由大肠杆菌中提取重组猪生长激素的研究

 本文通过研究提出了一种由大肠杆菌细胞中提取重组猪生长激素的方法。提取回收率为52.4％,纯度为94.1％,具有生物学活性。     

A novel fluorescent ceramide analogue for studying membrane traffic in animal cells: accumulation at the Golgi apparatus results in altered spectral properties of the sphingolipid precursor

A series of ceramide analogues bearing the fluorophore boron dipyrromethene difluoride (BODIPY) were synthesized and evaluated as vital stains for the Golgi apparatus, and as tools for studying lipid traffic between the Golgi apparatus and the plasma membrane of living cells. Studies of the spectral properties of several of the BODIPY-labeled ceramides in lipid vesicles demonstrated that the fluorescence emission maxima were strongly dependent upon the molar density of the probes in the membrane. This was especially evident using N-[5-(5,7-dimethyl BODIPY)-1-pentanoyl]-D-erythro-sphingosine (C5-DMB-Cer), which exhibited a shift in its emission maximum from green (integral of 515 nm) to red (integral of 620 nm) wavelengths with increasing concentrations. When C5-DMB-Cer was used to label living cells, this property allowed us to differentiate membranes containing high concentrations of the fluorescent lipid and its metabolites (the corresponding analogues of sphingomyelin and glucosylceramide) from other regions of the cell where smaller amounts of the probe were present. Using this approach, prominent red fluorescent labeling of the Golgi apparatus, Golgi apparatus-associated tubulovesicular processes, and putative Golgi apparatus transport vesicles was seen in living human skin fibroblasts, as well as in other cell types. Based on fluorescence ratio imaging microscopy, we estimate that C5-DMB-Cer and its metabolites were present in Golgi apparatus membranes at concentrations up to 5-10 mol %. In addition, the concentration-dependent spectral properties of C5-DMB-Cer were used to monitor the transport of C5-DMB-lipids to the cell surface at 37 degrees C.     

IL-4 down-regulates the expression of J11d on murine B cells.

T cell-derived IL-4 has many effects on murine B cells, including the up-regulation of class II antigens and the induction of isotype switching. The development of memory B cells and the decreased expression of J11d antigens on these cells are influenced by T cells. In this report, we determined whether the decreased expression of J11d can also be mediated by T cell-derived lymphokines and, in particular, IL-4. We found that IL-4 can down-regulate the expression of J11d on both large and small B cells, that this effect becomes significant after 48 hr of culture and occurs at doses of IL-4 that are similar to those required to up-regulate murine class II MHC antigens encoded by I-region alpha genes (IA). Anti-IL-4 antibody completely blocks this effect but IFN-gamma does not. Other lymphokines (IL-1, IL-2, IL-3, IL-5, IL-6) neither induce a decrease in J11d nor alter the ability of IL-4 to down-regulate J11d expression. The decrease in J11d expression on B cells is not due to a preferential survival of cells expressing lower levels of J11d, although IL-4 has a more pronounced effect on these B cells. Finally, the down-regulation of J11d and up-regulation of IA by IL-4 occurs on all inducible B cells.     

四川省自贡市幼儿园3至5岁儿童乙型肝炎病毒感染情况及影响因素

为了解四川省自贡地区3～5岁幼儿乙型肝炎病毒(HBV)感染情况,并探索与感染有关的因素,1985年调查了1167名幼儿,其HBV总感染率为41.13％,HBsAg阳性率12.68％。幼儿的HBV感染与母亲HBsAg阳性密切相关。共检查母亲409例,38例HBsAg阳性,其幼儿HBsAg阳性率为50％(19/38),HBsAg阴性的母亲371例,其幼儿HBsAg阳性率9.97％(37/371),来自HBsAg阳性母亲的阳性子女占33.3％(19/56)。1986年随访HBV易感幼儿448例,HBV年感染率为12.95％(58/448),HBsAg年阳转率3.79％。HBV年感染率与原幼儿班级HBsAg阳性率的高低有关。     

Effect of matrices on affinity purification of protein C

One of the critical problems in scale-up of affinity chromatography is the mechanical strength of the support matrix against pressure. Because the costs of both the gel matrix and the ligand for the affinity chromatography are very high, the reusability of gel matrices is directly related to the total production cost. In certain cases, where the source material is viscous (e.g., blood plasma), irreversible deformation of gel matrices can readily occur, necessitating severe constraints in the flow rate. Consequently, productivity is low.We have characterized the system parameters and investigated the performance of various matrices that are commercially available. The experimental system used for this study was the immunoaffinity purification of protein C (an anticoagulant protein) from human blood plasma. The support matrices studied were cross-linked agarose, polymethyl acrylic, cellulose, and polyvinyl alcohol polymers. The major system parameters studied were pressure tolerance, coupling efficiency, adsorption efficiency, and batch adsorption/desorption kinetics of protein C to/from the monoclonal antibody (MAb)-Matrix complex. In addition, the apparent equilibrium constant and bandwidth of the product concentration profile in the eluate were characterized by performing pulse tests.A methodology was developed for evaluating the immunoaffinity colum performance for the separation of protein C. By utilizing the experimentally measured parameters, the flow rate limitation for each purification step was computed. Then, the purification performance of the matrices were evaluated in terms of productivity per unit time. Among the matrices tested, cellulose was superior in overall performance for the immunoaffinity purification of protein C using a 10 cm x 10 cm column.     

Immunohistochemical localization of 36 and 29 kDa proteins in sparganum.

Antigenic proteins of 36 and 29 kDa were localized in Spirometra mansoni plerocercoid (sparganum) immunohistochemically by avidin biotin complex (ABC) staining. When polyclonal antibodies such as BALB/c mouse serum immunized with crude saline extract of sparganum or confirmed sparganosis sera were reacted as primary antibodies, the positive chromogen (3-amino, 9-ethylcarbazole) reactions were recognized at syncytial tegument, tegumental cells, muscle and parenchymal cells and lining cells of excretory canals. A monoclonal antibody (MAb) which was reacting to 36 and 29 kDa proteins in the extract of the worm was localized at the syncytial tegument and tegumental cells. The present results suggested that the potent antigenic proteins of 36 and 29 kDa in sparganum were produced at the tegumental cells and transported to the syncytial tegument.     

Wild-type p53 restores cell cycle control and inhibits gene amplification in cells with mutant p53 alleles.

Loss of cell cycle control and acquisition of chromosomal rearrangements such as gene amplification often occur during tumor progression, suggesting that they may be correlated. We show here that the wild-type p53 allele is lost when fibroblasts from patients with the Li-Fraumeni syndrome (LFS) are passaged in vitro. Normal and LFS cells containing wild-type p53 arrested in G1 when challenged with the uridine biosynthesis inhibitor PALA and did not undergo PALA-selected gene amplification. The converse occurred in cells lacking wild-type p53 expression. Expression of wild-type p53 in transformants of immortal and tumor cells containing mutant p53 alleles restored G1 control and reduced the frequency of gene amplification to undetectable levels. These studies reveal that p53 contributes to a metabolically regulated G1 check-point, and they provide a model for understanding how abnormal cell cycle progression leads to the genetic rearrangements involved in tumor progression.     

Studies on glycosaminoglycans of regenerating rabbit ear cartilage

Cartilage regeneration in the adult rabbit ear was examined with respect to glycosaminoglycan (GAG) synthesis at various stages of the regeneration process. Increased hyaluronic acid and chondroitin sulfate synthesis was first seen 31 days after wounding, when a metachromatic cartilage matrix could be distinguished from blastemal cells. Analysis of cartilage and the overlying skin separately showed that 90% of the labeled chondroitin sulfate was found in the cartilage being regenerated. DEAE-cellulose chromatography of GAG preparations from 35-day regenerating cartilages showed hyaluronic acid and chondroitin sulfate peaks eluting in the same position as those isolated from normal cartilages. The identity of the hyaluronic acid and chondroitin sulfate peaks was confirmed by their susceptibility to Streptomyces hyaluronidase and chondroitinase ABC, respectively. Although the degree of sulfation in normal and regenerated cartilages was similar, the ratio of chondroitin 6-sulfate to chondroitin 4-sulfate was increased in regenerated cartilages. GAG preparations from unlabeled cartilages were digested with chondroitinase ABC and the disaccharide digestive products were identified and quantitiated. Normal cartilage had a $\text{Δ}\text{Di-6S}\text{Δ}\text{Di-4S}$ ratio of 0.27; the same ratio for the regenerated cartilage was 1.58.     

Electron paramagnetic and electron nuclear double resonance of the hydrogen peroxide compound of cytochrome c peroxidase

We have collected electron paramagnetic resonance (EPR) and electron nuclear double resonance (ENDOR) spectra from the hydrogen peroxide compound of yeast cytochrome c peroxidase, termed ES, employing EPR microwave frequencies of 9.6 and 11.6 GHz. We have measured and analyzed the temperature dependence of the spin-lattice relaxation rate (1/T1) of the paramagnetic center of ES over the temperature range 1.9 to 4 K. In addition, an upper bound to exchange coupling between the ferryl heme and EPR-visible centers of ES has been calculated and expressions for the dipolar interaction between a ferryl heme and a free radical have been derived. These results all confirm that the EPR signal of ES is not associated with an aromatic amino acid radical, and in particular not with a tryptophanyl radical. This conclusion has led us to consider an explanation of the EPR signal in terms of a nucleophilically stabilized methionyl radical.