转化生长因子β的提取、纯化及其对肝细胞凋亡的诱导作用

用酸/醇法从新鲜的牛血小板中粗提TGF-β。经离子交换色谱和凝胶色谱纯化后,收集经SDS-PAGE鉴定相对分子质量为25×10     

凝胶过滤与镍柱亲和纯化金葡菌a-溶血素重组蛋白的生物学特性比较

以在宿主菌株BL21(DE3)中成功表达的重组金黄色葡萄球菌a-溶血素蛋白为研究对象, 分析比较通过凝胶过滤层析(Gel filtration chromatography)和镍柱亲和层析纯化试剂盒(Ni-NTA spin columns)纯化所得到的重组蛋白的蛋白含量和生物特性方面的差异。SDS-PAGE分析检测纯化产物, Bradford法测定蛋白含量, 兔红细胞测定半数溶血效价。结果显示这2种方法得到的纯化产物在53 kD处均呈现单一清晰带, 达电泳级纯度。与此同时, 凝胶过滤对目的蛋白的纯化率为14.04%, 蛋白含量为0.337 mg/mL, 溶血活性为1519 HU/mg; 镍柱亲和层析的纯化率为17.5%, 蛋白含量为0.35 mg/mL, 溶血活性为1463 HU/mg。由此可见, 凝胶过滤得到的纯化产物在蛋白含量和蛋白活性方面丝毫不亚于镍柱亲和层析纯化试剂盒。     

Histocompatibility lymphocytic antigen (HLA) typing in patients with acute exacerbation of Hashimoto's thyroiditis

Histocompatibility lymphocytic antigen (HLA) typing was performed in 6 patients with acute exacerbation of Hashimoto's thyroiditis whose diagnoses were established on the basis of typical histological findings, and was compared with those of 12 with subacute thyroiditis, 33 with general Hashimoto's thyroiditis and also with a control group. There was a high incidence of BW35 in patients with subacute thyroiditis, although it was only seen in 1 of 6 patients with acute exacerbation. The difference was statistically significant (p less than 0.01). Four of 6 patients with acute exacerbation had DR2 and none of them had DR4, which was the reverse of the findings for Hashimoto's thyroiditis patients in general, and the difference in the incidence of DR2 was significant (p less than 0.001). None of the HLA types in patients with acute exacerbation was significantly different from those of the control group. In conclusion, HLA typing in patients with acute exacerbation was different from those of subacute thyroiditis and general Hashimoto's thyroiditis. Acute exacerbation was considered to involve quite a limited and rather unique population among patients with Hashimoto's thyroiditis.     

重组人核苷二磷酸激酶A的理化性质

对重组人核苷二磷酸激酶A（rhNDPK-A）进行纯化,并对重组产物的理化性质及在溶液中的聚合状态进行鉴定。NDPK-A工程菌发酵后的菌体高压匀浆,然后微孔过滤、超滤浓缩,所得样品经DEAE阴离子交换、Cibacron Blue亲和层析、分子筛层析三步纯化后,以SDS-PAGE和RP-HPLC分析纯化产物的纯度,RP-HPLC测定酶活性。合格制品以基质辅助激光解析飞行时间质谱测定相对分子质量（MW）;Edman降解法测定N末端序列;多角度激光散射法测定重组产物在溶液中的表观分子量。结果表明,rhNDPK-A纯化产物的SDS-PAGE纯度为97.3%,RP-HPLC纯度为99.2%;比活性为（900±100）u/mg;单体相对分子质量为17017,与NDPKA分子量理论值相差132。测序结果表明,rhNDPK-A N末端缺失Met残基,其理论分子量为17017,与飞行质谱测定结果完全一致。表观分子量测定结果表明,rhNDPK-A在溶液中形成六聚体,表观分子量为102kD。上述结果说明, NDPK-A重组产物具与天然产物相同的自发形成六聚体性质,这为NDPK-A新药开发和机理研究打下了良好基础。     

Effects of haematocrit value and glucagon on the metabolism of perfused rat liver

Liver from adult male rats were perfused in situ for 30 min with either undiluted, defibrinated rat blood (haematocrit value 38%) or the same blood diluted with buffer to give a haematocrit of 20%. Perfusion with diluted blood lowered the PO2 of the effluent perfusate but this was insufficient to prevent the fall in O2 consumption due to the reduction in haematocrit. Glucagon (5 X 10(-9) M) increased hepatic O2 consumption with whole blood but not with diluted blood. perfusate K+ was increased by perfusion with diluted blood and glucagon. Bile flow was depressed and biliary K+ increased by glucagon but only in experiments with whole blood. Perfusate glucose was raised by lowering of hepatic O2 consumption but the hormonal stimulation of glucose output was the same at both haematocrits. Net ketogenesis was increased with perfusion with diluted blood and by glucagon. In the absence of glucagon there was a net secretion of triacylglycerols which was depressed by lowering of the haematocrit. Glucagon inhibited triacylglycerol secretion and the effect was greater with whole blood so that there was net uptake. While effects of glucagon were obtained during perfusion at a lower haematocrit, it would appear that whole blood was the medium that allowed their fullest expression.     

Virulence of Agrobacterium tumefaciens strains with Brassica napus and Brassica juncea

Summary Brassica napus and Brassica juncea were infected with a number of Agrobacterium tumefaciens strains. Tumourigenesis was very rapid and extremely efficient on B. juncea with all but one of the strains. Tumourigenesis on B. napus varied widely. It was very efficient with the nopaline strains, was reduced with the succinamopine strain A281 and was very weak with the octopine strains. The latter observation was confirmed with six different B. napus rapeseed cultivars. The selectivity was due to differences in the virulence of Ti plasmids with B. napus, rather than the tumourigenicity of the T-DNA or virulence of the chromosomal genes associated with the strains. An exception was strain LBA4404. The virulence of the octopine strains was increased by coinfection with more virulent disarmed strains and by induction with acetosyringone.     

Hyperreactivity of the bronchial tree in patients with hay fever before and after immunotherapy

Immunotherapy with Pollinex was performed in 46 patients with hay fever. Hyperreactivity of the bronchi determined with inhalation histamine test was noted in 36% of the patients. Histamine test was positive in 24% of patients treated with Pollinex. The difference was statistically insignificant.     

Inhalation anesthesia in adult cattle

An inhalation technique was used for anesthesia during ileal cannulation in five adult cows. Following sedation with intravenous acepromazine, anesthesia was induced intravenously with thiopental sodium in 5% glyceryl guaiacolate solution. Endotracheal intubation was performed and anesthesia maintained with halothane in oxygen via a circle system with a precision vaporizer. In all cases, induction was smooth and no difficulties were experienced during the maintenance of anesthesia. Total anesthesia time was 1.5 to 2.5 hours. Following completion of the surgical procedure, which was performed with the animal in left lateral recumbency, each cow was rolled to a sternal position and supported, if necessary. The endotracheal tube was left in place, with oxygen administration continued, until the animal was able to swallow. Recoveries were rapid and all animals were ambulatory within 30 minutes after completion of the surgery. The only post-operative complication due to anesthesia was transient mouth soreness in two cases, attributed to the use of a mouth speculum during intubation.     

The Effect of Nonanal on Dipodascus aggregatus IV. Studies with Different Carbon Sources

Nonanal (80 μ) in ethanolic solution stimulated the growth of Dipodascus aggregatus with fructose (55.5 mM) as carbon source (inoculum grown with fructose or glucose). If the inoculum had been grown with galactose, neither growth with galactose nor growth with glucose was affected by nonanal. If the inoculum had been grown with glucose, growth with galactose was weakly. stimulated. —Growth with galactose (galactose-grown inoculum) was strongly stimulated by nonanal if xylose at a low concentration (0.53 mM) was added. — The oxygen uptake of glucose grown cells with glucose as substrate was stimulated by 200 μM nonanal in the absence of ethanol. The respiratory activity of galactose-grown cells was also stimulated with galactose as well as with glucose as substrate. In the absence of exogenous substrate the oxygen uptake of glucose-grown cells was weakly stimulated by nonanal whereas that of galactose-grown cells was strongly stimulated.     

Modification of cyclosporin A and conjugation of its derivative to HPMA copolymers

Cyclosporin A (CsA) was epoxidized with m-chloroperoxybenzoic acid in the presence of sodium carbonate or with tert-butyl hydroperoxide in the presence of dioxomolybdenum iminodiethanoxide. The CsA epoxide was not stable and rearranged into a compound with a more stable five-member ring structure. An amino group containing cyclosporin A derivative (CsA amine) was obtained by the reaction of CsA epoxide with excess ethylenediamine. The yield of the CsA amine was 30--40% based on the CsA. An HPMA copolymer--CsA conjugate was prepared by the reaction of the CsA amine with an HPMA and MA-Gly-Phe-Leu-Gly-ONp copolymer. The content of CsA amine in the conjugate was 8.7 wt %. The CsA amine was released from the copolymer by enzymatic hydrolysis with papain.     

肝素钠树脂精制工艺的研究

利用 D2 5 4 树脂可以将原始粗品肝素钠的效价提高到 179.8U SP U/m g,进一步纯化可得 2 0 1.5U SP U/m g的精品 ,其 H2 O2 用量 ( 1.5% )和氧化时间 ( 2 4 h)均少于传统工艺 ,产品光吸收和收率 ( 85.8% )最优     

Reduction of nitro-BT by various chloroform-methanol fractions obtained from the grey matter of the rat brain

Histochemical studies have shown, that nerve fibers can not be visualised in the grey matter of the brain by the reduction of Nitro-BT, if prior to incubation an extraction with chloroform-methanol of the brain slices was performed. According with this finding the grey matter of the rat brain was homogenized and extracted with chloroform-methanol. After centrifugation the obtained supernatant was three times evaporated. Each time before evaporation the supernatant was dissolved in chloroform-methanol solution v/v 2:1. The substance obtained was of teer-like consistency and brown colour. The substance was weight and used for further experiments. Similar extraction was done with phosphate buffer. It was found, that the phosphate buffer does not extract the searched for substance. The comparison of the extinction curves of various fractions has shown that the highest concentration of the substance which reduces Nitro-BT can be found in that fraction which was three times washed with chloroform-methanol. The same concentration was observed in the water fraction which was the result of purification with hexanmethanol. From our investigations it may be concluded that the substance which reduces Nitro-BT in tissue is bound with lipids and purified form lipids passes into water.     

Stem diameter variations and cold hardiness in walnut trees

The effect of freezing temperatures on stem diameter was measured in the field and in climatic chambers using linear variable differential transformers (LVDT sensors). In acclimated stems, there was reversible stem shrinkage associated with freeze-thaw cycles. The maximum shrinkage correlated with stem diameter (thickness of the bark). The wood was responsible for only 15% of the shrinkage associated with a freeze event, and experiments with isolated bark showed that connection with the wood was not necessary for most of the freeze-induced shrinkage to occur. Considering the amount of stem shrinkage associated with summer drought in walnut, the amount of contraction of the bark with freezing was actually much less than might be predicted by water relations theory. Reversible stem shrinkage occurred in living tissues, but not in autoclaved tissues. For the latter, swelling was observed with freezing and this swelling could be explained by the bark alone. Similar swelling was observed during September and October for non-acclimated plants. Water was lost with each freeze-thaw cycle starting with the first, and freezing injury of the bark, with discoloration of tissues, was also observed in non-acclimated plants. Given that the diameter fluctuation patterns were dramatically different for acclimated versus non-acclimated plants, and for living versus autoclaved tissues, LVDT sensors could represent a novel, non-invasive approach to testing cold hardiness.     

三种方法诊断不孕症患者子宫内膜息肉的临床价值研究

目的:探讨单用阴道超声(TVS)、子宫输卵管造影(HSG)、超声子宫水造影(SIS)以及三种方法联合诊断不孕症患者子宫内膜息肉(EP)的临床价值。方法:以206例行宫腔镜联合诊刮或病检的不孕症患者为研究对象,回顾性分析各种检查方法对EP的筛查结果,评价各种检查方法的真实性、可靠性以及预测值。结果:206例不孕症中,共确诊EP患者60例,阳性率29.1%。三种检查方法中,TVS的灵敏度最高(70.0%),特异度最低(73.3%),漏诊率最低(30.0%),误诊率最高(26.7%),正确诊断指数最高(43.3%),阴性似然比最小(0.409),阴性预测值最高(85.6%);SIS检查的灵敏度最低(38.7%),漏诊率最高(61.3%),但是特异性最高(93.3%),误诊率最低(6.7%),阳性似然比最大(4.284),阳性预测值最大(66.6%),正确诊断指数最低(32.0%);HSG检查的上述各项评价指标均介于TVS和SIS之间。TVS和SIS与金标准的符合率低,Kappa值均小于0.4;HSG符合率最高(86.2%),Kappa值0.647。三种检查联合诊断的灵敏度89.3%,漏诊率10.7%,特异度91.4%,误诊率8.6%,正确诊断指数80.7%,阳性似然比10.384,阴性似然比0.117,符合率89.3%,Kappa值0.792,阳性预测值83.3%,阳性预测值94.6%。结论:对于宫腔可能存在内膜息肉的不孕症患者,单一采用阴道超声检查、子宫输卵管造影或超声子宫水造影方法的灵敏度均较低,漏诊率高,与金标准的一致性较差,而三种方法联合用于诊断不孕症患者EP的真实性、可靠性及预测值均较好。     

Optimum conditions for high-resolution gradient analysis

The effect of experimental variables in the photometric scanning of centrifuged sucrose gradients on the apparent resolution was studied. Better resolution was obtained with a flow cell with a comparatively large diameter flow path that was designed for bulk flow than with a flow cell with a comparatively small diameter flow path that was designed for laminar flow. Degradation of resolution caused by an increase in the flow rate was more apparent with the laminar flow cell than with the bulk flow cell. The resolution increased as the illuminated volume of the flow cell decreased but was relatively insensitive to illuminated volume at the lower values tested. Resolution was determined by the number of pea plyribosomes resolved and the shape of their peaks and by the deterioration in shape of peaks given by zones of southern bean mosaic virus during repetitive scanning. It was found that resolution of ribosomal peaks is not a good quantitative method for characterizing instrumental performance.     

Amino acid sequence of the diazooxonorleucine binding site of Acinetobacter and Pseudomonas 7A glutaminase--asparaginase enzymes

Acinetobactor glutaminase-asparaginase was treated with [6-14C]diazo-5-oxonorleucine, reduced with sodium borohydride, and cleaved with cyanogen bromide. Radioactivity was present only in a 96-residue-N-terminal peptide which eluted as the second peptide peak on Sephadex G-50. Radioactivity was released with the threonine in position 12 during automatic sequencing of this peptide. The amino acid sequence of a 60-residue tn-terminal segment and a 16-residue C-terminal segment of this peptide was determined. Pseudomonas 7 A glutaminase-asparaginase was treated with [6-14C]diazo-5-oxonorleucine and reduced with sodium borohydride. Radioactivity was released with the threonine in residue 20 during automatic sequencing of the whole enzyme. Analysis of 26 N-terminal residues showed that an 8-residue segment containing the radioactive threonine was identical with that in Acinetobacter glutaminase-asparaginase and in Escherichia coli asparaginase. Additional identical residues were noted in the N-terminal regions of these enzymes.     

Development of biosensor based on immobilized L-glutamate oxidase for determination of monosodium glutamate in food

A monosodium glutamate (MSG) biosensor with immobilized L-glutamate oxidase (L-GLOD) has been developed and studied for analysis of MSG in sauces, soup etc. The immobilized enzymatic membrane was attached with oxygen electrode with a push cap system. The detection limit of the sensor was 1 mg/dl and the standard curve was found to be linear upto 20 mg/dl. Response time of the sensor was 2 min. Cross-linking with glutaraldehyde in presence of Bovine Serum Albumin (BSA) as a spacer molecule has been used for immobilization. Optimization of the sensor was done with an increase in L-GLOD concentration (6.3-31.5 IU) and also with increase in loading volume of enzyme solution (5-20 microl). Optimization of pH and temperature was also studied. The permeability of O2 through different membrane was studied with and without immobilized L-GLOD. The enzymatic membrane was used for over 20 measurements and stability of the membrane was observed.     

Fe3+络合萃取法从野葛根中分离葛根素

利用Fe3 + 能够和葛根素生成可溶性络合物的性质建立了一种从中药野葛根中萃取葛根素的新型分离方法。以甲醇冷浸从野葛根中提取葛根总黄酮 ,将其进行水解、中和 ,再给水解葛根总黄酮中加入FeCl3 使葛根素与Fe3 + 络合溶解 ,过滤除去其它不溶性物质 ,用盐酸解聚Fe3 + 葛根素络合物 ,则得葛根素粗品 ,将其重结晶可得葛根素。同时 ,利用分光光度法确定了Fe3 + 葛根素络合物解聚的最佳酸度。利用TLC标准品对照、IR和分光光度法对产品进行了定性和定量分析。该方法从葛根中提取葛根素收率为 1 2 % ,纯度为 96 5 % ,具有操作简便、工艺流程简单 ,容易实现工业化的优点。     

The reactivation of nitrate reductase from spinach (Spinacia oleracea L.) inactivated by NADH and cyanide,using trivalent manganese either generated by illuminated chloroplasts or as manganipyrophosphate

Nitrate reductase of spinach (Spinacia oleracea L.) leaves which had been inactivated in vitro by treatment with NADH and cyanide, was reactivated by incubation with oxidant systems and measured as FMNH₂-dependent activity. Reactivation was produced with trivalent manganese compounds represented either by manganipyrophosphate or produced by oxidation of Mn²⁺ ions in the presence of illuminated chloroplasts and compared with reactivation obtained with ferricyanide. Reactivation in the chloroplast system was equivalent to that with ferricyanide when orthophosphate was used but was variable and weak in the presence of pyrophosphate, although manganipyrophosphate was formed, freely. Reactivation by manganipyrophosphate in dark reaction conditions was less effective than with ferricyanide but was not inhibited by the addition of pyrophosphate. Reactivation with illuminated unheated chloroplasts was dependent on added manganese and oxidation of manganese in the presence of pyrophosphate was abolished by boiling the chloroplasts. In the presence of orthophosphate however, boiled, illuminated chloroplasts reactivated the enzyme in the absence of added manganese. Reactivation occurred spontaneously in air, more slowly than with the other oxidants, but to a similar extent to that produced by manganipyrophosphate. The results provide a possible model for physiological reactivation mechanisms.     

Intermittent cyclophosphamide in refractory rheumatoid arthritis.

Three patients with refractory rheumatoid arthritis were treated with oral cyclophosphamide; in two cases this was supplemented with pulse treatment with methylprednisolone. Long term remission was induced in all three patients and was sustained until follow up at least nine months after the methylprednisolone was stopped. Leucopenia occurred but resolved when cyclophosphamide was reduced from daily to intermittent dosing. Intermittent treatment with cyclophosphamide, possibly in conjunction with pulses of methylprednisolone, may induce remission in patients with rheumatoid arthritis refractory to other forms of treatment.