The Japanese journal of experimental medicine最新文献

A hybrid plasmid was constructed by insertion of the HBc gene encoding HBcAg into the pKK223-3 plasmid at the SmaI cleavage site in the correct direction just downstream from the tac promoter and upstream from the rrnB terminator. The recombinant plasmid carrying the HBc gene was introduced into E. coli and cloned. HBcAg was synthesized in E. coli by using the expression plasmid under the regulation of the tac promoter and rrnB terminator. The tac promoter, derived from sequences of trp and lac UV5 promoters, has identical sequences in two domains (-35 and -10 regions) with optimal distance, and the Shine-Dalgarno sequence, which enables protein synthesis to start at the ATG of the adjacent HBc gene. The nucleotide sequence of the HBc gene and its predicted amino acid sequence were almost identical to those previously reported. Purified HBcAg has a molecular weight of 21,500. This polypeptide gave a positive reaction with anti-HBcAg and anti-HBe antibodies, and was assembled into spherical particles 37 nm in diameter. The recombinant plasmid, carrying the HBc gene between the tac promoter (trp-lac hybrid promoter) and the rrnB terminator in expression plasmid pKK223-3, was useful for efficient expression of the HBc gene and production of HBcAg particles in E. coli.

T-2 toxin (T-2(4)), one of the trichothecene mycotoxins produced by various gerera of Fusarium spp., is a potent inhibitor to the syntheses of protein and DNA in mammalian cells. The selective cytotoxicity of T-2 toxin-conjugated anti-EL-4 monoclonal antibodies (T-2-mAb) was investigated against murine thymoma EL-4 cells in vitro and in vivo systems. At first T-2 was converted to T-2 hemiglutarate by glutaric anhydride. Then T-2 hemiglutarate was activated to 3-[4-(N-succinimidoxycarbonyl)-butyryl]-T-2 (T-2-G-OSu) by N-hydroxysuccinimide. Thus obtained T-2-G-OSu was conjugated with mAb specific for EL-4 cells. The T-2-mAb markedly inhibited the proliferation of cultured EL-4 cells, but no such cytotoxic effect was observed against irrelevant SP2/0 cells. The cytotoxicity of T-2-conjugated normal gamma globulin (T-2-nGG) against EL-4 cells was far less than that of the above T-2-mAb. Ammonium chloride and monensin, inhibitors of lysosomal enzymes, enhanced the cytotoxicity of T-2-mAb. The presence of both 2-deoxyglucose together with sodium azide, inhibitors of energy-dependent reaction, reduced the cytotoxicity of T-2-mAb. Therefore, the selective binding to the target cells followed by an energy-dependent endocytosis and an intracellular liberation of T-2 by hydrolysis may account for the cytotoxicity of the T-2-mAb. In mice pre-transplanted with EL-4 cells, T-2-mAb increased the mean survival time (MST) with a direct dosage dependence.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of oxygen free radicals in inhibition of proliferation of Meth A tumor cells was studied. Diethyldithiocarbamate (DDC), a chelator which inactivates superoxide dismutase, was used to examine the effect of tumor necrosis factor (TNF), lymphotoxin preparation derived from a human lymphoid cell line (c-LT) and glycyrrhizin (GL) on in vitro proliferation of Meth A tumor cells. High degree of antiproliferative effect was observed when DDC was added simultaneously with TNF to target cells. Similar effect was obtained by the addition of GL. However, augmentation of antiproliferative effect was not observed when c-LT was added. However, augmentation of antiproliferative effect was observed when the target cells had been treated with DDC in advance of the addition of TNF, GL or c-LT. Roles of oxygen free radicals in inhibition of tumor cell proliferation were discussed.

A comparative study of specific and non-specific immunosuppression has been carried out in acute and chronic Plasmodium berghei infected mice in an in vivo system. In our previous studies, immunosuppression during acute P. berghei infection was attributed to T lymphocytes when we studied modulation of blastogenic response of lymphocytes in an in vitro system. In the present study, delayed type hypersensitivity (DTH) was evident from the homing of radiolabelled bone marrow cells into the delayed lesions of antigen challenged foot pads during chronic infection. This response was suppressed during acute infection especially in early stages. A greater concentration of bone marrow cells in the liver and spleen occurred during chronic infection in comparison with acute infection. When radiolabelled bone marrow cells from infected mice were injected into the normal mice previously given malaria antigen in foot pads, no significant change in homing pattern in liver, spleen or foot pads was observed. Contact sensitivity to picryl chloride was suppressed during acute infection, but was intact during chronic infection. Since these responses are mediated by T lymphocytes, significance of these responses is discussed.

Modulation of c-myc gene expression by extracellular stimuli in H4IIE rat hepatoma cells was investigated by Northern blot analysis. Treatment of these cells with phorbol 12-O-tetradecanoate 13-acetate (TPA), insulin and concanavalin A (Con A) resulted in transient accumulation of c-myc transcripts within 2 hours. The induction of c-myc mRNA was dose dependent with similar responses for all three agents. The maximally induced c-myc mRNA levels varied from 5- to 15-fold of the control. Treatment with cycloheximide (10 micrograms/ml) and H7, a protein kinase C inhibitor (20 microM), inhibited this induction, suggesting that c-myc induction by these agents requires protein synthesis and protein kinase C activation.

A female infant with early-onset GM1-gangliosidosis type I was investigated. The lymphocytes, transformed lymphocytes and cultured skin fibroblasts of the patient were demonstrated to have severe beta-D-galactosidase deficiency. The beta-D-galactosidase activities of these cells from the patient's father and mother were at the lower limit of the normal range. The oligosaccharide accumulation in urine of the patient showed the typical type I GM1-gangliosidosis pattern, but no GM1 ganglioside was detected in the patient's urine or transformed lymphocytes. The clinical features were compatible with infantile GM1-gangliosidosis. The mixture of homogenates from the cultured fibroblasts or transformed lymphocytes of the patient and controls showed no complementation of beta-D-galactosidase activity against the controls.

In order to establish an animal model and to identify a causative virus of enterically transmitted non-A, non-B hepatitis, Macaca fascicularis was inoculated with a fecal extract obtained from Myanmar patients with acute sporadic non-A, non-B hepatitis. The primates developed acute hepatitis exhibited by a transient elevation of aminotransferases in the sera and occurrence of hepatic necroinflammation between 2 and 4 weeks postinoculation. Subsequent second passage of the fecal extract made from first-passage primates into another Macaca fascicularis and Macaca mulatta induced acute hepatitis. Likewise, third passage was also successfully performed. Immune electron microscopy of the stool extract incubated with the primate serum at the acute phase of hepatitis showed an aggregation of virus-like particles. These particles consisted of full and empty round particles without an envelope, measuring approximately 27 nm in diameter. A dispersion of similar particles was found ultrastructurally in the hyaloplasm of hepatocytes surrounding the focal necrosis. This putative causative virus appears to be a new hepatitis virus.

Generation of reactive oxygen species (ROS) by blood monocytes and Kupffer cells of normal and Plasmodium berghei infected mice treated at different levels of parasitaemia with chloroquine, were studied. Cells isolated at lower level of parasitaemia (less than 2%) produced ROS within the range of normal animals, whereas ROS production by the cells isolated from the animals at higher level of parasitaemia (greater than 20%), was significantly higher even without stimulation with latex particles. The ROS generation capacity of both normal and infected animals was less after the chloroquine treatment. This inhibitory effect of chloroquine may be beneficial in protecting the host from the adverse effect of reactive oxygen species by controlling their overproduction.

On weeks 7 and 9 post infection (PI), the number of Schistosoma japonicum in mice maintained at low (5 degrees C) and high (32 degrees C and 35 degrees C) temperatures did not differ from those in controls (25 degrees C), but shorter males were observed in hosts at 5 degrees C. Further, both male and female worms recovered on week 7 PI from mice kept at 35 degrees C were shorter than those recovered from controls. Staining analysis revealed that worm maturation was not affected in mice kept at 5 degrees C and 32 degrees C. However, some female worms from mice kept at 35 degrees C had no eggs in the uterus. The day of patency of worms in the temperature-stressed mice did not differ from that in controls. EPG in mice kept at 32 degrees C rose to a high level on weeks 7-8 PI which was about 10 times as many as those in control mice, while EPG in mice kept at 5 degrees C appeared to be lower than in controls. The 7-week tissue egg count revealed that the hot- and cold-stressed mice supported significantly less egg productivity per female worm than control mice. The environmental temperature could not alter the migration pattern of schistosomula from the skin to lungs in mice.