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Biomedical subjects

C Li

Publications and source records attributed to C Li.

At least 127 records · Page 7Linked to original sources
PubMed2001

Increased serum VEGF in 13 children with Wilms' tumour falls after surgery but rising levels predict poor prognosis.

Vascular endothelial cell growth factor (VEGF, a potent endothelial cell mitogen in vitro) may be important in tumour development and its spread in vivo. In this preliminary study, we tested the hypotheses that (i) raised serum levels in Wilms' tumour fall after surgery, and (ii) rising levels predict adverse outcome. Serum VEGF was measured (ELISA) in 13 children about to undergo surgery, and serially on the following day, a week later, and finally 3-6 months after surgery. A simple follow-up at 6 months was also performed. The control group was 60 healthy adults. Before surgery, the median (inter-quartile range) VEGF in the children was 20 ng/ml (10.4-70.5) and was 1 ng/ml (0.5-4.0) in the adults. This difference is statistically highly significant (P=0.0001). After surgery, levels in the children fell significantly to 1.3 ng/ml (0.5-7.95) the following day and to 1.9 ng/ml (0.5-5.0) the following week (P<0.001, ANOVA). Six months after surgery, three of the children had died. A level of >10 ng/ml measured 3 months after surgery correctly identified all three deaths: levels were 1.3 ng/ml (0.5-1.9) in the survivors and were 20.0 ng/ml (15-104) in those who died. These data indicate a rapid reduction in raised VEGF following surgery for Wilms' tumour, but that rising levels predict poor prognosis.

Adolescent↗
PubMed2001

Direct extraction of specific pharmacophoric flavonoids from gingko leaves using a molecularly imprinted polymer for quercetin.

A new straightforward approach to extract active components from Chinese traditional herb was reported by using a molecularly imprinted polymer (MIP) as the sorbent material. The MIP was prepared using quercetin, a typical active compound of the flavonoid family in gingko leaves as the template. Acrylamide was used as the functional monomer and two polar solvent tetrahydrofuran was used as the porogen. Afterwards, the polymer was evaluated by chromatographic analysis. It exhibited high selectivity for quercetin and good affinity to its structural analogues. Specific binding amount of quercetin on the MIP in the solvent methanol was 12 microg/g polymer by the cartridge test. The MIP cartridge could directly trap a specific class of compounds including quercetin and kaempferol from the hydrolyzate of gingko leaves. The result demonstrated the possibility of direct extraction of certain pharmacophoric constituents from herb by MIP technology. The compounds extracted were confirmed by time-of-flight mass spectrometry.

Chromatography, High Pressure Liquid↗
PubMed2001

Induction of melanoma in murine macrophage inflammatory protein 2 transgenic mice heterozygous for inhibitor of kinase/alternate reading frame.

The molecular and genetic events that contribute to the genesis and progression of cutaneous malignant melanoma are poorly understood, attributable in large part to the different genetic alterations accompanying tumorigenesis. Inhibitor of kinase 4a (INK4a) is often inactivated in families with hereditary melanoma. Loss of INK4a/alternate reading frame (ARF) in mice is associated with increased incidence of other tumors such as lymphoma and fibrosarcoma. However, the incidence of melanoma in INK4a/ARF-deficient mice is very low. Our previous studies have revealed that the CXC chemokine, CXCL1, is overexpressed in human malignant melanoma cells and is linked to transformation of immortalized murine melanocytes. To study the direct role of CXCL1 on the genesis of primary melanoma lesions, transgenic mouse lines were established that express the murine homologue of CXCL1, murine macrophage inflammatory protein 2 (MIP-2), under the transcriptional control of the tyrosinase promoter/enhancer (Tyr-MIP-2) in the mice that were deficient or not deficient for INK4a/ARF. Strong MIP-2 immunoreactivity was associated with pigmented melanocytes in the hyperproliferative hair follicles in the Tyr-MIP-2 transgenic mice, and the level of MIP-2 expression was similar in both INK4a/ARF heterozygous or wild-type mice. After treatment of mice with 7,12-dimethylbenz(a)anthracene, cutaneous melanomas formed in 12% (17/145) of the Tyr-MIP-2 transgene-positive mice, whereas only 2% (3/146) of the Tyr-MIP-2 transgene-negative mice developed melanoma. When melanocytes cultured from MIP-2 transgenic mice null for INK4a/ARF were transplanted into nude mice, melanoma formation occurred in 83% (10/12) of the cases with a latency period of 3 months. However, no melanoma lesions arose in nude mice injected with INK4a/ARF -/- melanocytes, which did not express the MIP-2 transgene. Our results demonstrate that constitutive expression of MIP-2 in INK4a/ARF-deficient melanocytes facilitates formation of malignant melanoma.

Animals↗
PubMed2001

Classification of human lung carcinomas by mRNA expression profiling reveals distinct adenocarcinoma subclasses.

We have generated a molecular taxonomy of lung carcinoma, the leading cause of cancer death in the United States and worldwide. Using oligonucleotide microarrays, we analyzed mRNA expression levels corresponding to 12,600 transcript sequences in 186 lung tumor samples, including 139 adenocarcinomas resected from the lung. Hierarchical and probabilistic clustering of expression data defined distinct subclasses of lung adenocarcinoma. Among these were tumors with high relative expression of neuroendocrine genes and of type II pneumocyte genes, respectively. Retrospective analysis revealed a less favorable outcome for the adenocarcinomas with neuroendocrine gene expression. The diagnostic potential of expression profiling is emphasized by its ability to discriminate primary lung adenocarcinomas from metastases of extra-pulmonary origin. These results suggest that integration of expression profile data with clinical parameters could aid in diagnosis of lung cancer patients.

Adenocarcinoma↗
PubMed2001

Human adenovirus proteinase: DNA binding and stimulation of proteinase activity by DNA.

The interaction of the human adenovirus proteinase (AVP) with various DNAs was characterized. AVP requires two cofactors for maximal activity, the 11-amino acid residue peptide from the C-terminus of adenovirus precursor protein pVI (pVIc) and the viral DNA. DNA binding was monitored by changes in enzyme activity or by fluorescence anisotropy. The equilibrium dissociation constants for the binding of AVP and AVP-pVIc complexes to 12-mer double-stranded (ds) DNA were 63 and 2.9 nM, respectively. DNA binding was not sequence specific; the stoichiometry of binding was proportional to the length of the DNA. Three molecules of the AVP-pVIc complex bound to 18-mer dsDNA and six molecules to 36-mer dsDNA. When AVP-pVIc complexes bound to 12-mer dsDNA, two sodium ions were displaced from the DNA. A Delta of -4.6 kcal for the nonelectrostatic free energy of binding indicated that a substantial component of the binding free energy results from nonspecific interactions between the AVP-pVIc complex and DNA. The cofactors altered the interaction of the enzyme with the fluorogenic substrate (Leu-Arg-Gly-Gly-NH)2-rhodamine. In the absence of any cofactor, the Km was 94.8 microM and the kcat was 0.002 s(-1). In the presence of adenovirus DNA, the Km decreased 10-fold and the kcat increased 11-fold. In the presence of pVIc, the Km decreased 10-fold and the kcat increased 118-fold. With both cofactors present, the kcat/Km ratio increased 34000-fold, compared to that with AVP alone. Binding to DNA was coincident with stimulation of proteinase activity by DNA. Although other proteinases have been shown to bind to DNA, stimulation of proteinase activity by DNA is unprecedented. A model is presented suggesting that AVP moves along the viral DNA looking for precursor protein cleavage sites much like RNA polymerase moves along DNA looking for a promoter.

Adenoviruses, Human↗
PubMed2001

Functional cloning of Src-like adapter protein-2 (SLAP-2), a novel inhibitor of antigen receptor signaling.

In an effort to identify novel therapeutic targets for autoimmunity and transplant rejection, we developed and performed a large-scale retroviral-based functional screen to select for proteins that inhibit antigen receptor-mediated activation of lymphocytes. In addition to known regulators of antigen receptor signaling, we identified a novel adaptor protein, SLAP-2 which shares 36% sequence similarity with the known Src-like adaptor protein, SLAP. Similar to SLAP, SLAP-2 is predominantly expressed in hematopoietic cells. Overexpression of SLAP-2 in B and T cell lines specifically impaired antigen receptor-mediated signaling events, including CD69 surface marker upregulation, nuclear factor of activated T cells (NFAT) promoter activation and calcium influx. Signaling induced by phorbol myristate acetate (PMA) and ionomycin was not significantly reduced, suggesting SLAP-2 functions proximally in the antigen receptor signaling cascade. The SLAP-2 protein contains an NH2-terminal myristoylation consensus sequence and SH3 and SH2 Src homology domains, but lacks a tyrosine kinase domain. In antigen receptor-stimulated cells, SLAP-2 associated with several tyrosine phosphorylated proteins, including the ubiquitin ligase Cbl. Deletion of the COOH terminus of SLAP-2 blocked function and abrogated its association with Cbl. Mutation of the putative myristoylation site of SLAP-2 compromised its inhibitory activity and impaired its localization to the membrane compartment. Our identification of the negative regulator SLAP-2 demonstrates that a retroviral-based screening strategy may be an efficient way to identify and characterize the function of key components of many signal transduction systems.

Adaptor Proteins, Signal Transducing↗
PubMed2001

Differential localization of the neurofibromatosis 1 (NF1) gene product, neurofibromin, with the F-actin or microtubule cytoskeleton during differentiation of telencephalic neurons.

The protein product of the neurofibromatosis 1 gene, neurofibromin, is abundantly expressed in the cerebral cortex during development, but its physiological role remains unknown. To gain insights into the functions of neurofibromin in neurons, we examined patterns of expression and subcellular localization of neurofibromin during neuronal differentiation. Western blot analysis of telencephali homogenates throughout chick embryogenesis revealed that neurofibromin expression increased during embryonic development. Further analysis showed that telencephalic neurons were also enriched in neurofibromin in culture and that a biphasic gain in expression correlated well with both phases of differentiation in culture, first with a massive outgrowth of processes and gains in neurotransmitter phenotype differentiation, and then with synapse formation. Compared to proteins associated with distinct cytoskeleton systems, the pattern of neurofibromin expression correlated closely with that of the cortical cytoskeleton protein paxillin. Moreover, analysis of immunofluorescence staining of neurofibromin showed that in the presence of a protein crosslinker which preserves both soluble and filamentous cytoskeleton proteins after extraction with Triton X-100, neurofibromin colocalized with F-actin only during the first differentiation phase. This colocalization persisted when the actin cytoskeleton was collapsed with cytochalasin D treatment. In contrast, during the second phase of differentiation neurofibromin colocalized with microtubules, but not F-actin, and the staining pattern was disrupted with nocodazole, but not cytochalasin. A constant finding under all conditions was the presence of neurofibromin in the nucleus, which supports the idea that the bipartite nuclear targeting sequence between residues 2555 and 2572 of neurofibromin may be functional. In summary, we have shown that telencephalic neurons and astroblasts are enriched in neurofibromin and that the subcellular targeting of neurofibromin toward the actin or the microtubule cytoskeleton is developmentally regulated.

Actin Cytoskeleton↗
PubMed2001

A comparison between microsatellite analysis and cytology of urine for the detection of bladder cancer.

We have studied, in a blinded fashion, cytology and microsatellite analysis on urine sediments of 143 Chinese with or without bladder lesions. Microsatellite analysis succeeded in all 143 individuals, while cytology could not be performed on 42 subjects due to insufficient urine samples. Both microsatellite analysis and routine urine cytology succeeded in 81 bladder cancer patients. Seventy-three patients (90%) were identified by microsatellite analysis, while only 41 patients (51%) were identified by cytology. However, four patients were identified by routine cytology but not by microsatellite analysis. Our results proved that microsatellite analysis is more sensitive than routine cytology but the two methods are complementary to each other.

Adult↗
PubMed2001

Interaction of the human adenovirus proteinase with its 11-amino acid cofactor pVIc.

The interaction of the human adenovirus proteinase (AVP) and AVP-DNA complexes with the 11-amino acid cofactor pVIc was characterized. The equilibrium dissociation constant for the binding of pVIc to AVP was 4.4 microM. The binding of AVP to 12-mer single-stranded DNA decreased the K(d) for the binding of pVIc to AVP to 0.09 microM. The pVIc-AVP complex hydrolyzed the substrate with a Michaelis constant (K(m)) of 3.7 microM and a catalytic rate constant (k(cat)) of 1.1 s(-1). In the presence of DNA, the K(m) increased less than 2-fold, and the k(cat) increased 3-fold. Alanine-scanning mutagenesis was performed to determine the contribution of individual pVIc side chains in the binding and stimulation of AVP. Two amino acid residues, Gly1' and Phe11', were the major determinants in the binding of pVIc to AVP, while Val2' and Phe11' were the major determinants in stimulating enzyme activity. Binding of AVP to DNA greatly suppressed the effects of the alanine substitutions on the binding of mutant pVIcs to AVP. Binding of either or both of the cofactors, pVIc or the viral DNA, to AVP did not dramatically alter its secondary structure as determined by vacuum ultraviolet circular dichroism. pVIc, when added to Hep-2 cells infected with adenovirus serotype 5, inhibited the synthesis of infectious virus, presumably by prematurely activating the proteinase so that it cleaved virion precursor proteins before virion assembly, thereby aborting the infection.

Adenoviruses, Human↗
PubMed2001

The dynamics of hepatitis C virus binding to platelets and 2 mononuclear cell lines.

Hepatitis C virus (HCV) binds to platelets in chronically infected patients where free HCV constitutes only about 5% of total circulating virus. Free HCV preferentially binds to human mononuclear cell lines but free and complexed virus binds equally to platelets. The extent of free HCV binding to human Molt-4 T cells (which express CD81) and to human promonocytic U937 cells or to platelets (which do not express CD81) was similar. The binding of free HCV to the cell lines was saturated at a virus dose of 1 IU HCV RNA per cell but binding to platelets was not saturable. Human anti-HCV IgG, but not anti-CD81, markedly inhibited HCV binding to target cells in a dose-dependent manner. Human antibodies to HCV hypervariable region 1 of E2 glycoprotein partially inhibited viral binding to target cells. Recombinant E2 also inhibited viral binding to target cells in a dose-dependent manner, with the efficacy of this decreasing in the rank order of Molt-4 cells more than U937 cells more than platelets. In contrast to HCV, recombinant E2 bound to Molt-4 cells to an extent markedly greater than that apparent with U937 cells or platelets. These results suggest that the binding of HCV to blood cells is mediated by multiple cell surface receptors and that recombinant E2 binding may not be representative of the interaction of the intact virus with target cells.

Antibodies, Viral↗
PubMed2001

Differential regulation of interleukin-12 and interleukin-10 by heat shock response in murine peritoneal macrophages.

Heat shock response is a conserved stress response and has been shown to have anti-inflammatory effects. We investigated the effect of heat shock response on LPS-induced production of IL-12 and IL-10, which are two important cytokines playing contradictory roles in regulation of immune response, by murine peritoneal macrophages. The data showed that induction of heat shock response strongly suppressed LPS-induced production of IL-12 while augmented that of IL-10, suggesting the pleiotropic effects of heat shock response on immune regulatory gene expression. Also, the novel observation on up-regulation of IL-10 by heat shock response adds to the mechanism by which heat shock response exerts its anti-inflammatory effects.

Animals↗
PubMed2001

An ossified Meckel's cartilage in two Cretaceous mammals and origin of the mammalian middle ear.

An ossified Meckel's cartilage has been recovered from two early Cretaceous mammals from China. This element is similar to Meckel's cartilage in prenatal and some postnatal extant mammals and indicates the relationship of Meckel's cartilage with the middle ear in early mammals. The evidence shows that brain expansion may not be the initial factor that caused the separation of postdentary bones from the dentary as middle ear ossicles during mammalian evolution. The failure of the dentary to seize reduced postdentary elements during ontogeny of early mammals is postulated as an alternative mechanism for the separation. Modifications of both feeding and hearing apparatuses in early mammals may have led to the development of the definitive mammalian middle ear.

Animals↗
PubMed2001

Isolation and preliminary biological assessment of AADGAPLIRFamide and SVPGVLRFamide from Caenorhabditis elegans.

To date, 9 FMRFamide-related peptides (FaRPs) have been structurally characterised from Caenorhabditis elegans. Radioimmunometrical screening of an ethanolic extract of C. elegans revealed the presence of two additional FaRPs that were purified by reverse-phase HPLC and subjected to Edman degradation analysis and gas-phase sequencing. Unequivocal primary structures for the two FaRPs were determined as Ala-Ala-Asp-Gly-Ala-Pro-Leu-Ile-Arg-Phe-NH(2) and Ser-Val-Pro-Gly-Val-Leu-Arg-Phe-NH(2). Using MALDI-TOF mass spectrometry, the molecular masses of the peptides were found to be 1032 Da (MH) and 875 Da (MH)(+), respectively. Two copies of AADGAPLIRFamide are predicted to be encoded on the precursor gene termed flp-13, while one copy of SVPGVLRFamide is located on flp-18. Synthetic replicates of the peptides were tested on Ascaris suum somatic muscle to assess bioactivity. ADDGAPLIRFamide had inhibitory effects on A. suum muscle strips, which occurred over a range of concentrations from a threshold for activity of 10 nM to 10 microM. SVPGVLRFamide was excitatory on A. suum somatic musculature from a threshold concentration for activity of 1 nM to 10 microM. The inhibitory and excitatory effects of AADGAPLIRFamide and SVPGVLRFamide, respectively, were the same for dorsal and ventral muscle strips as well as innervated and denervated preparations, suggesting that these physiological effects are not nerve cord dependent. Addition of ADDGAPLIRFamide (10 microM) to muscle strips preincubated in high-K(+) and -Ca(2+)-free medium resulted in a normal inhibitory response. Peptide addition to muscle strips preincubated in Cl(-)-free medium showed no inhibitory response, suggesting that the inhibitory response of the peptide may be chloride mediated. A normal excitatory response was noted following the addition of 10 microM SVPGVLRFamide to muscle strips preincubated in high-K(+), Ca(2+)- and Cl(-)-free media.

Animals↗
PubMed2001

A monomer is the minimum functional unit required for channel and ATPase activity of the cystic fibrosis transmembrane conductance regulator.

The cystic fibrosis transmembrane conductance regulator (CFTR) normally functions as a phosphorylation-regulated chloride channel on the apical surface of epithelial cells, and lack of this function is the primary cause for the fatal disease cystic fibrosis (CF). Previous studies showed that purified, reconstituted CFTR can function as a chloride channel and, further, that its intrinsic ATPase activity is required to regulate opening and closing of the channel gate. However, these previous studies did not identify the quaternary structure required to mediate conduction and catalysis. Our present studies show that CFTR molecules may self-associate in CHO and Sf9 membranes, as complexes close to the predicted size of CFTR dimers can be captured by chemical cross-linking reagents and detected using nondissociative PAGE. However, CFTR function does not require a multimeric complex for function as we determined that purified, reconstituted CFTR monomers are sufficient to mediate regulated chloride conduction and ATPase activity.

Adenosine Triphosphatases↗
PubMed2001

Molecular mechanisms for aberrant expression of the human breast cancer specific gene 1 in breast cancer cells: control of transcription by DNA methylation and intronic sequences.

Breast cancer specific gene 1 (BCSG1), also referred as synuclein gamma, is the third member of a neuronal protein family synuclein. BCSG1 is not expressed in normal breast tissues but highly expressed in advanced infiltrating breast carcinomas. When over expressed, BCSG1 significantly stimulates breast cancer metastasis. To elucidate the molecular mechanisms underlying the abnormal transcription of BCSG1 in breast cancer cells, in this study, we isolated a 2195 base pair fragment of human BCSG1 gene. This fragment includes 1 kb 5'-flanking region, exon 1, and intron 1. By analysing the promoter activity and the methylation status of the exon 1 region, we show that (1) Intron 1 plays critical roles in the control of BCSG1 gene transcription through cis-regulatory sequences that affect BCSG1 transcription in cell type-specific and cell type-nonspecific manners. (2) The activator protein-1 (AP-1) is functionally involved in BCSG1 transcription in breast cancer cells through its binding to an AP-1 motif located in the intron 1. (3) The exon 1 region of BCSG1 gene contains a CpG island that is unmethylated in BCSG1-positive SKBR-3 and T47D cells but densely methylated in BCSG1-negative MCF-7 cells. (4) Treating MCF-7 cells with a demethylating agent 5-Aza-2'-deoxycytidine specifically activated BCSG1 transcription. Thus, our results suggest that while the cellular content of transcription activators and repressors that interact with the cis-regulatory sequences present in the intron 1 contribute prominently to the tissue-specific expression of BCSG1, demethylation of exon 1 is an important factor responsible for the aberrant expression of BCSG1 in breast carcinomas.

Amino Acid Motifs↗
PubMed2001

Inhibition of MDM2 by hsp90 contributes to mutant p53 stabilization.

Stabilization and overexpression are hallmarks of mutant p53 found in nearly 50% of human tumors. Mutations in the conformation-sensitive core domain of p53 often lead to association with molecular chaperones such as hsp70 and hsp90. Inhibition of hsp90 function accelerates mutant p53 degradation. We recently found that expression of p53 core domain mutants inhibits MDM2 degradation, suggesting that mutant p53 can modulate MDM2 functions. In this report, we show that mutant p53 mediates formation of MDM2-p53-hsp90 complexes. Release of MDM2 from the p53-hsp90 complex after DNA damage restores MDM2 but not p53 turnover, whereas dissociation of hsp90 by geldanamycin increases the degradation of both MDM2 and mutant p53. Mutant p53 degradation after hsp90 inhibition requires MDM2 expression. The interaction between MDM2 and hsp90 is disrupted by the 2A10 antibody, which recognizes a site on MDM2 important for binding to alternative reading frame (ARF). Expression of mutant p53 prevents MDM2 from binding ARF and accumulating in the nucleolus in an hsp90-dependent fashion. These results suggest that hsp90 recruited by mutant p53 conceals the ARF-binding site on MDM2 and inhibits its ubiquitin-protein isopeptide ligase function, resulting in the stabilization of both mutant p53 and MDM2.

Benzoquinones↗
PubMed2001

Model-based analysis of oligonucleotide arrays: model validation, design issues and standard error application.

BACKGROUND: A model-based analysis of oligonucleotide expression arrays we developed previously uses a probe-sensitivity index to capture the response characteristic of a specific probe pair and calculates model-based expression indexes (MBEI). MBEI has standard error attached to it as a measure of accuracy. Here we investigate the stability of the probe-sensitivity index across different tissue types, the reproducibility of results in replicate experiments, and the use of MBEI in perfect match (PM)-only arrays. RESULTS: Probe-sensitivity indexes are stable across tissue types. The target gene's presence in many arrays of an array set allows the probe-sensitivity index to be estimated accurately. We extended the model to obtain expression values for PM-only arrays, and found that the 20-probe PM-only model is comparable to the 10-probe PM/MM difference model, in terms of the expression correlations with the original 20-probe PM/MM difference model. MBEI method is able to extend the reliable detection limit of expression to a lower mRNA concentration. The standard errors of MBEI can be used to construct confidence intervals of fold changes, and the lower confidence bound of fold change is a better ranking statistic for filtering genes. We can assign reliability indexes for genes in a specific cluster of interest in hierarchical clustering by resampling clustering trees. A software dChip implementing many of these analysis methods is made available. CONCLUSIONS: The model-based approach reduces the variability of low expression estimates, and provides a natural method of calculating expression values for PM-only arrays. The standard errors attached to expression values can be used to assess the reliability of downstream analysis.

Cluster Analysis↗