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Biomedical subjects

X Zhou

Publications and source records attributed to X Zhou.

At least 631 records · Page 35Linked to original sources
PubMed1995

A subunit interaction in chloroplast ATP synthase determined by genetic complementation between chloroplast and bacterial ATP synthase genes.

F1F0-ATP synthases utilize protein conformational changes induced by a transmembrane proton gradient to synthesize ATP. The allosteric cooperativity of these multisubunit enzymes presumably requires numerous protein-protein interactions within the enzyme complex. To correlate known in vitro changes in subunit structure with in vivo allosteric interactions, we introduced the beta subunit of spinach chloroplast coupling factor 1 ATP into a bacterial F1 ATP synthase. A cloned atpB gene, encoding the complete chloroplast beta subunit, complemented a chromosomal deletion of the cognate uncD gene in Escherichia coli and was incorporated into a functional hybrid F1 ATP synthase. The cysteine residue at position 63 in chloroplast beta is known to be located at the interface between alpha and beta subunits and to be conformationally coupled, in vitro, to the nucleotide binding site > 40 A away. Enlarging the side chain of chloroplast coupling factor 1 beta residue 63 from Cys to Trp blocked ATP synthesis in vivo without significantly impairing ATPase activity or ADP binding in vitro. The in vivo coupling of nucleotide binding at catalytic sites to transmembrane proton movement may thus involve an interaction, via conformational changes, between the amino-terminal domains of the alpha and beta subunits.

Adenosine Triphosphate↗
PubMed1995

Disruption of the architectural factor HMGI-C: DNA-binding AT hook motifs fused in lipomas to distinct transcriptional regulatory domains.

Lipomas are one of the most common mesenchymal neoplasms in humans. They are characterized by consistent cytogenetic aberrations involving chromosome 12 in bands q14-15. Interestingly, this region is also the site of rearrangement for other mesenchymally derived tumors. This study demonstrates that HMGI-C, an architectural factor that functions in transcriptional regulation, has been disrupted by rearrangement at the 12q14-15 chromosomal breakpoint in lipomas. Chimeric transcripts were isolated from two lipomas in which HMGI-C DNA-binding domains (AT hook motifs) are fused to either a LIM or an acidic transactivation domain. These results, identifying a gene rearranged in a benign neoplastic process that does not proceed to a malignancy, suggest a role for HMGI-C in adipogenesis and mesenchyme differentiation.

Amino Acid Sequence↗
PubMed1995

Heat-inactivated Sendai virus can enter multiple MHC class I processing pathways and generate cytotoxic T lymphocyte responses in vivo.

We have earlier described an alternative MHC class I processing pathway for Sendai virus (SV) in H-2Kb-transfected T2 cells (T2Kb). These cells have deleted genes for transporters associated with Ag processing (TAP1/2) and proteasome subunits LMP2/7 but can still process SV for the presentation of an immunodominant nucleoprotein CTL epitope (nucleoprotein peptide 324-332, FAPGNYPAL, SV9), even in the presence of the fungal metabolite brefeldin A (BFA). Presently we have compared live and heat-inactivated SV to investigate whether infectious virus, including early events such as binding and fusion at the host cell membrane, is important for nonclassical MHC class I processing and immunogenicity. We have found that heated virus (56 degrees C, boiled or autoclaved) with no fusion and hemagglutinin-neuraminidase activities, behaves similar to live SV in T2kb cells by entering a TAP-independent and BFA-resistant pathway. In EL-4 cells, which do not express this nonclassical TAP-independent and BFA-resistant pathway, heat-treated SV is processed in a BFA-sensitive way. In T1Kb- and TAP1/2-transfected T2Kb cells, as in T2Kb cells, processing of heat-inactivated SV was completely BFA resistant. Heat-inactivated SV was also found to prime CTLs in vivo. We conclude that heat-inactivated SV can enter both BFA-sensitive and -resistant MHC class I processing pathways and that SV in this respect may be particularly efficient. What property in the SV that is important for this characteristic is presently not clear but might be useful for the deliberate generation of CTL responses in vivo.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1995

Generation of cytotoxic and humoral immune responses by nonreplicative recombinant Semliki Forest virus.

The Semliki Forest virus (SFV) expression system can be used to package recombinant RNA into infectious suicide particles. Such RNA encodes only the SFV replicase and the heterologous protein but no structural proteins of SFV, and it is thus deficient in productive replication. We demonstrate here that infection of C57BL/6 (H-2b) and BALB/c (H-2d) mice with recombinant SFV expressing the nucleoprotein (NP) of influenza virus (SFV-NP) resulted in efficient priming of influenza virus-specific CD8+ cytotoxic T-cell (CTL) responses. The generated CTLs lysed both homologous (A/PR/8/34) and heterologous (A/HK/68) influenza virus-infected, or peptide-coated, target cells to a similar degree as CTLs induced by wild-type influenza virus in a major histocompatibility complex class I-restricted fashion. As few as 100 infectious units of virus induced a strong CTL response. Induction of CTL by SFV-NP could also be achieved in CD4 gene-targeted mice, demonstrating the independence of the primary CTL response of CD4+ helper T cells. One immunization generated a CTL response that peaked after 1 week, and an additional booster injection generated a CTL memory, which was still detectable after 40 days. SFV-NP immunizations also generated high-titered IgG humoral responses that remained significant after several months. These results demonstrate that the recombinant SFV suicide system is highly efficient in antigen presentation and suggest that it may have a potential as a recombinant vaccine.

Amino Acid Sequence↗
PubMed1995

Glycosylphosphatidylinositol-linked Db does not induce an influenza-specific cytotoxic T lymphocyte response or recycle membrane-bound peptides.

Major histocompatibility complex (MHC) class I molecules, as well as MHC class I-bound peptides, are known to recycle between the cell surface and an undefined, endosomal-like compartment. Little is known about the functional significance of this process. We have explored this using two different forms of the H-2Db molecule expressed in transgenic mice, either transmembranous (Db-tm) or with a glycophosphatidylinositol (GPI)-lipid anchor (Db-GPI). The recycling capacity of peptides bound to Db-tm and Db-GPI was investigated using glycosylated Db-binding glycopeptides, which were detected by flow cytometry. Only the tm form of Db was found to readily internalize and recycle glycopeptides to the cell surface. When transgenic mice were immunized with influenza A virus (PR8) strain and tested for cytotoxic T lymphocyte (CTL) responses against an immunedominant nucleoprotein epitope (366-374, ASNENMETM), only Db-tm mice were found to generate specific CTL responses. The results support the idea that membrane recycling of MHC class I-bound peptides on antigen-presenting cells may be important for the generation of certain CTL responses.

Animals↗
PubMed1995

Genomic DNA and messenger RNA expression alterations of the CDKN2B and CDKN2 genes in esophageal squamous carcinoma cell lines.

The genes CDKN2B (MTS2) and CDKN2 (MTS1) encoding the proteins p15 and p16 are both located on chromosomal band 9p21, a locus at which frequent homozygous and heterozygous deletions occur in many primary human tumors, including esophageal carcinoma. CDKN2 and CDKN2B belong to a family of cyclin-dependent kinase 4 inhibitors (INK41) and control cell proliferation during the G1 phase of the cell cycle. Their inactivation may contribute to uncontrolled growth in human cancers. To investigate whether CDKN2B and CDKN2 are involved in esophageal tumorigenesis, we studied homozygous deletion, intragenic mutation, and messenger RNA (mRNA) expression of CDKN2 and CDKN2B in nine esophageal squamous cancer cell lines. Polymerase chain reaction (PCR) amplification revealed that five of the nine cell lines (55%) manifested homozygous deletions of CDKN2B, CDKN2, and/or flanking loci on chromosomal band 9p21. Reverse transcriptase-PCR (RT-PCR) was used to examine CDKN2 and CDKN2B mRNA in the nine cell lines. Lack of CDKN2 and CDKN2B mRNA correlated perfectly with homozygous deletion involving these genes. No subtle intragenic mutations of CDKN2B or CDKN2 were detected by DNA sequencing of their entire coding sequences in any cell lines lacking homozygous deletion. Two of the cell lines manifested homozygous deletions excluding CDKN2; one of these two deletions also excluded CDKN2B. These results suggest that inactivation of CDKN2B and CDKN2 may contribute to the malignant phenotype in esophageal cells and that homozygous deletion may be the predominant mechanism for inactivation of CDKN2B and CDKN2. Alternatively, a gene or genes adjacent to CDKN2B/CDKN2 may constitute the target(s) of deletion at this locus.

Alleles↗
PubMed1995

Mechanisms of beta 1 integrin-dependent adherence of granulocytic HL60 to fibronectin.

We investigated the mechanism of beta 1 integrin-mediated adherence of stimulated granulocytic HL60 cells to fibronectin using a monoclonal antibody (15/7) that recognizes beta 1 integrins only when the receptors are active for ligand binding. Phorbol myristate acetate (PMA) stimulated expression of the 15/7 epitope on granulocytic HL60 by nearly fivefold but had an insignificant effect on the expression of the epitope on undifferentiated HL60 cells. These results paralleled the effect of PMA on HL60 and granulocytic HL60 adhesion to fibronectin, indicating that activation of beta 1 integrins is important for beta 1-mediated adherence of granulocytic HL60 cells to fibronectin. Agonists that stimulate alpha 5 beta 1-dependent human polymorphonuclear leukocyte (PMN) adhesion to fibronectin (C5a and PMA) also upregulated the 15/7 epitope on purified human PMNs. Although PMA rapidly induces increased levels of filamentous actin (F-actin) in granulocytic HL60 cells and a decrease in F-actin levels in undifferentiated HL60 cells, depolymerization of the actin cytoskeleton with cytochalasin B did not affect increased expression of the 15/7 epitope on granulocytic HL60 cells. Cytochalasin B did, however, inhibit granulocytic HL60 adherence to fibronectin by 50%, demonstrating that actin polymerization is important for optimal beta 1-dependent granulocytic adherence.

Actins↗
PubMed1995

Detection of bromobenzene-induced hepatocellular necrosis using magnetic resonance microscopy.

The authors used magnetic resonance (MR) microscopy to assess hepatic tissue damage induced by bromobenzene both in living rats and in fixed rat liver tissues. Experiments were conducted at 7 Tesla on three groups of Fisher rats treated with bromobenzene at a single dose of 68, 135, and 269 mg/kg, respectively. Optical microscopy of hematoxylin and eosin stained sections showed liver damage only at the highest dose, whereas with MR microscopy, tissue alterations were detected at all three doses both in vivo and ex vivo. The contrast mechanism of the superior sensitivity of MR microscopy is believed to be related to the changes in local diffusion coefficients that accompany cellular degeneration and death, although other contrast mechanisms may also be involved. The superior sensitivity of MR microscopy, as demonstrated in this study, has many implications for potential use of MR techniques to perform in vivo histology.

Animals↗
PubMed1995

Interaction of singlet oxygen with 5'-nucleotidase in rat hearts.

This study was aimed to determine whether singlet oxygen (1O2) attenuates 5'-nucleotidase activity in the ischemic myocardium. Isolated rat hearts were exposed to either exogenous 1O2 produced by irradiating rose bengal or 40-min ischemia and reperfusion. Ecto-5'-nucleotidase activity was inhibited by exogenous 1O2 (3.74 +/- 0.38 mumol/min/g dry weight), when compared with normal control (7.52 +/- 0.41 mumol/min/g dry weight; P < 0.05). The enzymatic activity was significantly preserved by histidine (25 mM)--a 1O2 scavenger (7.04 +/- 0.61 mumol/min/g dry weight; P < 0.05 v rose bengal group). After ischemia, the activity of ecto-5'-nucleotidase was greatly reduced (2.51 +/- 0.25 mumol/min/g dry weight), when compared with normal control. Histidine significantly enhanced ecto-5'-nucleotidase activity (6.55 +/- 0.52 mumol/min/g dry weight, P < 0.05 v ischemic control). Adenosine release was consistent with ecto-5'-nucleotidase activity. The time course studies of effects of 1O2 on coronary flow, cardiac function, and LDH release revealed that the damage by 1O2 to ecto-5'-nucleotidase activity and adenosine release primarily accounted for impaired coronary flow, cardiac dysfunction, and impaired cardiac metabolism. Lipid peroxidation induced by exogenous 1O2 or ischemia was in parallel with ecto-5'-nucleotidase deactivation by 1O2. It is concluded that 1O2 causes inactivation of ecto-5'-nucleotidase and attenuation of adenosine release which could possibly be one of the important mechanisms of oxygen radical-mediated myocardial injury.

5'-Nucleotidase↗
PubMed1995

An impedimetric method for rapid screening of cosmetic preservatives.

An efficient impedance method was developed for rapid evaluation of cosmetic preservatives. The method used decimal reduction time or D-value to assess preservative efficacies. The D-value, which was calculated from the plot of Log CFU ml-1 versus time by linear regression analysis, could be obtained within 48 h. Thus, the time required for the challenge test was reduced from 4-8 weeks with the standard procedures (eg US Pharmacopeia), to 2 days with the current method. A calibration curve (r = -0.95) was established by plotting the Log CFU ml-1 versus capacitance detection time (DT) of 108 samples. With the calibration, CFU can be estimated directly from the impedance test without plating. Two commercial biocides and several other chemicals were evaluated in a shampoo by the impedance procedure against Pseudomonas aeruginosa. The D-values obtained from the impedance test were not significantly different from those produced by the conventional plate count method. The technique was found to be particularly useful when screening a large number of compounds to find novel preservatives and synergistic preservative combinations.

Cosmetics↗
PubMed1995

Changes in the serum concentrations of gonadotrophins and oestradiol in peripubertal Chinese Beijing black gilts.

Four gifts of the Chinese Beijing Black breed, all from the same litter, had their blood serum concentrations of luteinizing hormone (LH), follicle stimulating hormone (FSH) and oestrogens (E2) measured by radioimmunoassay at intervals of 20 days between the ages of 5 and 185 days. The concentrations of LH declined sharply between days 5 to 25 and remained low until rising again from day 85 and reaching a peak at day 105 before declining again and remaining constant. The concentrations of FSH increased from day 85, peaking at day 105, followed by a slow decline to day 185. Serum E2 concentrations were high at day 5 then decreased dramatically, but increased considerably at 185 days. These results are similar to those reported from European breeds.

Animals↗
PubMed1995

Glutathione adducts, not carbamylated lysines, are the major modification of lens alpha-crystallins from renal failure patients.

alpha-Crystallins from the water-soluble and the water-insoluble, guanidine-soluble portions of lenses from four renal failure patients and two normal donors of similar age were isolated and enzymatically digested into peptides. Molecular weights of the peptides, determined by fast atom bombardment mass spectrometry, indicated modifications specifically associated with renal failure. The only modifications observed in the alpha-crystallins from renal failure patients, but not in the normal old lenses, were glutathione adducts to Cys 131 and Cys 142. These adducts were present in the lenses of all four renal failure patients, but not in the two normal old lenses. The four lenses from the renal failure patients were searched for evidence of carbamylation at lysyl or cysteinyl residues: carbamylation was not detected. Because the same mass spectrometric methods had previously demonstrated sufficient sensitivity and specificity to detect as little as 5% modification in the examination of in vitro carbamylated bovine lenses, these results indicated that carbamylation is not a major modification of the lens alpha-crystallins of renal failure patients.

Aged↗
PubMed1995

A system for production of commercial quantities of human lactoferrin: a broad spectrum natural antibiotic.

We previously reported the production of limited quantities of biologically active recombinant human lactoferrin in the filamentous fungus Aspergillus oryzae. In the present study, we report a modification of this production system combined with a classical strain improvement program that has enabled production of levels of recombinant human lactoferrin in excess of 2 g/l. The protein was expressed in Aspergillus awamori as a glucoamylase fusion polypeptide which was secreted into the growth medium and processed to mature human lactoferrin by an endogenous KEX-2 peptidase. The recombinant protein retains full biological activity in terms of its ability to bind iron and human enterocyte receptors. Furthermore, the recombinant protein functions as a potent broad spectrum antimicrobial protein.

Animals↗
PubMed1995

Intragenic mutations of CDKN2B and CDKN2A in primary human esophageal cancers.

The CDKN2A and CDKN2B genes, encoding p16 and p15 respectively, are located on chromosome 9p21, a locus at which frequent homozygous and heterozygous deletions occur in many primary human tumors, including esophageal carcinoma. CDKN2A and CDKN2B inhibit cyclin dependent kinase 4 (CDK4) and CDK6 and control cellular proliferation by preventing entry into the S phase of the cell cycle. Their inactivation may contribute to uncontrolled growth in human cancer. We previously described CDKN2A exon 2 mutations in a pilot study of 43 esophageal cancers. In order to determine whether CDKN2A and CDKN2B are frequent targets of 9p21 deletion in esophageal carcinogenesis, we have now analyzed 60 primary esophageal cancers for mutations in both exons 1 and 2 of CDKN2A and CDKN2B by direct sequencing of PCR amplified genomic DNAs. In conjunction with our previously published data, we have identified a total of eight nucleic acid substitutions among 60 esophageal carcinomas; here, we describe one new CDKN2B nonsense mutation and one new silent CDKN2B mutation that occurred somatically. Taken together, these results suggest that intragenic mutations in CDKN2A and CDKN2B occur in esophageal cancer, but that they are infrequent events. In view of the known high frequency of loss of heterozygosity at the chromosome 9p21 locus in esophageal cancers, the current data suggest that intragenic mutation is not the predominant mode of inactivation of CDKN2A and CDKN2B or that other genes are targets of deletion at this locus in these cancers.

Base Sequence↗
PubMed1995

Cell-specific localization of apolipoprotein E messenger ribonucleic acid in the testis and epididymis of the rat.

Apolipoprotein (apo) E, a 35-kDa protein found on the surface of several lipoproteins, has been detected in many peripheral tissues and is postulated to function in facilitating the transfer of cholesterol/lipids between cells. We examined the expression of apo E mRNA in the testes and epididymides of juvenile rats (21 days old), prepubescent rats (34-36 days old), and sexually mature rats (75-80 days old). In situ hybridization using 35S-labeled rat apo E riboprobes was used to identify cells containing apo E mRNA. Such cells were located in the interstitial area of testes obtained from rats of all ages. This cell population consisted of primarily Leydig cells with occasional macrophages, according to immunoreactivity to 3 beta-hydroxysteroid dehydrogenase and antimacrophage antibodies, respectively. Caput epididymides obtained from sexually mature and prepubescent rats contained apo E mRNA-positive cells located in the basal region of the epididymal tubules and within the interstitial stroma. Our data are consistent with the concept that locally produced apo E plays a role in the physiologic function of the rat testis and epididymis.

Aging↗