Electrophoretic investigation of nuclear membrane ribonucleases of rat liver and hepatoma-27 cells

Preparations of the nuclear membranes were obtained from purified nuclei of rat liver and hepatoma-27 cells, and from them enzyme-containing extracts of acid-soluble proteins were then prepared. The protein extracts were subjected to disk electrophoresis in 15% polyacrylamide gel. Ribonucleases (RNases) which are constituents of the acid-soluble proteins of the nuclear membranes of normal liver were found to be present as several different components which differed in their electrophoretic mobility and in several physicochemical properties from crystalline bovine RNase and the RNase of nuclear chromatin.Institute of Problems in Oncology, Academy of Sciences of the Ukrainian SSR, Kiev. N. K. Kol'tsov Institute of Developmental Biology, Academy of Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR R. E. Kavetskii.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 84, No. 12, pp. 670–672, December, 1977.     

应用激光电泳技术对MA737小鼠乳腺癌细胞膜表面电荷特性的研究

本工作应用激光多普勒电泳技术对ＭＡ７３７小鼠乳腺癌细胞膜表面电荷特性及高温（４２．５℃）对癌细胞的热效应进行了深入的研究。结果发现，ＭＡ７３７癌细胞的电泳迁移率（ＥＰＭ）明显高于相应正常细胞的ＥＰＭ（约２０％），表明癌细胞表面负电荷增高；实验发现唾液酸可能是ＭＡ７３７癌细胞膜表主要负电基团的来源之一。研究还发现，ＭＡ７３７癌细胞的热敏性比相应的正常细胞强，随高温受热时间的延长，癌细胞的ＥＰＭ呈下降趋势。     

Human myeloma light chains with increased molecular weight: high frequency among lambda chains

The discovery of a human myeloma protein comprising a kappa L-chain with an increased mol. wt of 30,000) (Bouvet et. al., 1980) prompted investigations on the incidence of such heavier L-chains among other human myeloma proteins. In 105 samples examined, 34 were found to have L-chains heavier than normal (23,000-24,000), ranging from 25,000 up to 31,000, and five of lighter mol. wt (21,000-22,000). These mol. wt abnormalities were detected by electrophoresis in sodium dodecyl sulfate 10% polyacrylamide gels (SDS-PAGE) after reduction with 2-mercaptoethanol. The mol. wt of three of the heavier kappa or lambda chains was also estimated by filtration through a Sephadex G100 column and by sedimentation equilibrium. All three methods indicated a mol. wt increase of about 15-25% as compared with the usual mol. wt. The distribution of the high mol. wt chains among all L-chains examined was found to be 11 out of 62 kappa chains (17.7%) and 23 out of 43 lambda chains (53%) (P less than 0.001). A preferential association of such L-chains with H-chains producing multiple bands in SDS-PAGE (P less than 0.01) and an association between multiple L-chain and multiple H-chain band (P less than 0.05) were also observed. In contrast, no abnormal L-chain was found in immunoglobulins from normal subjects. Spontaneous degradation of the normal H-chains sometimes yielded fragments of 30,000 mol. wt. These fragments were easily distinguishable from abnormal L-chains. The nature of extra mol. wt in heavy L-chains was investigated for the presence of carbohydrate moiety. Four large and three normal size L-chains were examined for amino-sugar and sialic acid content. A small amount (one residue per molecule) of amino-sugar was detected only in two normal and two heavy L-chains, whereas sialic acid was only found in the heaviest (27,000-30,000) L-chains (Lh) and in small percentage (one or two residues per molecule). Total sugar estimation in one Lh chain indicated a proportion not exceeding three or four residues per L-chain (mol. wt 1,000) and this is insufficient to explain the 15-25% (3,600-6,000) mol. wt increase. It is therefore possible that, at least in some heavy myeloma L-chains, an additional peptide is expressed. Whatever the nature of the increase it would be of interest to elucidate whether this is a marker of malignant process or of an intermediate step of normal Ig synthesis.     

β Chain deficiency in three patients with dysfunctional C8 molecules

Structural and functional studies were performed on a dysfunctional C8 molecule present in the serum of two siblings and an unrelated individual. The C8 in these three sera exhibited a pattern of partial immunologic identity with C8 in normal serum but was devoid of functional activity. The C8 was immunoprecipitated from the three sera and from a control serum with an antihuman C8 antiserum and analyzed by SDS-PAGE using highly purified human C8 as a reference. A selective absence of a band of 62,000 mol. wt was observed in the immunoprecipitates from the sera containing dysfunctional C8. Experiments performed with the purified α-γ and γ subunits showed that the hemolytic activity of the C8 deficient sera could be reconstituted by the addition of the β chain but not the α-γ dimer. Binding of the dysfunctional C8 to C$\text{567}$ was excluded by the following observations: (1) EAC 1–7 treated with the C8 deficient sera and then washed could not be lysed after the addition of the β subunit and C9; and (2) the abnormal molecules did not interfere with the consumption of normal C8 by the soluble complex SC5b-7.     

三氧化二砷对肝癌细胞系HepG2生长及核基质蛋白的影响

目的 探讨三氧化二砷对人肝癌细胞系ＨｅｐＧ２的生长及核基质蛋白的影响。方法 应用生长指数分析和高分辨ＳＤＳ－聚丙烯酰胺交双向电泳比较用Ａｓ２Ｏ３处理后的ＨｅｐＧ２细胞主核基质蛋白的改变。结果 Ａｓ２Ｏ３抑制ＨｅｐＧ２的生长,双向电泳显示经Ａｓ２Ｏ３处理后ＨｅｐＧ２细胞内出现新的核基质蛋白,并且该蛋白是细胞特异性的。结论 核基质蛋白是某些抗癌药物的作用靶子,Ａｓ２Ｏ３抑帛 癌细胞的生长可望有用于临床     

定向克隆构建RAB5A基因真核细胞表达载体

目的 探讨ＲＡＢ５Ａ作为蛋白质人胞信号传导调控者在肿瘤转移中的作用及影响。方法 采用双酶切、定向克隆技术建立了人ＲＡＢ５Ａ基因真核细胞表达载体ｐｃＤＮＡ３．１（－）０ＲＡＢ５Ａ。结果 测定分析证实了ＲＡＢ５Ａ基因正向插入ｐｃＤＮＡ３．１（－表达载体,免疫组化分析表明转染了ｐｃＤＮＡ３．１（－）－ＲＡＢ５Ａ表达载体的ＡＧＺＹ８３－ａ细胞系细胞内的荧光亮度明显强于转染前,蛋白电泳显示近２３ＫＤ的蛋白质     

高效毛细管电泳分离测定头孢克罗血浆浓度

采用乙腈直接沉淀血浆蛋白,以头孢拉定为内标,建立了适合人血浆中头孢克罗分离测定的高效毛细管电泳方法。在48cm×50μm未涂层石英毛细管柱上,选择50mmol／L磷酸二氢钠-12．5mmol／L硼砂(pH8．08)为运行缓冲液,操作电压20kV(＋)→(－),压力进样12psi×s,柱上紫外265nm检测,头孢克罗和内标头孢拉定在6．5min内获得了理想的分离,内源性物质不干扰高效毛细管电泳定量分析。血药浓度在0．25～25．0μg／ml范围内线性关系良好(r＝0．9993,n＝5),绝对回收率大于90．5％,日内及日间精密度均小于9．5％,并成功地应用于健康志愿者单剂量口服头孢克罗普通胶囊后血药浓度的动态分析。     

新生儿脐血血液流变学观察

目的 :观察新生儿脐血的血液粘度、血浆粘度、红细胞压积、血沉、红细胞电泳情况。方法 :新生儿分娩断脐后立即抽取脐血 4ml+肝素抗凝 ,并立即送检 ,采用北京普利生 L BY- N6 A自清洗旋转式粘度计检验。结果 :母体为正常生理妊娠的新生儿组 :全血低切粘度 8.91± 6 .79m pa· s,全血高切粘度 5 .6 6± 2 .0 mpa· s,血浆粘度 1 .46± 0 .2 4m pa· s,红细胞压积 5 0 .1±1 8.4% ,血沉 2 .1 4± 1 .36 mm/ h,红细胞电泳 1 5 .82± 3.79s;母体为妊高征的新生儿组 :全血低切粘度 1 1 .96± 7.38m pa· s,全血高切粘度 6 .48± 1 .5 7mpa· s,血浆粘度 2 .5 3± 1 6 .4mpa· s,红细胞压积 31 .47± 0 .35 % ,血沉 1 .6 8± 1 .43mm/ h,红细胞电泳1 5 .6 8± 3.43s;正常妇女组 :全血低切粘度 2 2 .33± 6 .88mpa· s,全血高切粘度 4.5 0± 0 .5 4mpa· s,血浆粘度 1 .73± 0 .0 9mpa· s,红细胞压积 41 .0 9± 2 .6 0 % ,血沉 1 7.0 9± 6 .35 mm/ h,红细胞电泳 2 1 .0 4± 1 .44 s。结论 :孕期母体血液流变学变化对新生儿影响较小     

Nerve-induced release of nitric oxide in the rabbit gastrointestinal tract as measured by in vivo microdialysis

1. Nitric oxide (NO) has been suggested as a gastrointestinal neurotransmitter, mediating the gastric receptive relaxation and the relaxation in the peristaltic reflex. The aim of the present study was to measure nerve-induced NO formation in vivo in the gastrointestinal tract.
2. Formation of the nitric oxide oxidation products nitrite and nitrate during vagal nerve stimulation were measured in the anaesthetized rabbit. Microdialysis probes were inserted into the wall of the stomach and proximal colon, and nitrite and nitrate in dialysate measured by capillary electrophoresis.
3. During bilateral vagal nerve stimulation there was an increase in nitrite and nitrate formation at the level of the stomach and in nitrite formation at the level of the colon. This increase was inhibited by intravenous administration of the NO synthase inhibitor N^ω-nitro-L-arginine methyl ester (L-NAME 30 mg kg⁻¹). Furthermore, L-NAME significantly increased nerve-induced gastric and colonic contractions, as well as spontaneous colonic contractions.
4. In summary, we present a new methodological procedure for quantification of small changes in nitric oxide formation in vivo. This study provides evidence that nitric oxide is released in the stomach and colonic wall during vagal nerve activity, at concentrations able to cause inhibition of smooth muscle contractions in vivo.

     

琼脂糖火箭电泳法测定唾液溶菌酶活性

本文采用琼脂糖火箭电泳法测定唾液溶菌酶活性,将琼脂扩散方法和电泳技术结合一起,使溶菌酶在电场中泳动,在通过含有作用底物的凝胶时与溶壁微球菌发生水解作用,形成可见的溶菌透明峰,根据峰值与溶菌酶浓度的时数值成正比的关系求得样品溶菌酶含量。该方法简便、快速、稳定性好,灵敏度高,结果准确可靠。在本实验条件下最低检出量为1.25 μg/ml,在5.O-100.0μg/ml范围内呈良好的浓度-峰高的线性关系,r=O.9947,P<0.001,批内重复变异系数CV=4.15％,批间重复变异系数CV=7.42％,平均回收率为95％～114％。