28a-rHRF,不同IPTG浓度、诱导温度和时间对重组蛋白的可溶性表达水平无明显影响。但是,与pET-30a-rHRF转化菌相比,pET-28a-rHRF转化菌所表达的重组蛋白因其N端和C端均带有6×his标签,增强了目的蛋白与树脂的结合力,而使亲和层析纯化效果(重组蛋白的浓度和纯度)显著提高。结论rHRF重组蛋白的可溶性表达水平不受IPTG浓度、诱导表达温度和时间的影响,但是可溶性重组蛋白两端均携带6×his标签可使亲和层析纯化效果显著提高,从而有利于获得足量rHRF用于进一步的生物学功能研究。     

新生大鼠嗅球嗅鞘细胞的纯化方法研究

目的 探讨新生大鼠嗅球嗅鞘细胞（OECs）的体外培养和纯化方法，为研究脊髓损伤后神经再生获得丰富OECs来源奠定基础。方法 用原代培养的方法分离、培养新生大鼠嗅球OECs，比较p75抗体吸附、阿糖胞苷抑制、差速贴壁三种方法的纯化效果，根据GFAP免疫细胞化学染色结果统计所得OECs的纯度，同时在相差显微镜下对不同培养时期的OECs的形态进行观察。结果 体外培养的新生大鼠嗅球OECs主要为双极或三级细胞，其突起细长。未经纯化的OECs则成纤维细胞生长迅速而占优势，三种纯化方法均能使接种14d后的OECs纯度均值大于75％。p75抗体吸附法和阿糖胞苷抑制法的纯化率均值略高于差速贴壁法。结论 新生大鼠嗅球OECS的原代培养中细胞纯化是必要的；在脊髓损伤再生研究中，较之p75抗体吸附法和阿糖胞苷抑制法，差速贴壁法是一种简单、经济、实用的OECs纯化方法。     

缺血再灌注对大鼠睾丸脑红蛋白表达的影响

目的探讨大鼠睾丸缺血再灌注对脑红蛋白（neuroglobin,NGB）表达的影响及意义。方法48只健康雄性SD大鼠随机分为正常对照组、假手术组和实验组,建立大鼠睾丸缺血再灌注模型,分别于缺血再灌注后3、6、12、24、48、96h脱颈处死大鼠,提取睾丸组织,常规HE染色进行形态学观察,应用实时荧光定量PCR和Westernblot技术检测NGB的表达变化。结果大鼠睾丸缺血再灌注性损伤后睾丸生精细胞水肿,96h后可见细胞层数减少,排列紊乱,管腔内精子细胞减少。NGB在睾丸缺血再灌注后表达升高,其中mRNA水平6h组和48h组与正常对照组相比差异显著（P〈0．05）,蛋白水平6h组、12h组和96h组与正常对照组相比差异显著（P〈0．05）。结论NGB在大鼠睾丸缺血再灌注后表达升高可能对睾丸损伤有一定的保护作用。     

大鼠睾丸和附睾的去甲肾上腺素能神经支配

应用改良的乙醛酸诱发儿茶酚胺(CA)的荧光组织化学方法,证明去甲肾上腺素(NA)能神经终末在大鼠睾丸和附睾内的分布及其构筑形式。实验发现,大鼠皋丸间质结缔组织及血管壁接受NA能神经支配:此种带膨体荧光神经纤维发出分枝,伸进曲细精管壁;神经终末包绕着睾丸网内小管壁;NA能神经终末延伸进入附睾头部的输出小管,并在管腔内形成网织状结构;在附睾体,有神经纤维伸入附睾管内;附睾尾部,荧光膨体纤维分布更为丰富。