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Biomedical subjects

X Zou

Publications and source records attributed to X Zou.

At least 19 recordsLinked to original sources
PubMed2004

Involvement of peripheral neuropeptide Y receptors in sympathetic modulation of acute cutaneous flare induced by intradermal capsaicin.

In a recent study, we have demonstrated that the dorsal root reflex (DRR)-mediated acute cutaneous neurogenic inflammation following intradermal injection of capsaicin (CAP) is sympathetically dependent and subject to modulation by peripheral alpha(1)-adrenoceptors. Postganglionic sympathetic neurons contain not only adrenergic neurotransmitters, but also non-adrenergic substances, including neuropeptide Y (NPY). In this study, we examined if peripheral NPY receptors participate in the flare following CAP injection. Different NPY receptor subtypes were studied by using relatively specific agonists and antagonists for the Y(1) and Y(2) subtypes. Changes in cutaneous blood flow on the plantar surface of the foot were measured using a laser Doppler flowmeter. Following CAP injection, cutaneous flare spread more than 20 mm away from the site of CAP injection. Removal of the postganglionic sympathetic nerves by surgical sympathectomy reduced dramatically the CAP-evoked flare. If the foot of sympathectomized rats was pretreated with either NPY or Y(2) receptor agonists by intra-arterial injection, the spread of flare induced by CAP injection could be restored and prolonged. However, if the spinal cord was pretreated with a GABA(A) receptor antagonist, bicuculline, to prevent DRRs, NPY or an Y(2) receptor agonist no longer restored the CAP-evoked flare. A Y(1) receptor agonist did not affect the CAP-evoked flare in sympathectomized rats. In sympathetically intact rats, blockade of either peripheral NPY or Y(2) receptors with [D-Trp(32)]-NPY or BIIE0246 markedly reduced the flare induced by CAP injection, whereas blockade of peripheral Y(1) receptors by BIBP3226 did not obviously affect the flare. It is suggested that NPY is co-released with NE from the postganglionic sympathetic terminals to activate NPY Y(2) and alpha(1) receptors following CAP injection. Both substances are involved, at least in part, in modulation of the responses of CAP sensitive afferents thereby affecting their ability to evoke the release of inflammatory agents from primary afferents.

Animals↗
PubMed2002

Role of protein kinase A in phosphorylation of NMDA receptor 1 subunits in dorsal horn and spinothalamic tract neurons after intradermal injection of capsaicin in rats.

Protein phosphorylation is a major mechanism for regulation of N-methyl-D-aspartate (NMDA) receptor function. The NMDA receptor 1 subunit (NR1) is phosphorylated by protein kinase A (PKA) on serine 890 and 897. We have recently reported that there is enhanced phosphorylation of NR1 on serine 897 in dorsal horn and spinothalamic tract (STT) neurons after intradermal injection of capsaicin (CAP) in rats [Zou et al. (2000) J. Neurosci. 20, 6989-6997]. Whether or not this phosphorylation, which develops during central sensitization following CAP injection, is mediated by PKA remains to be determined. In this study, western blots and immunofluorescence staining were employed to observe if pretreatment with a PKA inhibitor, N-[2-((p-bromocinnamyl)amino)ethyl]-5-isoquinolinesulfonamide, HCl (H89), blocks the enhanced phosphorylation of NR1 on serine 897 following injection of CAP into the glabrous skin of one hind paw of anesthetized rats. Western blots showed that pretreatment with H89 caused a decrease in CAP-induced phosphorylation of NR1 protein in spinal cord segments L(4)-S(1). In experiments using immunofluorescence staining, the numbers of phospho-NR1-like immunoreactive (p-NR1-LI) neurons seen after CAP injection were significantly decreased in the dorsal horn of the L(4)-L(5) segments on the side ipsilateral to the injection after PKA was inhibited. When STT cells were labeled by microinjection of the retrograde tracer, fluorogold, we found that the proportion of p-NR1-LI STT cells on the side ipsilateral to the injection in the superficial laminae of spinal cord segments L(4)-L(5) was markedly reduced when H89 was administered intrathecally before CAP injection. However, the proportion of p-NR1-LI STT cells in deep laminae was unchanged unless the PKC inhibitor, chelerythrine chloride, was co-administered with H89. Combined with our previous findings, the present results indicate that NR1 in spinal dorsal horn neurons, including the superficial dorsal horn STT cells, is phosphorylated following CAP injection and that this phosphorylation is due to the action of PKA. However, the phosphorylation of deep STT cells involves both PKA and PKC.

Animals↗
PubMed2002

Production of antigen-specific human monoclonal antibodies: comparison of mice carrying IgH/kappa or IgH/kappa/lambda transloci.

Here we compare human monoclonal antibody (MAb) production from mouse strains that carry disruptions of their endogenous mouse IgH/IgK loci and harbor human IgM + Igkappa(BABkappa) or human IgM + Igkappa + IgA transloci (BABkappa,lambda). We found that whereas both strains proved effective for the isolation of antigen-specific IgM antibodies, many of the IgM MAbs elicited from BABkappa comprise human mu chains that are associated with mouse lambda chains. In contrast, BABkappa,lambda mice gave rise to fully functional, polymeric human IgM antibodies comprising both human IgH and human IgL chains. Therefore, the inclusion of a human Iglambda translocus (in addition to the human IgH + Igkappa transloci) not only diminishes problems of endogenous mouse Iglambda expression but also provides a strain of mice that yields fully human MAbs to a wide range of antigens, as witnessed by the isolation of MAbs to human blood cells, tumor cell lines, and an immunoglobulin idiotype.

Animals↗
PubMed2001

Enzymatic activity of coenzyme B(12) derivatives with altered axial nucleotides: probing the mechanochemical triggering hypothesis in ribonucleotide reductase.

Theoretical studies (J. Inorg. Biochem. 2001, 83, 121) of the involvement of the bulky 5,6-dimethylbenzimidazole (Dmbz) ligand of coenzyme B(12) (5'-deoxyadenosylcobalamin, AdoCbl) in the mechanism of activation of the carbon-cobalt bond of the coenzyme for homolytic cleavage by AdoCbl-dependent enzymes (the "mechanochemical triggering" mechanisms) have shown that a purely steric, ground-state mechanism can supply only a few kilocalories per mole (of the observed 13-16 kcal mol(-1)) of activation, but that an electronic mechanism, operating to stabilize the transition state, can explain all of the observed catalytic effect. To address these mechanisms experimentally, analogues of AdoCbl in which the Dmbz ligand is replaced by benzimidazole (Ado(Bzim)Cbl) or by imidazole (Ado(Im)Cbl) have been prepared and characterized. Both of these analogues support turnover in the AdoCbl-dependent ribonucleoside triphosphate reductase (RTPR) from Lactobacillus leichmannii at 100% of the activity of AdoCbl itself, but the Ado(Im)Cbl analogue has a significantly higher K(m). 5'-Deoxyadenosylcobinamide, the analogue in which the axial nucleotide has been chemically removed, in contrast, is inactive in the spectrophotometric assay, which indicates that it has at most 1% of the activity of AdoCbl. Stopped-flow spectrophotometric measurements of the formation of cob(II)alamin at the enzyme active site show that RTPR binds Ado(Bzim)Cbl slightly more weakly than it does AdoCbl, but binds Ado(Im)Cbl 8-fold more weakly. While the equilibrium constant for cob(II)alamin formation is nearly the same for Ado(Bzim)Cbl and AdoCbl, it is 5-fold smaller for Ado(Im)Cbl. Finally, the forward rate constant for enzyme-induced Co-C bond homolysis was about the same for Ado(Bzim)Cbl and for AdoCbl but was 17-fold smaller for Ado(Im)Cbl. These results are consistent with a small contribution from ground-state mechanochemical triggering, but they do not in themselves rule out transition-state mechanical triggering.

Adenine Nucleotides↗
PubMed2001

Equilibrium and kinetic studies on the reactions of alkylcobalamins with cyanide.

Ligand substitution equilibria of different alkylcobalamins (RCbl, R = Me, CH(2)Br, CH(2)CF(3), CHF(2), CF(3)) with cyanide have been studied. It was found that CN(-) first substitutes the 5,6-dimethylbenzimidazole (Bzm) moiety in the alpha-position, followed by substitution of the alkyl group in the beta-position trans to Bzm. The formation constants K(CN) for the 1:1 cyanide adducts (R(CN)Cbl) were found to be 0.38 +/- 0.03, 0.43 +/- 0.03, and 123 +/- 9 M(-1) for R = Me, CH(2)Br, and CF(3), respectively. In the case of R = CH(2)CF(3), the 1:1 adduct decomposes in the dark with CN(-) to give (CN)(2)Cbl. The unfavorable formation constants for R = Me and CH(2)Br indicate the requirement of very high cyanide concentrations to produce the 1:1 complex, which cause the kinetics of the displacement of Bzm to be too fast to follow kinetically. The kinetics of the displacement of Bzm by CN(-) could be followed for R = CH(2)CF(3) and CF(3) to form CF(3)CH(2)(CN)Cbl and CF(3)(CN)Cbl, respectively, in the rate-determining step. Both reactions show saturation kinetics at high cyanide concentration, and the limiting rate constants are characterized by the activation parameters: R = CH(2)CF(3), DeltaH = 71 +/- 1 kJ mol(-1), DeltaS = -25 +/- 4 J K(-1) mol(-1), and DeltaV = +8.9 +/- 1.0 cm(3) mol(-1); R = CF(3), DeltaH = 77 +/- 3 kJ mol(-1), DeltaS = +44 +/- 11 J K(-1) mol(-1), and DeltaV = +14.8 +/- 0.8 cm(3) mol(-1), respectively. These parameters are interpreted in terms of an I(d) and D mechanism for R = CH(2)CF(3) and CF(3), respectively. The results of the study enable the formulation of a general mechanism that can account for the substitution behavior of all investigated alkylcobalamins including coenzyme B(12).

Cyanides↗
PubMed2001

Novel control motif cluster in the IgH delta-gamma 3 interval exhibits B cell-specific enhancer function in early development.

The majority of the human Ig heavy chain (IgH) constant (C) region locus has been cloned and mapped. An exception is the region between C delta and C gamma 3, which is unstable and may be a recombination hot spot. We isolated a pBAC clone (pHuIgH3'delta-gamma 3) that established a 52-kb distance between C delta and C gamma 3. Sequence analysis identified a high number of repeat elements, explaining the instability of the region, and an unusually large accumulation of transcription factor-binding motifs, for both lymphocyte-specific and ubiquitous transcription activators (IKAROS, E47, Oct-1, USF, Myc/Max), and for factors that may repress transcription (Delta EF1, Gfi-1, E4BP4, C/EBP beta). Functional analysis in reporter gene assays revealed the importance of the C delta-C gamma 3 interval in lymphocyte differentiation and identified independent regions capable of either enhancement or silencing of reporter gene expression and interaction with the IgH intron enhancer E mu. In transgenic mice, carrying a construct that links the beta-globin reporter to the novel delta-gamma 3 intron enhancer (E delta-gamma 3), transgene transcription is exclusively found in bone marrow B cells from the early stage when IgH rearrangement is initiated up to the successful completion of H and L locus recombination, resulting in Ab expression. These findings suggest that the C delta-C gamma 3 interval exerts regulatory control on Ig gene activation and expression during early lymphoid development.

Amino Acid Motifs↗
PubMed2001

Design, docking, and evaluation of multiple libraries against multiple targets.

We present a general approach to the design, docking, and virtual screening of multiple combinatorial libraries against a family of proteins. The method consists of three main stages: docking the scaffold, selecting the best substituents at each site of diversity, and comparing the resultant molecules within and between the libraries. The core "divide-and-conquer" algorithm for side-chain selection, developed from an earlier version (Sun et al., J Comp Aided Mol Design 1998;12:597-604), provides a way to explore large lists of substituents with linear rather than combinatorial time dependence. We have applied our method to three combinatorial libraries and three serine proteases: trypsin, chymotrypsin, and elastase. We show that the scaffold docking procedure, in conjunction with a novel vector-based orientation filter, reproduces crystallographic binding modes. In addition, the free-energy-based scoring procedure (Zou et al., J Am Chem Soc 1999;121:8033-8043) is able to reproduce experimental binding data for P1 mutants of macromolecular protease inhibitors. Finally, we show that our method discriminates between a peptide library and virtual libraries built on benzodiazepine and tetrahydroisoquinolinone scaffolds. Implications of the docking results for library design are explored.

Amino Acids↗
PubMed2001

Helicobacter pylori seropositivity and subsite-specific gastric cancer risks in Linxian, China.

BACKGROUND: Helicobacter pylori carriage (i.e., persistent exposure to the organism without gastric epithelial cell invasion) is an established risk factor for noncardia gastric cancer. However, its association with the risk of cancer of the gastric cardia is controversial. Consequently, we designed this prospective, nested case-control study to further explore the subsite-specific gastric cancer risks associated with H. pylori seropositivity (a surrogate marker for persistent exposure). METHODS: A total of 99 patients with gastric cardia cancer, 82 patients with noncardia gastric cancer, and 192 cancer-free subjects were selected from among the participants (n = 29 584) of a nutrition intervention trial previously conducted in Linxian, China. H. pylori seropositivity was determined by assaying for the presence of H. pylori whole cell and CagA antibodies in baseline serum samples from all subjects. Seropositivity was defined as one or both serum assays being positive. Odds ratios (ORs) for subsite-specific gastric cancer were estimated by multivariate logistic regression analyses. All statistical comparisons were two-sided (alpha =.05). RESULTS: H. pylori seropositivity rates for subjects with gastric cardia cancer, noncardia gastric cancer, and gastric cardia and noncardia cancers combined were 70% (P =.02), 72% (P: =.01), and 71% (P =.003) compared with 56% for cancer-free control subjects. OR estimates for H. pylori seropositivity were 1.87 (95% confidence interval [CI] = 1.10 to 3.17) for gastric cardia cancer, 2.29 (95% CI = 1.26 to 4.14) for noncardia gastric cancer, and 2.04 (95% CI = 1.31 to 3.18) for gastric cardia and noncardia cancers combined. CONCLUSIONS: H. pylori seropositivity was associated with increased risks for both gastric cardia cancer and noncardia gastric cancer in this well-characterized cohort. Thus, H. pylori carriage may increase the risk of cancer throughout the stomach.

Adenocarcinoma↗
PubMed2001

Complete nucleotide sequence and characterization of pUA140, a cryptic plasmid from Streptococcus mutans.

Approximately 5% of strains of Streptococcus mutans contain plasmid DNA. Strain UA140 harbors a 5.6-kb cryptic plasmid, pUA140, with an overall G+C content of 32.7%. Five open reading frames (ORF), encoding peptides of larger than 100 amino acid residues, were initially designated as ORF1 to ORF5. These five ORFs were located on the same strand of pUA140. ORF1 (258 amino acids) resembled a replication protein, Rep. Upstream of the putative Rep gene, a double-stranded origin for plasmid replication that showed strong similarity to those of a number of plasmids in the pT181 family was identified. Further upstream was a region constituting the single-stranded origin of replication. A single-stranded DNA intermediate was detected during plasmid replication. Taken together, these results suggest that pUA140 replicated by the rolling circle replication mechanism but exhibited several characteristics that differ from those of other members of the pT181 plasmid family.

Amino Acid Motifs↗
PubMed2001

Structured population on two patches: modeling dispersal and delay.

We derive from the age-structured model a system of delay differential equations to describe the interaction of spatial dispersal (over two patches) and time delay (arising from the maturation period). Our model analysis shows that varying the immature death rate can alter the behavior of the homogeneous equilibria, leading to transient oscillations around an intermediate equilibrium and complicated dynamics (in the form of the coexistence of possibly stable synchronized periodic oscillations and unstable phase-locked oscillations) near the largest equilibrium.

Age Factors↗
PubMed2001

NMDA or non-NMDA receptor antagonists attenuate increased Fos expression in spinal dorsal horn GABAergic neurons after intradermal injection of capsaicin in rats.

GABAergic neurons play an important role in the generation of primary afferent depolarization, which results in presynaptic inhibition and, if large enough, triggers dorsal root reflexes. Recent electrophysiological studies by our group have suggested that increased excitation of spinal GABAergic neurons by activation of N-methyl-D-aspartate (NMDA) and non-NMDA receptors following intradermal injection of capsaicin results in the generation of DRRs that contribute to neurogenic inflammation. The present study was to determine if changes in the expression of Fos protein occur in GABAergic neurons in the lumbosacral spinal cord following injection of capsaicin into the glabrous skin of one hind paw of anesthetized rats and if pretreatment with an NMDA receptor antagonist, D-(-)-2-amino-7-phosphonoheptanoic acid (AP7) or a non-NMDA receptor antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) blocks Fos expression in these neurons. The experiments used western blots and immunofluorescence double labeling staining following capsaicin or vehicle injection. Western blots showed that Fos protein was increased on the ipsilateral side in spinal cord tissue 0.5 h after capsaicin injection. Pretreatment with AP7 or CNQX caused a decrease in capsaicin-induced Fos expression. Immunofluorescence double labeling showed that the proportion of Fos-positive GABAergic neuronal profiles was significantly increased following capsaicin injection (48.8+/-4.8%) compared to the vehicle injection (23.8+/-5.1%) in superficial laminae on the ipsilateral side in lumbosacral spinal cord (P<0.05). However, when the spinal cord was pretreated with AP7 (5 microg) or CNQX (0.2 microg), only 9.1+/-0.6% or 7.1+/-0.8% of GABA-immunoreactive neuronal profiles were stained for Fos following capsaicin injection. The blockade of the capsaicin-evoked Fos staining was dose-dependent. These findings suggest that GABAergic neurons take part in dorsal horn circuits that modulate nociceptive information and that the function of GABAergic neurons following capsaicin injection is partially mediated by NMDA and non-NMDA receptors.

2-Amino-5-phosphonovalerate↗
PubMed2001

Production of cloned goats from enucleated oocytes injected with cumulus cell nuclei or fused with cumulus cells.

This study was designed to produce cloned goats from cumulus cells. Cloning donor nuclei were from cumulus cells either freshly isolated or cultured in vitro. Enucleated oocytes were either injected with cumulus cell nuclei without piezo-driven manipulator (injection method) or fused with cumulus cells (fusion method). The survival rate of cloned embryos, obtained by injection, was higher than that derived from fusion (62.7 and 45.9%, respectively). Two cloned goats were derived by fusion with in vitro cultured cumulus cells without starvation, but died shortly after natural birth, from respiratory difficulties. Their birth weights (2.23 kg and 2.03 kg) were within the normal range (2.0-2.7 kg) and postmortem analysis revealed no morphological abnormalities. The third cloned goat, derived by injection of nuclei from freshly isolated cumulus cells, weighed 3.3 kg at birth, and was 37% overweight compared with the average weight of the same species. This goat is healthy and well as this paper is being prepared. Nested PCR-RFLP analysis confirmed that all the cloned goats were derived from the donor cells.

Animals↗
PubMed2001

Truncation of the mu heavy chain alters BCR signalling and allows recruitment of CD5+ B cells.

Ig are multifunctional molecules with distinct properties assigned to individual domains. To assess the importance of IgM domain assembly in B cell development we generated two transgenic mouse lines with truncated muH chains by homologous integration of the neomycin resistance gene (neo(r)) into exons C(mu)1 and C(mu)2. Upon DNA rearrangement shortened muH chain transcripts, V(H)-D-J(H)-C(mu)3-C(mu)4, are produced independent of the transcriptional orientation and termination signals provided by neo(r). The truncated muH chain of approximately 52 kDa associates non-covalently with the L chain to form a monovalent HL heterodimer. Surface IgM is assembled into a defective BCR complex which has lost important signalling capacity. In immunizations with T-dependent and T-independent antigens, specific IgM antibodies cannot be detected, whilst IgG responses remain normal. B cell development in the bone marrow is characterized by an increase in early B cells, but a decrease of B220(+) cells from the stage when muH chain rearrangement is completed. The peritoneal lymphocyte population has elevated levels of CD5(+) B cells and their expansion may be the result of a negative feedback mechanism. The results show that antigenic stimulation is compromised by truncated monovalent IgM and that this deficit in stimulation leads to reduced levels of conventional B-2 lymphocytes, but dramatically increased levels of B-1 cells.

Alternative Splicing↗
PubMed2001

The cell cycle-regulatory CDC25A phosphatase inhibits apoptosis signal-regulating kinase 1.

CDC25A phosphatase promotes cell cycle progression by activating G(1) cyclin-dependent kinases and has been postulated to be an oncogene because of its ability to cooperate with RAS to transform rodent fibroblasts. In this study, we have identified apoptosis signal-regulating kinase 1 (ASK1) as a CDC25A-interacting protein by yeast two-hybrid screening. ASK1 activates the p38 mitogen-activated protein kinase (MAPK) and c-Jun NH(2)-terminal protein kinase-stress-activated protein kinase (JNK/SAPK) pathways upon various cellular stresses. Coimmunoprecipitation studies demonstrated that CDC25A physically associates with ASK1 in mammalian cells, and immunocytochemistry with confocal laser-scanning microscopy showed that these two proteins colocalize in the cytoplasm. The carboxyl terminus of CDC25A binds to a domain of ASK1 adjacent to its kinase domain and inhibits the kinase activity of ASK1, independent of and without effect on the phosphatase activity of CDC25A. This inhibitory action of CDC25A on ASK1 activity involves diminished homo-oligomerization of ASK1. Increased cellular expression of wild-type or phosphatase-inactive CDC25A from inducible transgenes suppresses oxidant-dependent activation of ASK1, p38, and JNK1 and reduces specific sensitivity to cell death triggered by oxidative stress, but not other apoptotic stimuli. Thus, increased expression of CDC25A, frequently observed in human cancers, could contribute to reduced cellular responsiveness to oxidative stress under mitogenic or oncogenic conditions, while it promotes cell cycle progression. These observations propose a mechanism of oncogenic transformation by the dual function of CDC25A on cell cycle progression and stress responses.

Animals↗
PubMed2001

[Human cytomegalovirus inhibits the proliferation of CFU-MK in vitro].

OBJECTIVE: To investigate the effect of human cytomegalovirus (HCMV) on the proliferation of colony forming unit-megakaryocyte (CFU-MK). METHODS: Semi-solid CFU-MK culture system was used to observe the effect of HCMV AD169 strain on CFU-MK growth of 20 cord blood samples. HCMV DNA and immediate early antigen (IEA) mRNA in CFU-MK were detected by in situ-polymerase chain reaction (IS-PCR) and reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: HCMV AD169 suppressed the differentiation and proliferation of CFU-MK in vitro significantly. The suppression was in a dose-dependent fashion. HCMV DNA was successfully detected in colony cells from viral infection group, and did the expression of HCMV IEA mRNA. CONCLUSION: HCMV AD169 can directly infect megakaryocyte progenitor and suppress their proliferation and differentiation.

Cell Proliferation↗