SC-19220, a prostaglandin E2 antagonist, inhibits osteoclast formation by 1,25-dihydroxyvitamin D3 in cell cultures.
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1,25 Dihydroxy vitamin D3 (1,25(OH)2D3), prostaglandin (PG) E2 and parathyroid hormone (PTH) induce osteoclast formation in cell cultures. Previously, we have shown that SC-19220, an antagonist of the EP1 subtype of PGE receptors, inhibited tartrate-resistant acid phosphatase (TRAP)-positive cell formation by PGE2 and PTH in adherent cell cultures taken from neonatal rats. Since 1,25(OH)2D3 has been shown to induce osteoclast formation through PGE2 synthesis, in this study we have examined the effect of SC-19220 on osteoclast formation induced by 1,25(OH)2D3 in cell cultures by measuring bone resorption as well as TRAP-positive cell formation. SC-19220 inhibited osteoclast formation by 1,25(OH)2D3 as well as by PGE2 in cell cultures. The addition of SC-19220 to the later half but not to the earlier half of the culture inhibited 1,25(OH)2D3-induced formation. In the culture in which hydroxyurea was added in the later half period, SC-19220 inhibited osteoclast formation by 1, 25(OH)2D3. Under these conditions, 17-phenyl PGE1, an EP1 agonist, induced osteoclast formation. Thus, SC-19220 inhibits certain reactions in the later processes of osteoclast formation induced by 1,25(OH)2D3. In addition, SC-19220 also inhibited osteoclast formation induced by interleukin (IL)-11 and IL-6 as well as by PTH. It is suggested that the SC-19220 inhibiting reactions are shared by all the inducers including 1,25(OH)2D3 and are essential for osteoclast formation. (+info)
Effects of anti-epileptic drugs on glutamine synthetase activity in mouse brain.
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1. Glutamine synthetase (GS) is a key enzyme in the regulation of glutamate neurotransmission in the central nervous system. It is responsible for the conversion of glutamate to glutamine, and for the detoxification of ammonia. 2. We have investigated the effects of single and repeated intraperitoneal administration of a range of established and new anti-epileptic drugs on GS activity in mouse brain. 3. Four hours after the final dose, animals were sacrificed and the brains removed for analysis of GS activity. 4. Both single and repeated doses of phenytoin and carbamazepine were found to reduce enzyme activity (P<0.05). 5. Single doses of phenobarbitone, felbamate and topiramate were without effect, however repeated administration of these drugs dose-dependently reduced GS activity (P<0.05). 6. Single and repeated doses of sodium valproate, vigabatrin, lamotrigine, gabapentin, tiagabine, levetiracetam and desglycinyl-remacemide were found to have no effect on GS activity. 7. The reduction in enzyme activity demonstrated is unlikely to be related to the anti-epileptic actions of these drugs, but may contribute to their toxicity. (+info)
Role of group II secretory phospholipase A2 in atherosclerosis: 1. Increased atherogenesis and altered lipoproteins in transgenic mice expressing group IIa phospholipase A2.
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Some observations have suggested that the extracellular group IIa phospholipase A2 (sPLA2), previously implicated in chronic inflammatory conditions such as arthritis, may contribute to atherosclerosis. We have examined this hypothesis by studying transgenic mice expressing the human enzyme. Compared with nontransgenic littermates, the transgenic mice exhibited dramatically increased atherosclerotic lesions when maintained on a high-fat, high-cholesterol diet. Surprisingly, the transgenic mice also exhibited significant atherosclerotic lesions when maintained on a low-fat chow diet. Immunohistochemical staining indicated that sPLA2 was present in the atherosclerotic lesions of the transgenic mice. On both chow and atherogenic diets, the transgenic mice exhibited decreased levels of HDLs and slightly increased levels of LDLs compared with nontransgenic littermates. These data indicate that group IIa sPLA2 may promote atherogenesis, in part, through its effects on lipoprotein levels. These data also provide a possible mechanism for the observation that there is an increased incidence of coronary artery disease in many chronic inflammatory diseases. (+info)
Immunogenicity, genetic stability, and protective efficacy of a recombinant, chimeric yellow fever-Japanese encephalitis virus (ChimeriVax-JE) as a live, attenuated vaccine candidate against Japanese encephalitis.
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Yellow fever (YF) 17D vaccine virus, having a 60-year history of safe and effective use, is an ideal vector to deliver heterologous genes from other medically important flaviviruses. A chimeric YF/Japanese encephalitis (JE) virus (ChimeriVax-JE virus) was constructed by insertion of the premembrane and envelope (prME) genes of an attenuated human vaccine strain (SA14-14-2) of Japanese encephalitis (JE) virus between core and nonstructural (NS) genes of a YF 17D infectious clone. The virus grew to high titers in cell cultures and was not neurovirulent for 3- to 4-week-old mice at doses /=10(3) pfu of ChimeriVax-JE virus were solidly protected against intraperitoneal challenge with a virulent JE virus. Genetic stability of the chimera was assessed by sequential passages in cell cultures or in mouse brain. All attenuating residues and the avirulent phenotype were preserved after 18 passages in cell cultures or 6 passages in mouse brains. (+info)
Nuclear translocation of nuclear factor kappa B in early 1-cell mouse embryos.
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Nuclear factor kappa B (NF-kappaB) is a transcription factor that controls the expression of a number of genes under cellular redox potential. It has recently been found that NF-kappaB plays a pivotal role in morphogenesis and embryonic development, e.g., in formation of Drosophila malanogaster ventral structures and chicken limb buds. However, the role of NF-kappaB in preimplantation development in mammals is not yet understood. In this study, we show that RelA, one of the subunits of NF-kappaB, is expressed in mouse eggs and embryos from the metaphase II (MII) oocyte to the blastocyst stage. Therefore, it is thought that RelA is maternally expressed and that it continues to be expressed during preimplantation development. Immunofluorescence analysis showed that RelA protein was mainly distributed in the cytoplasm of embryos, whereas nuclear translocation of RelA, evidence for NF-kappaB activation, was observed only at the early 1-cell stage. Finally we studied the effects of NF-kappaB inhibitors, pyrrolidine dithiocarbamate and N-acetyl-L-cysteine, on the preimplantation development of mouse embryos. When these inhibitors were added to the culture medium from the early 1-cell stage, subsequent development through the 2-cell stage was inhibited. However, little, if any, influence on the progression through the 2-cell stage was observed when the inhibitors were added at the late 1-cell or the 2-cell stage. Taken together, the results suggest that the activation of NF-kappaB at the early 1-cell stage is required for the development of mouse embryos beyond the 2-cell stage. (+info)
Immunolocalization of CRES (Cystatin-related epididymal spermatogenic) protein in the acrosomes of mouse spermatozoa.
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The CRES (cystatin-related epididymal spermatogenic) protein is a member of the cystatin superfamily of cysteine protease inhibitors and exhibits highly restricted expression in the reproductive tract. We have previously shown that CRES protein is present in elongating spermatids in the testis and is synthesized and secreted by the proximal caput epididymal epithelium. The presence of CRES protein in developing germ cells and in the luminal fluid surrounding maturing spermatozoa prompted us to examine whether CRES protein is associated with spermatozoa. In the studies presented, indirect immunofluorescence, immunogold electron microscopy, and Western blot analysis demonstrated that CRES protein is localized in sperm acrosomes and is released during the acrosome reaction. Interestingly, while the 19- and 14-kDa CRES proteins were present in testicular and proximal caput epididymal spermatozoa, the 14-kDa CRES protein was the predominant form present in mid-caput to cauda epididymal spermatozoa. Furthermore, following the ionophore-induced acrosome reaction, CRES protein localization was similar to that of proacrosin/acrosin in that it was detected in the soluble fraction as well as associated with the acrosome-reacted spermatozoa. The presence of CRES protein in the sperm acrosome, a site of high hydrolytic and proteolytic activity, suggests that CRES may play a role in the regulation of intraacrosomal protein processing or may be involved in fertilization. (+info)
DNA-mediated immunization of mice with plasmid encoding HBs antigen.
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In order to develop an experimental DNA vaccine for the prevention and treatment of hepatitis B virus infection, hepatitis B virus surface antigen (HBsAg) DNA was subcloned into an E. coli-eukaryotic cell shuttle vector and was expressed in the Baculovirus expression system. Intramuscular, intradermal, and intraperitoneal injections of 30 microg of the plasmid DNA expressing HBsAg induced humoral and cellular immune responses in ICR mice. The first IgG antibodies were detected after ten days and specific IgG antibody titers peaked after two months of a single intramuscular DNA injection. Anti-HBs antibody titers gradually increased and peaked at four months following intradermal DNA injection, and in case of intraperitoneal injection they peaked at seven months. Generation of HBs-specific helper T lymphocytes was also investigated through the production of interleukin-2 by T helper cells. Boosting effects of HBs DNA were investigated without much results. In general, DNA-mediated HBs immunization induced humoral and cellular immune responses in mice that appears to simulate immune responses in human during the course of HBV vaccination. (+info)
Effect of intracellular Ca2+ chelation with the acetoxymethyl ester-derived form of bis(o-aminophenoxy)ethane-N,N,N,N',N'-tetraacetic acid on meiotic division and chromosomal segregation in mouse oocytes.
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PURPOSE: Our purpose was to ascertain the effect of intracellular Ca2+ chelation on the chromosomal distribution and segregation of mouse oocytes during maturation in vitro. METHODS: Germinal vesicle oocytes were loaded with the acetoxymethyl ester-derived form of bis(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA-AM). Chromosomal distribution and segregation of control and BAPTA-AM-treated metaphase II (MII) oocytes were evaluated at 16 hr, and intracellular ATP content at 0, 1, and 16 hr after BAPTA-AM loading. RESULTS: BAPTA-AM treatment decreased (P < or = 0.05) the potential for in vitro maturation, increased (P < or = 0.0001) the percentage of oocytes displaying an abnormal distribution of metaphase II chromosomes in the meiosis II spindle and aneuploidy, and decreased (P < or = 0.005) the ATP content at 0, 1, and 16 hr of culture compared to the control groups. CONCLUSIONS: These findings raise some concern about any other condition/drug that may directly or indirectly decrease the intracellular Ca2+ concentration in human oocytes. (+info)