利用带有原核增强子样序列的新型载体在大肠杆菌中高效表达人α肿瘤坏死因子

将去除信号肽的人肿瘤坏死因子-α(Tumor necrosis factor-α,TNF-α)cDNA插入到带有原核增强子样序列Px的新型表达载体pBV320中,使TNF cDNA 5′端直接置于大肠杆菌trp启动子下游,采用37℃恒温培养,使TNF在大肠杆菌中获得了高效表达,表达活性达1.35(±0.17)×10~6U/L菌液。表达的TNF-α对L929细胞的毒性作用可被抗人肿瘤坏死因子-α的单克隆抗体所中和。表达菌裂解液作SDS-聚丙烯酰胺凝胶电泳,显示有一条分子量与TNF分子量吻合、约为17000道尔顿的蛋白带。利用DEAE-Sepharose阴离子交换层析及Sephacryl S-200凝胶过滤对上述重组人TNF-α进行纯化,获得电泳纯产品,比活性为1.48×10~6U/mg。     

Utilization of alpha-ketoglutarate as a precursor for transmitter glutamate in cultured cerebellar granule cells

Alpha-ketoglutarate together with an amino group donor (alanine) was shown to be able to serve as a precursor for the glutamate pool which is released by potassium-induced depolarization (i.e., transmitter glutamate) in cerebellar granule cells. However, these compounds could not be utilized as precursors for intracellular glutamate or for release of transmitter aspartate. The formation of transmitter glutamate was inhibited by the transamination inhibitor aminooxyacetic acid but not by phenylsuccinate, an inhibitor of the dicarboxylate carrier in the mitochondrial membrane. Both of these inhibitors have previously been found to inhibit synthesis of transmitter glutamate from glutamine. The results support the hypothesis that alpha-ketoglutarate and alanine undergo transamination in the cytosol to form pyruvate and glutamate, and that this glutamate pool is available for transmitter release of glutamate but does not constitute the major intracellular pool of glutamate.     

Alpha-2-adrenergic modulation of the parathyroid hormone-inhibition of phosphate uptake in cultured renal (OK) cells

Parathyroid hormone enhances the formation of cAMP and decreases the Na+-dependent uptake of phosphate in cultured renal cells derived from the American opossum (OK cells). Epinephrine, acting as an alpha 2-adrenergic agonist, inhibits the PTH-induced synthesis of cAMP by a pertussis toxin-sensitive mechanism and blunts the inhibition of phosphate transport by PTH. Na+-dependent alpha-methylglucoside and Na+ uptakes by the cells are unaffected by PTH and epinephrine. These findings suggest that alpha 2-adrenergic agonists may selectively modulate PTH-sensitive phosphate transport in the renal proximal tubule.     

Computer aided analysis for biosensing and screening

Biosensors with animal and microbial cells immobilized close to the tip of a membrane electrode have been developed for chemical and drug testing. Our experimental results show that biosensors can be used for drug screening and to provide useful information about various cell-chemical interactions. A computer aided analysis (CAA) software package is being developed here using the biosensor for various screening purposes. This software package enables us to use a computer to analyze the biosensor dynamic responses. Computer simulation and parameter estimation techniques are used to select the best model and to describe the biochemical and pharmacologic effects of various chemicals and drugs on different cell lines.     

To early distinguish neonatal transient leukemoid proliferation from congenital leukemia by in vitro cell growth

To differentiate neonatal transient leukemoid proliferation from congenital leukemia at an early stage is often difficult. Bone marrow culture is found to be helpful in this aspect. A normal in vitro growth pattern suggests transient leukemoid proliferation, while an abnormal growth pattern indicates congenital leukemia. A neonate who manifested with pictures mimicking acute myeloblastic leukemia (M1), had a karyotype of 46, XY/46, XY, i(21 q). However, the in vitro growth pattern was normal and so only supportive treatment was given. All the leukemoid manifestations disappeared several months later and he is now a healthy 2 year old boy remaining in complete remission. A second neonate who also displayed features of acute myeloblastic leukemia (M2), had a karyotype of 46, XY/47, XY, + 21 and abnormal in vitro growth pattern. This neonate died at 18 days of age.     

Aneuploidy in mammalian somatic cells in vivo

Aneuploidy is an important potential source of human disease and of reproductive failure. Nevertheless, the ability of chemical agents to induce aneuploidy has been investigated only sporadically in intact (whole-animal) mammalian systems. A search of the available literature from the EMCT Aneuploidy File (for years 1970-1983) provided 112 papers that dealt with aneuploidy in mammalian somatic cells in vivo. 59 of these papers did not meet minimal criteria for analysis and were rejected from subsequent review. Of the remaining 53 papers that dealt with aneuploidy induction by chemical agents in mammalian somatic cells in vivo, only 3 (6%) contained data that were considered to be supported conclusively by adequate study designs, execution, and reporting. These 3 papers dealt with 2 chemicals, one of which, mercury, was negative for aneuploidy induction in humans, and the other, pyrimethamine, was positive in an experimental rodent study. The majority of papers (94%) were considered inconclusive for a variety of reasons. The most common reasons for calling a study inconclusive were (a) combining data on hyperploidy with those on hypoploidy and/or polyploidy, (b) an inadequate or unspecified number of animals and/or cells per animal scored per treatment group, and (c) poor data presentation such that animal-to-animal variability could not be assessed. Suggestions for protocol development are made, and the future directions of research into aneuploidy induction are discussed.     

Studies of structure-activity relationships of human interleukin-2

Human interleukin-2 (IL-2) has 3 cysteine residues; cysteines 58 and 105 form an intramolecular disulfide bridge, whereas cysteine 125 has a free sulfhydryl group. In this study, site-specific mutagenesis has been used to modify the cysteine residues of recombinant Escherichia coli-derived IL-2 (rIL-2) to evaluate the functional structure of IL-2. Substitution or deletion of cysteine 105 disrupted the disulfide bridge and yielded a mutant protein which was 8-10 times less active than wild type rIL-2. A similar modification at position 58, however, reduced the activity of rIL-2 by more than 250-fold. Although substitution of serine for cysteine 125 did not affect IL-2 activity, deletion of cysteine 125 or deletion of amino acids in the vicinity of this cysteine yielded mutant proteins with little, if any, activity. These results indicate that the protein structure in the vicinity of both positions 58 and 125 is more critical than that close to position 105. These findings may provide a clue to the understanding of the functional structure of human IL-2.     

广东楝科植物分类的初步研究

本文是对广东楝科植物种类的初步整理、研究,记载了16属33种7交种,包括本省产的13属27种7变种和外来引种的3属6种。其中有1个新变种(封开地黄连Munronia hainanensis How et T. Chen var. microphyllina X. M. Chen),3个新组合(曾椤Amoora tsangii (Merr.) X. M. Chen, 雷楝Reinwardtiodendron dubium (Merr.) X. M. Chen, 毛香椿Toona sinensis (A. Juss.) M. J. Roem. var. schensiana (C. DC.) X. M. Chen), 3个分布新记录[雷楝属Reinwardtiodendron (我国新记录),鹧鸪花Trichilia connaroides (W. et A.) Bentv. var. connaroides(广东新记录),海南(木坚)木Dysoxylvm hainanense Merr. var. hainanense(广东大陆新记录)].