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Biomedical subjects

W Jiang

Publications and source records attributed to W Jiang.

At least 127 records · Page 7Linked to original sources
PubMed1999

Sequence analysis and expression of the aspartokinase and aspartate semialdehyde dehydrogenase operon from rifamycin SV-producing amycolatopsis mediterranei.

A approximately 4.8 kb KpnI fragment, from the upstream region of the methylmalonyl-CoA mutase gene (mutAB) of rifamycin SV-producing Amycolatopsis mediterranei, was cloned and partially sequenced. Codon preference analysis showed three complete ORFs. ORF2 is internal to ORF1, and encodes a polypeptide corresponding to 172 amino acids, whereas ORF1 encodes a polypeptide of 421 amino acids. They were identified as the encoding genes of aspartokinase alpha- and beta-subunits by comparing the amino acid sequences with those in the database. The downstream ORF3, whose start codon was overlapped with the stop codon of both ORF1 and ORF2 by 1 bp, was identified as the aspartate semialdehyde dehydrogenase gene (asd), encoding a polypeptide of 346 amino acids. Subclones containing either the ask gene or the asd gene were constructed, in which the genes could be expressed under Lac promoters. Two subclones could transform E. coli CGSC 5074 (ask-) and E. coli X6118 (asd-) to prototrophy, supporting the functional assignments. Southern hybridisation indicated that the approximately 4.8 kb sequenced region represented a continuous segment in the A. mediterranei chromosome. It is concluded that ask and asd genes are present in an operon in A. mediterranei, and therefore that organisation of these two genes is the same as in most gram-positive bacteria, such as Mycobacteria, Corynebacterium glutamicum and Bacillus subtilis, but is different from Streptomyces akiyoshiensis.

Actinobacteria↗
PubMed1999

Long-term cortical CBF recording by laser-Doppler flowmetry in awake freely moving rats subjected to reversible photothrombotic stroke.

This study aimed at developing a laser-Doppler flowmetry (LDF) device suitable for long-term cortical cerebral blood flow (cCBF) measurement in awake, freely moving rats. The device included a flow probe adapter for permanent fixation to the skull bone and a connector that held the flow probe in the adapter in exactly the same position during repeated cCBF recordings. With this LDF recording system, cCBF values were stable and unaltered in awake, freely moving rats up to 4 days after operation compared with initial recordings during anesthesia. Repeated cCBF measurements in rats after transient removal and reattachment of the flow probe revealed a coefficient of variation of 7.0-17.4%. The LDF recording system was applied to rats subjected to a photothrombotic ring stroke lesion. cCBF in the region-at-risk declined to 59-34-26-33% of baseline values (P < 0.01) at 1-2-24 48 h after irradiation with gradually restored cCBF values of 56-87% at 72-96 h post-irradiation (P < 0.01 vs. 24 h). Transcardial carbon black perfusion examination of the brains confirmed the sustained hypoperfusion in the region at risk up to 48 h post-ischemia followed by a consistently occurring late spontaneous reperfusion. In conclusion, a novel laser-Doppler cortical CBF recording system has been set up that allows stable long-term cortical CBF follow-up in awake, freely moving rats.

Animals↗
PubMed1999

Identification and localization of MEP1A-like sequences (MEP1AL1-4) in the human genome.

The human MEP1A gene encodes the meprin alpha subunit that consists of a protease domain conserved in the astacin family of metalloendopeptidases and several C-terminal interaction domains present in other proteins. Using the alpha subunit cDNA, we identified two clones from a human P1-derived artificial chromosome (PAC) library. Fluorescence in situ hybridization (FISH) mapped both PACs (1e12, 65a14) to chromosome 6p21, confirming the MEP1A location. FISH also mapped PAC 65a14 to chromosome 13cen, and to chromosome 9 in three different regions, 9p12-13, 9q21, and 9q22. Southern blot analysis showed that sequences of PAC 65a14 and MEP1A were similar in the 3' end but different in the 5' end, revealing for the first time that the human genome may encode multiple interaction domains highly similar to those of the meprin alpha subunit. The symbols of MEP1AL1, MEP1AL2, MEP1AL3, and MEP1AL4 have been designated for MEP1A-like sequences on 9p12-13, 9q21, 9q22, and 13cen, respectively.

Animals↗
PubMed1999

p53 protects against skin cancer induction by UV-B radiation.

To assess the role of the p53 tumor suppressor gene in skin carcinogenesis by UV radiation, mice constitutively lacking one or both copies of the functional p53 gene were compared to wild-type mice for their susceptibility to UV carcinogenesis. Heterozygous mice showed greatly increased susceptibility to skin cancer induction, and homozygous p53 knockout mice were even more susceptible. Accelerated tumor development in the heterozygotes was not associated with loss of the remaining wild-type allele of p53, as reported for tumors induced by other carcinogens, but in many cases was associated with UV-induced mutations in p53. Tumors arose on the ears and dorsal skin of mice of all three genotypes, and homozygous knockout mice also developed ocular tumors, mainly melanomas. Skin tumors in the p53 knockout mice were predominately squamous cell carcinomas and were associated with premalignant lesions resembling actinic keratoses, whereas those in the heterozygous and wild-type mice were mainly sarcomas. These results demonstrate the importance of p53 in protecting against UV-induced cancers, particularly in the eye and epidermis.

Animals↗
PubMed1999

On the approximation rate of hierarchical mixtures-of-experts for generalized linear models.

We investigate a class of hierarchical mixtures-of-experts (HME) models where generalized linear models with nonlinear mean functions of the form psi (alpha + xT beta) are mixed. Here psi (.) is the inverse link function. It is shown that mixtures of such mean functions can approximate a class of smooth functions of the form psi (h(x)), where h(.) epsilon W2;K infinity (a Sobolev class over [0, 1]s), as the number of experts m in the network increases. An upper bound of the approximation rate is given as O(m-2/s) in Lp norm. This rate can be achieved within the family of HME structures with no more than s-layers, where s is the dimension of the predictor x.

Algorithms↗
PubMed1999

Structure of the membrane domain of subunit b of the Escherichia coli F0F1 ATP synthase.

The structure of the N-terminal transmembrane domain (residues 1-34) of subunit b of the Escherichia coli F0F1-ATP synthase has been solved by two-dimensional 1H NMR in a membrane mimetic solvent mixture of chloroform/methanol/H2O (4:4:1). Residues 4-22 form an alpha-helix, which is likely to span the hydrophobic domain of the lipid bilayer to anchor the largely hydrophilic subunit b in the membrane. The helical structure is interrupted by a rigid bend in the region of residues 23-26 with alpha-helical structure resuming at Pro-27 at an angle offset by 20 degrees from the transmembrane helix. In native subunit b, the hinge region and C-terminal alpha-helical segment would connect the transmembrane helix to the cytoplasmic domain. The transmembrane domains of the two subunit b in F0 were shown to be close to each other by cross-linking experiments in which single Cys were substituted for residues 2-21 of the native subunit and b-b dimer formation tested after oxidation with Cu(II)(phenanthroline)2. Cys residues that formed disulfide cross-links were found with a periodicity indicative of one face of an alpha-helix, over the span of residues 2-18, where Cys at positions 2, 6, and 10 formed dimers in highest yield. A model for the dimer is presented based upon the NMR structure and distance constraints from the cross-linking data. The transmembrane alpha-helices are positioned at a 23 degrees angle to each other with the side chains of Thr-6, Gln-10, Phe-14, and Phe-17 at the interface between subunits. The change in direction of helical packing at the hinge region may be important in the functional interaction of the cytoplasmic domains.

Amino Acid Sequence↗
PubMed1999

Multistep regulation of DNA replication by Cdk phosphorylation of HsCdc6.

We have characterized HsCdc6, a human protein homologous to the budding yeast Cdc6p that is essential for DNA replication. We show that, unlike Cdc6p, the levels of HsCdc6 protein remain constant throughout the cell cycle in human cells. However, phosphorylation of HsCdc6 is regulated during the cell cycle. HsCdc6 is an excellent substrate for Cdk2 in vitro and is phosphorylated in vivo at three sites (Ser-54, Ser-74, and Ser-106) that are phosphorylated by Cdk2 in vitro, strongly suggesting that HsCdc6 is an in vivo Cdk substrate. HsCdc6 is nuclear in G1, but translocates to the cytoplasm at the start of S phase via Crm1-dependent export. An HsCdc6A1A2A3 mutant, which mimics unphosphorylated HsCdc6, is exclusively nuclear, and its expression inhibits initiation of DNA replication. An HsCdc6E1E2E3 mutant, which mimics phosphorylated HsCdc6, is exclusively cytoplasmic and is not associated with the chromatin/nuclear matrix fraction. Based on these results, we propose that phosphorylation of HsCdc6 by Cdks regulates DNA replication of at least two steps: first, by promoting initiation of DNA replication and, second, through nuclear exclusion preventing DNA rereplication.

Amino Acid Sequence↗
PubMed1999

Higher DNA adduct levels in urinary bladder and prostate of slow acetylator inbred rats administered 3,2'-dimethyl-4-aminobiphenyl.

Human epidemiological studies suggest associations between acetylator phenotype and aromatic amine-induced tumors. The aromatic amine carcinogen 3,2'-dimethyl-4-aminobiphenyl (DMABP) induces colon, prostate, and urinary bladder tumors in the rat, and a rapid and slow acetylator rat model has been characterized. The formation of DNA adducts has been used as a valuable biomarker in tumorigenesis. In order to examine the relationship between the acetylation polymorphism and aromatic amine-induced cancer, DNA adducts were measured in three target organs (colon, prostate, and urinary bladder) and two nontarget organs (liver and heart) of male rapid (F344) and slow (WKY) acetylator inbred rats administered DMABP. Two DMABP-DNA adducts, N-(deoxyguanosin-8-yl)-DMABP (C8-DMABP) and 5-(deoxyguanosin-N2-yl)-DMABP (N2-DMABP), were identified in each target and nontarget organ examined. C8-DMABP-DNA adduct levels were highest in liver and were dose related in liver, colon, urinary bladder, and prostate. DMABP-DNA adduct levels were significantly higher in the prostate and urinary bladder of slow acetylator vs rapid acetylator rats. These studies suggest that DMABP-induced DNA damage is acetylator phenotype-dependent in urinary bladder and prostate, two target organs for DMABP-induced tumorigenesis in the rat.

Acetylation↗
PubMed1999

Direct correlation between nitric oxide synthase II inducibility and metastatic ability of UV-2237 murine fibrosarcoma cells carrying mutant p53.

The relationship between nitric oxide synthase II (NOS II) inducibility and the metastatic ability of UV-2237 murine fibrosarcoma cells was determined. Highly metastatic cells survived to produce numerous lung metastases after i.v. injection in syngeneic C3H/HeN mice, whereas poorly metastatic cells did not. Highly metastatic clones exhibited higher levels of NOS II than did poorly metastatic clones in response to interleukin 1alpha and IFN-gamma stimulation. Furthermore, both poorly and highly metastatic clones contained an identical p53 mutation. Overexpression of NOS II in a highly metastatic clone by transfection with NOS II gene retarded tumor growth and completely suppressed metastasis. Our data indicate that a low to moderate level of NOS II expression directly correlates with metastatic ability of UV-2237 fibrosarcoma cells carrying mutant p53 and that a high level of nitric oxide production suppresses tumor growth and metastasis.

Animals↗
PubMed1999

Observation of transient disorder during myosin subfragment-1 binding to actin by stopped-flow fluorescence and millisecond time resolution electron cryomicroscopy: evidence that the start of the crossbridge power stroke in muscle has variable geometry.

The mechanism of binding of myosin subfragment-1 (S1) to actin in the absence of nucleotides was studied by a combination of stopped-flow fluorescence and ms time resolution electron microscopy. The fluorescence data were obtained by using pyrene-labeled actin and exhibit a lag phase. This demonstrates the presence of a transient intermediate after the collision complex and before the formation of the stable "rigor" complex. The transient intermediate predominates 2-15 ms after mixing, whereas the rigor complex predominates at time >50 ms. Electron microscopy of acto-S1 frozen 10 ms after mixing revealed disordered binding. Acto-S1 frozen at 50 ms or longer showed the "arrowhead" appearance characteristic of rigor. The most likely explanation of the disorder of the transient intermediate is that the binding is through one or more flexible loops on the surfaces of the proteins. The transition from disordered to ordered binding is likely to be part of the force-generating step in muscle.

Actins↗
PubMed1999

Effect of energy restriction on the expression of cyclin D1 and p27 during premalignant and malignant stages of chemically induced mammary carcinogenesis.

The restriction of energy intake has a profound inhibitory effect on carcinogenesis, yet the mechanism or mechanisms that account for this effect are unknown. In this experiment, the hypothesis tested was that energy restriction upregulates the expression of p27/kip1, a gene product associated with cell-cycle growth arrest, while downregulating cyclin D1, a protein that combines with cyclin-dependent kinases to promote phosphorylation of retinoblastoma protein and the progression of cells through the cell cycle. We studied levels of these proteins in uninvolved mammary epithelial cells and in mammary intraductal proliferations, ductal carcinomas in situ, and adenocarcinomas induced in response to administration of 1-methyl-1-nitrosourea in animals fed either ad libitum or 90%, 80%, or 60% of ad libitum intake. Protein levels were evaluated immunohistochemically by using computer-assisted image analysis to quantify differences in protein expression among treatment groups. The expression of p27 increased and the expression of cyclin D1 decreased dose-dependently in response to energy restriction. The effect was greater on p27 than on cyclin D1. The hypothesis proposed is that energy restriction inhibits carcinogenesis by arresting cell-cycle progression by regulating p27/kip1.

Adenocarcinoma↗
PubMed1999

Selenium-induced inhibition of angiogenesis in mammary cancer at chemopreventive levels of intake.

The trace element nutrient selenium (Se) has been shown to possess cancer-preventive activity in both animal models and humans, but the mechanisms by which this occurs remain to be elucidated. Because angiogenesis is obligatory for the genesis and growth of solid cancers, we investigated, in the study presented here, the hypothesis that Se may exert its cancer-preventive activity, at least in part, by inhibiting cancer-associated angiogenesis. The effects of chemopreventive levels of Se on the intra-tumoral microvessel density and the expression of vascular endothelial growth factor in 1-methyl-1-nitrosourea-induced rat mammary carcinomas and on the proliferation and survival and matrix metalloproteinase activity of human umbilical vein endothelial cells in vitro were examined. Increased Se intake as Se-enriched garlic, sodium selenite, or Se-methylselenocysteine led to a significant reduction of intra-tumoral microvessel density in mammary carcinomas, irrespective of the manner by which Se was provided: continuous exposure (7-wk feeding) with a chemoprevention protocol or acute bolus exposure (3 d) after carcinomas had established. Compared with the untreated controls, significantly lower levels of vascular endothelial growth factor expression were observed in a sizeable proportion of the Se-treated carcinomas. In contrast to the mammary carcinomas, the microvessel density of the uninvolved mammary glands was not altered by Se treatment. In cell culture, direct exposure of human umbilical vein endothelial cells to Se induced cell death predominantly through apoptosis, decreased the gelatinolytic activities of matrix metalloproteinase-2, or both. These results indicate a potential for Se metabolites to inhibit key attributes (proliferation, survival, and matrix degradation) of endothelial cells critical for angiogenic sprouting. Therefore, inhibition of angiogenesis associated with cancer may be a novel mechanism for the anticancer activity of Se in vivo, and multiple mechanisms are probably involved in mediating the anti-angiogenic activity.

Animals↗
PubMed1999

Mechanisms by which energy restriction inhibits carcinogenesis.

Cancer that occurs at numerous organ sites, including the colon and breast, is inhibited by energy restriction, and the inhibition is proportional to the degree of restriction imposed. In an effort to identify the mechanism(s) by which energy restriction exerts this effect, a short term model system of experimentally induced mammary carcinogenesis was used. Given that carcinogenesis is known to involve a dysregulation to tissue size homeostasis in which cell proliferation and cell death are in dysequilibrium, we hypothesized that energy restriction exerts its effect by altering one or more aspects of cell cycle regulation. It was observed that energy restriction inhibited cell proliferation and increased cell death due to apoptosis. Thus attention was next focused on aspects of cell cycle regulation that might be affected by energy restriction. It was observed that the amount of p27 protein, one member of the Cip/Kip family of genes that are involved in cell cycle arrest, was increased dose dependently by energy restriction. Based on this and related observations, the hypothesis is advanced that energy restriction inhibits carcinogenesis, at least in part, by delaying cell cycle progression via shifting cell populations into a G(0)/G(1)state. Ongoing work indicates that corticosteroids, which are produced in increased amounts in response to energy restriction, may be involved in mediating this effect.

Animals↗
PubMed1999

A photothrombotic ring stroke model in rats with sustained hypoperfusion followed by late spontaneous reperfusion in the region at risk.

In clinical thromboembolic stroke, spontaneous late recanalization is a common feature, but one which has been very sparsely studied experimentally. This study aimed at enabling the study of spontaneous reperfusion and exploring its consequences by modifying a recently developed photothrombotic-stroke model that focuses on the region-at-risk located within an ischemic ring-locus. The exposed crania of male Wistar rats (280-340 g) were subjected to a ring-shaped (5.0 mm outer diameter and 0.35 mm thick) laser-irradiation beam (514.5 nm; 0.89 W/cm2) for 2 min simultaneously with intravenous erythrosin B (17 mg/kg) infusion for 30 s. Transcardial carbon-black perfusion experiments revealed a ring-shaped cortical perfusion deficit at 4 h post-irradiation, which progressively increased at 10, 24, and 48 h, at which time the whole region-at-risk was pale with single distal branches of the middle cerebral artery being extensively narrowed, but not occluded. At 72 h, spontaneous reperfusion was observed in the region-at risk, which was even more pronounced at 7 and 28 days. Cortical cerebral blood flow (cCBF), measured by laser-Doppler flowmetry, was distinctly reduced at 2 min post-irradiation and further decreased slightly during 4 h of recording to ca. 24% of baseline values at the ring locus and 40% in the region-at-risk. In the region-at-risk, cCBF flow values were 23-30% of the baseline at 2448 h post-irradiation, followed by a relative cCBF increase to 71 and 77% at 72 and 96 h post-irradiation. Brain water content in the ischemic part of the cortex increased steadily from 4 to 48 h post-irradiation; at 72 h, it leveled off and returned to control values at 7 days. In conclusion, by employing a laser beam in the shape of a thin ring, critically sustained cCBF reduction was followed by late, consistent spontaneous reperfusion in the region-at-risk in this novel photochemically induced stroke-in-evolution model.

Animals↗
PubMed1999

A photothrombotic ring stroke model in rats with remarkable morphological tissue recovery in the region at risk.

The photothrombotic ring stroke model with sustained underperfusion followed by late spontaneous reperfusion (Gu et al. 1999) was employed to study its morphological consequences. The exposed crania of adult male Wistar rats were subjected to a ring-shaped laser irradiation beam simultaneously with intravenous erythrosin B infusion. The ischemic volume was calculated from serial sections throughout the ischemic lesions at 4, 10, 24, 48, and 72 h and 7 days and 28 days after irradiation. The ischemic volume, expressed as a percentage of the ipsilateral hemispheric volume, increased steadily from 4 to 10 to 24 h to reach its maximum value at 48 h after irradiation; at 3 days, 7 days, and 28 days, the ischemic volume was reduced to 75%, 24%, and 22% of the value at 48 h. Evaluation of ischemic volumes at different anteroposterior levels revealed that the reduced ischemic volume at 72 h and later was mainly due to morphological restoration in the centrally located, nonirradiated region at risk. An initial enlargement and development of cystic coagulation necrosis was observed in the cortical areas corresponding to the ring lesion itself. In the region at risk, a gradually deteriorating neuropil and nerve cell morphology were observed over time, with maximum severity at 48 h postirradiation. At this time, most laminae II and III neurons in the region at risk exhibited eosinophilia and pyknosis but no incrustations, with small islands of less damaged neurons randomly scattered. At 72 h and up to 28 days after irradiation, these cell characteristics were no longer observed and the region at risk was well populated with neurons that had a chiefly unremarkable cytological appearance. Neuronal counts in the central part of the region at risk were performed; no significant difference in neuronal density was observed between sham-operated controls and at 28 days after irradiation. In conclusion, the consistent, late spontaneous reperfusion coincided with remarkable tissue recovery as assessed morphologically in the region at risk. The data suggest that nerve cell repair may occur even after the detection, by conventional morphological methods, of prolonged critical ischemic neuronal damage in the setting of acute ischemic stroke.

Animals↗
PubMed1999

On the identifiability of mixtures-of-experts.

In mixtures-of-experts (ME) models, "experts" of generalized linear models are combined, according to a set of local weights called the "gating function". The invariant transformations of the ME probability density functions include the permutations of the expert labels and the translations of the parameters in the gating functions. Under certain conditions, we show that the ME systems are identifiable if the experts are ordered and the gating parameters are initialized. The conditions are validated for Poisson, gamma, normal and binomial experts.

Journal Article↗