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Biomedical subjects

X Mao

Publications and source records attributed to X Mao.

At least 55 records · Page 3Linked to original sources
PubMed2000

Expression of the RNA-binding protein TIAR is increased in neurons after ischemic cerebral injury.

T-cell restricted intracellular antigen-related protein (TIAR) is an RNA recognition motif-type RNA-binding protein that has been implicated in the apoptotic death of T-lymphocytes and retinal pigment epithelial cells. Western blots prepared with a monoclonal antibody against TIAR showed expression in normal rat hippocampus, and induction by 15 min of global cerebral ischemia. This increased expression was evident at 8 hr after ischemia and maximal at 24 hr, whereas expression at 72 hr was reduced below basal levels. Expression of TIAR protein was also increased in parietal cortex 6 and 24 hr after 90 min of focal cerebral ischemia induced by middle cerebral artery (MCA) occlusion, as well as in cultured cortical neurons and astroglia after exposure to hypoxia in vitro. Immunocytochemistry showed that increased expression of TIAR occurred mainly in the CA1 sector of hippocampus 24 hr after global ischemia, and in cortical and striatal neurons 24 hr after 20 or 90 min of focal ischemia. Double-labeling studies showed that TIAR protein expression was co-localized with DNA damage in neuronal cells. The findings suggest that TIAR may be involved in neuronal cell death after cerebral ischemic injury.

Animals↗
PubMed2000

Nine novel precursors of Buthus martensii scorpion alpha-toxin homologues.

The cDNAs encoding nine novel alpha-toxin homologues were isolated from the venom gland cDNA library of the Chinese scorpion Buthus martensii Karsch (BmK). They are rich in AAAA and TTTT elements at the 5' UTRs. The flanking region of the translation initiation codon ATG is AAAATGAA, which is highly conserved in scorpion Na(+), K(+) and Cl(-) channel toxin genes. These putative scorpion alpha-toxins shared 45.5-98.4% homology with the characterized BmK alpha-toxins, and were completely conserved in the positions of all eight cysteines. This showed, together with higher homology at nucleotide level than that at amino acid level, that these toxins may originate from a common ancestor. The discovery of a series of homologues of scorpion alpha-toxin with a different degree of natural mutation in the primary structure will provide us with a valuable system for studying the structure-function relationship of scorpion toxins.

Amino Acid Sequence↗
PubMed2000

Blood eicosanoids and immune indices during fasciolosis in water buffaloes.

The effects of trickle infections of water buffaloes with Fasciola hepatica (60 metacercariae daily during a period of 20 days) on the blood plasma levels of prostaglandin E(2) (PGE2), 6-keto-prostaglandin F(1alpha) (6-keto-PG F(1alpha)) and thromboxane B(2) (TXB2) were assessed. F. hepatica specific IgG and T- and B-lymphocyte ratios were evaluated as indicators of the immune response. Although the applied mode of infection did not result in clinical disease, changes in the plasma eicosanoid pattern were observed. Plasma PGE2 values were significantly elevated in the infected water buffaloes 11 weeks post-infection (w.p.i.). In contrast, transiently but significantly lower TXB2 values than in the uninfected controls were recorded in the phase of chronic fasciolosis. Plasma 6-keto-PGF(1alpha) values were not considerably altered by the infection throughout the study period. F. hepatica-specific IgG were detected from 4 to 21 w.p.i. The proportion of peripheral T- and B-lymphocytes shifted towards B-cells from 2 to 12 w.p.i., gradually returning to control values afterwards. Although the water buffaloes appeared to be rather resistant to trickle infection with F. hepatica, moderate changes in plasma eicosanoid patterns were observed, indicating tissue damage and/or inflammation. Induction of the immune response could be monitored by an increase of F. hepatica-specific IgG, which was paralleled by a relative increase of the B-lymphocyte population.

Animals↗
PubMed2000

Structure-function analysis of yeast mRNA cap methyltransferase and high-copy suppression of conditional mutants by AdoMet synthase and the ubiquitin conjugating enzyme Cdc34p.

Here we present a genetic analysis of the yeast cap-methylating enzyme Abd1p. To identify individual amino acids required for Abd1p function, we introduced alanine mutations at 35 positions of the 436-amino acid yeast protein. Two new recessive lethal mutations, F256A and Y330A, were identified. Alleles F256L and Y256L were viable, suggesting that hydrophobic residues at these positions sufficed for Abd1p function. Conservative mutations of Asp-178 established that an acidic moiety is essential at this position (i.e. , D178E was viable whereas D178N was not). Phe-256, Tyr-330, and Asp-178 are conserved in all known cellular cap methyltransferases. We isolated temperature-sensitive abd1 alleles and found that abd1-ts cells display a rapid shut-off of protein synthesis upon shift to the restrictive temperature, without wholesale reduction in steady-state mRNA levels. These in vivo results are consistent with classical biochemical studies showing a requirement for the cap methyl group in cap-dependent translation. We explored the issue of how cap methylation might be regulated in vivo by conducting a genetic screen for high-copy suppressors of the ts growth defect of abd1 mutants. The identification of the yeast genes SAM2 and SAM1, which encode AdoMet synthase, as abd1 suppressors suggests that Abd1p function can be modulated by changes in the concentration of its substrate AdoMet. We also identified the ubiquitin conjugating enzyme Cdc34p as a high-copy abd1 suppressor. We show that mutations of Cdc34p that affect its ubiquitin conjugation activity or its capacity to interact with the E3-SCF complex abrogate its abd1 suppressor function. Moreover, the growth defect of abd1 mutants is exacerbated by cdc34-2. These findings suggest a novel role for Cdc34p in gene expression and engender a model whereby cap methylation or cap utilization is negatively regulated by a factor that is degraded when Cdc34p is overexpressed.

Alanine↗
PubMed2000

Tetrandrine inhibits inward rectifying potassium current in cultured bovine aortic endothelial cells.

AIM: To study the effect of tetrandrine (Tet) on inward rectifying potassium current in cultured bovine aortic endothelial cells. METHODS: Inward rectifying potassium current (IRK) was observed by the whole cell patch-clamp technique. RESULTS: IRK was inhibited by Tet in a concentration-dependent manner and recovered to normal after wash with drug-free external solution. IRK was reduced from (582 +/- 48) pA to (221 +/- 40) pA at a holding potential of -70 mV by Tet 30 mumol/L. IC50 was 2.8 mumol/L. CONCLUSION: Tet inhibited inward rectifying potassium current in cultured bovine aortic endothelial cells.

Alkaloids↗
PubMed2000

[Studies on cell transformation and cell cycle in human embryo lungs induced by nickel compounds].

OBJECTIVE: To elucidate the association between nickel compounds and occupational lung cancer. METHODS: Human embryo lung cells (MRC-9 and IMR-90) were transformed by water-insoluble nickel oxide and nickel subsulfide, and changes in cell cycle of these transformed cells were analyzed by flow cytometric technique. RESULTS: Morphological transformation in MRC-9 and IMR-90 cells were induced after exposure to nickel compounds for 3 - 5 months, including formation of transforming foci, overlaying growth and epithelium-like changes in cells, etc. Some transformed cells could form colonies in soft agar media. Changes in cell cycle of the transformed cells were observed with flow cytometry. Proportions of cells at G(1) phase and G(2)/M phase were 62.61% and 16.37%, respectively, in the untreated MRC-9 cells and hyperploid cells were hardly seen in cell cycle plots. But, in the transformed MRC-9 cells, proportion of cells at G(1) phase reduced to about 50%, and that at G(2)/M phase increased to over 20% and that of hyperploid cells to over 10%. The same phenomenon was observed in the IMR-90 cells. CONCLUSIONS: Water-insoluble nickel compounds can induce morphological transformation of human embryo lung cells and those transformed cells appeared certain biological characteristics of tumor cells.

Cell Cycle↗
PubMed1999

Allelotype of uterine leiomyomas.

Uterine leiomyomas are the most common benign tumor that arise from smooth muscle cells of the myometrium. Little is known about the etiology and pathogenesis of this tumor. To investigate the molecular pathogenesis of these tumors, we have conducted an allelotype of 102 leiomyomas from 12 patients, using 67 fluorescently-tagged oligonucleotide primers amplifying microsatellite loci covering all autosomes. No areas of the genome showed frequent loss of heterozygosity (LOH); however, the highest rate of LOH (9%) was observed on 7q, consistent with previous cytogenetic observations. Uterine leiomyomas are sometimes multiple. In general, multiplicity of other types of neoplasm is associated with genetic predisposition to the disease. Because multiple tumors were available from each of the 12 patients studied, we looked for evidence of allele-specific LOH, which might indicate the presence of an underlying predisposition gene. However, no evidence for allele-specific LOH was detected, indicating that if cases of multiple uterine leiomyoma are due to an underlying predisposition gene, it is unlikely to be a recessive oncogene.

Alleles↗
PubMed1999

[Detection of cytomegalovirus antigens and virus DNA in the peripheral blood after organ transplantation].

OBJECTIVE: To diagnose timely the cytomegalovirus(CMV) infection of recipients after organ transplantation. METHODS: Thirty two organ transplanted recipients were demonstrated for CMV infection based on detection of antigenemia by immunoperoxidase assay, viremia with virus isolation, DNAemia by polymerase chain reaction (PCR) and in situ hybridization (ISH) in peripheral blood samples. RESULTS: Of 89 blood specimens examined, 35(39.3%) were positive for antigenemia, 25(28.1%) for viremia, 37(41.6%) for DNAemia by ISH, and 51(57.3%) for DNAemia by PCR. The results indicated that DNAemia assay with ISH and PCR and antigenemia assay were suitable methods for rapid, early, and sensitive detection of CMV infection. The results correlated with clinical symptoms. CONCLUSION: We suggest that detection of antigenemia, viremia, and DNAemia can monitor CMV infection of organ transplaned recipients and provide indication to antiviral therapy.

Adult↗
PubMed1999

Molecular cloning and sequencing of two 'short chain' and two 'long chain' K(+) channel-blocking peptides from the Chinese scorpion Buthus martensii Karsch.

Five full-length cDNAs encoding the precursors of two 'short chain' scorpion non-toxic peptides active on Ca(2+)-activated K(+) channels (BmP02 and BmP03) and two novel putative long chain K(+) channel-blocking peptides (named BmTXKbeta and BmTXKbeta2) were first isolated from the venom gland cDNA library of the Chinese scorpion Buthus martensii Karsch (BmK). BmTXKbeta2 showed a high similarity with AaTXKbeta, while BmTXKbeta was completely different in the deduced primary structure from the long chain and short chain scorpion toxins already characterized. Thus, BmTXKbeta expands the scorpion long chain K(+) channel-blocking peptide family. Although little sequence similarity exists between the above two short and two long peptides, they are similar at the positions of six cysteines, suggesting that they should all share a similar scaffold composed of an alpha-helix and a three-stranded beta-sheet.

3' Untranslated Regions↗
PubMed1999

Discovery of a small molecule insulin mimetic with antidiabetic activity in mice.

Insulin elicits a spectrum of biological responses by binding to its cell surface receptor. In a screen for small molecules that activate the human insulin receptor tyrosine kinase, a nonpeptidyl fungal metabolite (L-783,281) was identified that acted as an insulin mimetic in several biochemical and cellular assays. The compound was selective for insulin receptor versus insulin-like growth factor I (IGFI) receptor and other receptor tyrosine kinases. Oral administration of L-783,281 to two mouse models of diabetes resulted in significant lowering in blood glucose levels. These results demonstrate the feasibility of discovering novel insulin receptor activators that may lead to new therapies for diabetes.

Adenosine Triphosphate↗
PubMed1999

Improved reporter strain for monitoring Cre recombinase-mediated DNA excisions in mice.

Effective use of conditional Cre recombinase-loxP gene modification requires Cre-expressing mouse strains with defined patterns of expression. To assess the in vivo functionality of Cre-expressing mice, we have engineered an improved reporter strain for monitoring Cre-mediated excisions. The beta-galactosidase-neomycin phosphotransferase fusion gene (betageo)-trapped ROSA26 locus was modified by gene targeting such that betageo is expressed only after Cre-mediated excision of loxP-flanked DNA sequences. betageo from the excised ROSA26 allele is expressed ubiquitously in embryos and adult mice. By mating the reporter strain with Cre-expressing transgenic mice, we have shown that the loxP-flanked ROSA26 allele is accessible to Cre during early embryogenesis, as well as in a specific hematopoietic lineage (T lymphocytes). This improved reporter strain should facilitate monitoring in vivo Cre-mediated excision events in a variety of experimental contexts.

Animals↗
PubMed1999

Characterization of a neuronal kappaB-binding factor distinct from NF-kappaB.

Transcription factors that bind kappaB enhancer elements have begun to garner wide attention in neurobiology. Data suggest that activation of kappaB-binding factors in neurons can be protective against various neurotoxins, but other data have connected NF-kappaB to cell death. In electrophoretic mobility shift assays of kappaB-binding activity, we have found that the predominant activity in rat brain tissue, in primary neurons, and in neuronal cell lines has a mobility inconsistent with that of bona fide NF-kappaB (RelA-p50 heterodimer). We have tentatively termed this activity neuronal kappaB-binding factor (NKBF). Competition assays with various DNA probes distinguished NKBF from NF-kappaB. Probes that efficiently bind the p50 homodimer were able to compete with a conventional NF-kappaB probe for NKBF binding, but NKBF did not react with antibodies to p50 (or any other known Rel family members). Furthermore, UV-crosslinking indicated that NKBF is composed of two polypeptides of 82 kDa and 27 kDa. Although NKBF activity can be elevated in a manner independent of new macromolecular synthesis, it does not appear to be modulated by IkappaB. Finally, no NF-kappaB was induced by glutamate in highly enriched neuronal cultures, although it was induced in neuron-glia cocultures. These data suggest that the primary kappaB-binding transcription factor in neurons is a novel protein complex distinct from NF-kappaB.

Animals↗
PubMed1999

Vascular endothelial growth factor receptor KDR tyrosine kinase activity is increased by autophosphorylation of two activation loop tyrosine residues.

Vascular endothelial growth factor is an important physiological regulator of angiogenesis. The function of this endothelial cell selective growth factor is mediated by two homologous tyrosine kinase receptors, fms-like tyrosine kinase 1 (Flt-1) and kinase domain receptor (KDR). Although the functional consequence of vascular endothelial growth factor binding to the Flt-1 receptor is not fully understood, it is well established that mitogenic signaling is mediated by KDR. Upon sequencing several independent cDNA clones spanning the cytoplasmic region of human KDR, we identified and confirmed the identity of a functionally required valine at position 848 in the ATP binding site, rather than the previously reported glutamic acid residue, which corresponds to an inactive tyrosine kinase. The cytoplasmic domain of recombinant native KDR, expressed as a glutathione S-transferase fusion protein, can undergo autophosphorylation in the presence of ATP. In addition, the kinase activity can be substantially increased by autophosphorylation at physiologic ATP concentrations. Mutation analysis indicates that both tyrosine residues 1054 and 1059 are required for activation, which is a consequence of an increased affinity for both ATP and the peptide substrate and has no effect on kcat, the intrinsic catalytic activity of the enzyme. KDR kinase catalyzes phosphotransfer by formation of a ternary complex with ATP and the peptide substrate. We demonstrate that tyrosine kinase antagonists can preferentially inhibit either the unactivated or activated form of the enzyme.

DNA, Complementary↗
PubMed1999

Genetic aberrations in glioblastoma multiforme: translocation of chromosome 10 in an O-2A-like cell line.

We have examined the genetic aberrations in two near-diploid glioblastoma multiforme cell lines that appear to have arisen from different glial lineages. One cell line, Hu-O-2A/Gb1, expresses antigens and metabolic profiles characteristic of the oligodendrocyte-type-2 astrocyte (0-2A) lineage of the rat central nervous system. This line generates, in vitro, cells with characteristics of 0-2A progenitor cells, oligodendrocytes and astrocytes. The second cell line, IN1434, is derived from an astrocyte or a precursor cell restricted to astrocytic differentiation. In Hu-O-2A/Gb1 the sole homologue of chromosome 10 is disrupted at band 10p11-12.1 by translocation with chromosomes X and 15. The translocation breakpoint is localized between genetic markers D10S2103 and [D10S637, D10S1962, D10S355]. Other aberrations include a 5;14 translocation, deletion of the long and short arms of chromosome 16 and loss of one copy of the CDKN2 gene. IN1434 cells share some cytogenetic abnormalities with Hu-O-2A/Gb1 cells, despite their apparent derivation from a different biological origin, but also have translocations involving the long and short arms of chromosome 1 and the long arm of chromosome 7, and deletion of chromosome 13 at bands 13q12-21.

Animals↗
PubMed1999

In vivo post-translational processing and subunit reconstitution of cephalosporin acylase from Pseudomonas sp. 130.

Cephalosporin acylases are a group of enzymes that hydrolyze cephalosporin C (CPC) and/or glutaryl 7-amino cephalosporanic acid (GL-7ACA) to produce 7-amino cephalosporanic acid (7-ACA). The acylase from Pseudomonas sp. 130 (CA-130) is highly active on GL-7ACA and glutaryl 7-aminodesacetoxycephalosporanic acid (GL-7ADCA), but much less active on CPC and penicillin G. The gene encoding the enzyme is expressed as a precursor polypeptide consisting of a signal peptide followed by alpha- and beta-subunits, which are separated by a spacer peptide. Removing the signal peptide has little effect on precursor processing or enzyme activity. Substitution of the first residue of the beta-subunit, Ser, results in a complete loss of enzyme activity, and substitution of the last residue of the spacer, Gly, leads to an inactive and unprocessed precursor. The precursor is supposed to be processed autocatalytically, probably intramolecularly. The two subunits of the acylase, which separately are inactive, can generate enzyme activity when coexpressed in Escherichia coli. Data on this and other related acylases indicate that the cephalosporin acylases may belong to a novel class of enzymes (N-terminal nucleophile hydrolases) described recently.

Amino Acid Sequence↗
PubMed1999

Induction of IgE responses using a recombinant mosquito salivary allergen rAed a 2 without adjuvant in mice.

BACKGROUND: Reactions to mosquito bites are a global problem. Several salivary proteins from Aedes (Ae.) aegypti, the most common mosquito species, have been cloned and expressed. Plasmid DNA vaccination has been shown to be effective in the downregulation of IgE responses. To investigate the in vivo antigenicity of these recombinant proteins and to study the mechanisms underlying plasmid DNA vaccination, a mouse model sensitized with a recombinant antigen has been developed. METHODS: BALB/c and C57BL/6 mice were injected intradermally with a 37-kD recombinant Ae. aegypti salivary allergen (rAed a 2) in the absence of adjuvant twice weekly for 8 weeks and then challenged twice with rAed a 2 at weeks 10 and 12. Serum rAed a 2-specific IgE, IgG1 and IgG2a were measured by ELISA. Intradermal tests were performed every 4 weeks. The binding capacity of rAed a 2-specific IgE to the native Aed a 2 was examined by immunoblotting. RESULTS: In both strains, sensitization with rAed a 2 induced a significant increase in IgE and IgG1, but not IgG2a. In all sensitized mice, a positive immediate skin reaction was apparent, while delayed-type hypersensitivity reactions were not observed. BALB/c mice produced significantly higher levels of IgE and IgG1 and larger wheals than C57BL/6 mice. The IgE antibodies elicited by rAed a 2 bound to not only rAed a 2 but also its natural form in mosquito saliva. CONCLUSION: (1) Repeated injections of rAed a 2 without adjuvant induce predominant Th2-type responses in mice. (2) BALB/c mice are better responders for IgE production than C57BL/6 mice. (3) rAed a 2 has identical allergenicity to its natural form.

Adjuvants, Immunologic↗
PubMed1999

Inhibition of the activity of a neuronal kappaB-binding factor by glutamate.

Activation of transcription factors with affinity for kappaB enhancers is generally correlated with enhanced survival of neurons. In an apparent exception, excitotoxic concentrations of glutamate have been reported to elevate the activity of one such factor, nuclear factor-kappaB (NF-kappaB). Our data indicate that the constitutive neuronal kappaB-binding factor (NKBF) is distinct from bona fide NF-kappaB (RelA/p50 heterodimer). Therefore, we analyzed glutamate's effects on KB-binding activity in highly enriched primary neuronal cultures and in mixed neuron/glia cocultures. Electrophoretic mobility shift assays indicated that a 30-60-min exposure to 50-500 microM glutamate reduced NKBF activity by as much as 70%. Subtoxic doses of glutamate had little or no effect on this DNA-binding activity. Selective antagonists of either NMDA or AMPA [(RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate]/kai nat e receptors inhibited the influence of glutamate on NKBF activity. The effect of glutamate was mimicked by calcium ionophore, and it was blocked by lowering extracellular calcium concentrations or by cyclosporin A. Bona fide NF-kappaB was found only in cocultures containing significant numbers of glia, where it could be activated by glutamate. These data suggest that the primary influence of excitatory amino acids on neuronal KB-binding activity is an inhibitory one, strengthening the correlation between this transcriptional parameter and neuronal survival.

Animals↗