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Biomedical subjects

M Chen

Publications and source records attributed to M Chen.

At least 289 records · Page 16Linked to original sources
PubMed2000

[Effects of human chorionic gonadotropin on cellular cAMP levels in human gastro-adenocarcinoma SGC-7901 cells].

The purpose of this study was to determine the effects of hCG on cellular cAMP levels in gastric adenocarcinoma cells(SGC-7901). Radioimmunoassay was used. The results showed that hCG obviously increased cAMP levels in SGC-7901 cells in a dose-dependent and time-dependent manner. When the concentration of hCG increased from 10(-11) mol/L to 10(-6) mol/L, the cellular cAMP levels increased from about 2.1 pmol/mg protein to about 10 pmol/mg protein and were significantly different from the control(P < 0.01). These results suggest that hCG possibly exerts its effects on gastric cancer cells by activating adenylyl cyclase/cAMP signal transduction pathway.

Adenocarcinoma↗
PubMed2000

[Construction of genomic library from L. lactis AL2 and isolation of entire nisin biosynthesis gene cluster].

A total DNA library of Lactococcus lactis AL2 with high yield of nisin was successfully constructed using lambda EMBL3 as vector. 3900 plaques were obtained, which is much more than the required number of recombinants that represents an entire L. lactis genome according to Clarke and Carbon formula. Southern hybridization, PCR amplification and DNA sequencing revealed that the entire nisin biosynthesis gene cluster was isolated from the constructed library.

Base Sequence↗
PubMed2000

[A biomechanical study on the retrodiscal tissue of human temporomandibular joint].

In order to investigate the biomechanical properties and functional mechanism of the retrodiscal tissue (RT) of human temporomandibular joint (TMJ), we conducted a uniaxial tensile test on thirteen RT specimens taken from eight fresh TMJs of the human cadavers aged between 8-15 years (m = 11.8 yrs). The experimental data were analyzed in conjunction with the Quasi-Linear Viscoelastic Theory as proposed by Fung YC to characterize the time-dependent behaviour and the constitutive relationship of the RT. The results showed that the physiological biphasic zone of stress-strain curve lied within 20%-30% strain levels. The elastic modulus (MPa), tensile strength(MPa) and strain to failure(%) were high. There were significant differences in strain to yielding and energy resorption (N.mm) between lateral and medial RT. There was a good agreement between theoretical prediction value and experimental result within 6% strain level. These data suggest that there would be a higher passive deformation and lower safety storage of strength in the RT which could be related to anterior displacement of the TMJ disc and that Fung's Theory can adequately describe the stress-time behaviour and the constitutive relationship of the RT within lower strain rate.

Adolescent↗
PubMed2000

[The clinical significance of poststyloid space extension in nasopharyngeal carcinoma].

OBJECTIVE: To explore the clinical significance of poststyloid space extension in nasopharyngeal carcinoma. METHODS: One hundred and ninety-eight patients with newly treated nasopharyngeal carcinoma (stage I-IV a) were analyzed. These patients were divided into two groups depending on whether the poststyloid space was extended or normal. The 5-year survival rate, relapse and distant metastasis after the treatment were estimated using the Kaplan-Meier, Log-Rank methods. RESULTS: The rates of patients with oropharyngeal extension for these two groups were 72.1% and 16.8% respectively (P < 0.001). The rates of the cases with involvement of the 9, 10, 11, 12th cranial nerves were 20.9% and 6.2% respectively (P < 0.01). The rates of metastasis to cervical lymph node were 69.8% and 35.4% respectively (P < 0.01). The 5-year survival rates were 46.76% and 68.88% respectively (P = 0.0027). The local control rates were 63.7%, 78.7% respectively (P = 0.0862). The distant metastasis free survival rates were 71.7% and 84.2% respectively (P = 0.0468). These results showed that the survival rate, the local control rate were reduced and the distant metastasis was increased for patients with poststyloid space extension. CONCLUSIONS: The poststyloid space extension was an important prognostic factor with poorer treatment outcome. It is reasonable to classify such condition as stage T3 of nasopharyngeal carcinoma.

Adult↗
PubMed2000

[A computational spectral analysis method for multi-component drugs based on wavelet transform].

A novel computational spectral analysis method for multi-component drugs is proposed. The method applies the wavelet denoise processing technique to principle component regression(PCR). Since noise in the original spectral data is effectively filtered, the performance of PCR is obviously improved. A typical example in multi-component drug analysis has been used to verify the effectiveness of the novel algorithm. Compared with that obtained by PCR, the average of mean relative error which obtained by PCRW decreases to 0.46% from 1.48%.

Algorithms↗
PubMed1999

Expression of the casein kinase 2 subunits in Chinese hamster ovary and 3T3 L1 cells provides information on the role of the enzyme in cell proliferation and the cell cycle.

In order to investigate the in vivo functions of protein kinase CK2 (CK2), the expression of Myc-tagged versions of the subunits, Myc-CK2alpha and Myc-CK2beta, was carried out in Chinese hamster ovary cells (CHO cells) and in 3T3 L1 fibroblasts. Cell proliferation in these cells was examined. CHO cells that transiently overexpressed the Myc-CK2beta subunit exhibited a severe growth defect, as shown by a much lower value of [(3)H]thymidine incorporation than the vector controls, and a rounded shrunken morphology. In contrast, cells overexpressing Myc-tagged CK2alpha showed a slightly but consistently higher value of [(3)H]thymidine incorporation than the controls. The defect in cell growth and changes in morphology caused by Myc-CK2beta overexpression were partially rescued by coexpression of Myc-tagged CK2alpha. In parallel to the studies in CHO cells, the stable transfection of Myc-CK2alpha and Myc-CK2beta subunits was achieved in 3T3 L1 fibroblast cells. Similarly, the ectopic expression of Myc-CK2beta, but not Myc-CK2alpha, caused a growth defect. By measuring [(3)H]thymidine incorporation, it was found that expression of Myc-CK2beta prolonged the G(1) phase and inhibited up-regulation of cyclin D1 expression during G(1). In addition, a lower mitotic index and lower mitotic cyclin-dependent kinase activities were detected in Myc-CK2beta-expressing cells. Detailed analysis of stable cells that were synchronously released into the cell cycle revealed that the expression of Myc-CK2beta inhibited cells entering into mitosis and prevented the activation of mitotic cyclin-dependent kinases. Taken together, results from both transient and stable expression of CK2 subunits strongly suggest that CK2 may be involved in the control of cell growth and progression of the cell cycle.

3T3 Cells↗
PubMed1999

Activation of p53 tumor suppressor by hepatitis C virus core protein.

In addition to being a structural protein that packages the viral genomic RNA, hepatitis C virus (HCV) core protein possesses regulatory functions. In this report, we demonstrate that the HCV core protein could enhance the gene transactivation activity of the tumor suppressor p53, regardless of whether p53 was derived from an exogenous or an endogenous gene. The activation of p53 by the HCV core protein was supported by the observation that the HCV core protein could enhance the expression of p21(waf1/Cip1), a downstream effector gene of p53, in a p53-dependent manner. Further studies indicated that the HCV core protein could also suppress hepatocellular growth via p53. The HCV core protein and p53 could bind to each other in vitro, which was evidenced by the coimmunoprecipitation, the GST pull-down, and the Far-Western blot assays. The deletion-mapping analysis indicated that the carboxy-terminal sequence of p53 located between amino acids 366 and 380 was required for the core protein binding. These results raised the possibility that the HCV core protein might activate p53 through direct physical interaction. The persistent perturbation of p53 activity by the HCV core protein during chronic infection may have important consequences in HCV pathogenesis.

Carcinoma, Hepatocellular↗
PubMed1999

Age-related impairments in TCR/CD3 activation of ZAP-70 are associated with reduced tyrosine phosphorylations of zeta-chains and p59fyn/p56lck in human T cells.

The expression and catalytic activity of the protein tyrosine kinase (PTK) ZAP-70 are needed for normal intracellular signaling through the T-cell receptor (TCR)/CD3 complex. However, the possible effect of aging on the catalytic activity of ZAP-70 in human peripheral blood T cells stimulated via the TCR/CD3 complex is unknown. The current studies show that T cells from a substantial proportion of elderly humans (12) exhibit significant reductions in the catalytic activity, but not expression of ZAP-70 when stimulated by ligation of the TCR/CD3 with cross-linked anti-CD3epsilon monoclonal antibody OKT3. In addition, the reduced catalytic activity of ZAP-70 in T cells from elderly subjects was not restored to the normal levels in response to ligation of CD4 receptors, suggesting defects in PTKs linked to both CD3 and CD4 receptors. Other experiments demonstrated that the age-related impairments of ZAP-70 activation in anti-CD3-stimulated T cells were accompanied by decreased tyrosine phosphorylations of zeta-chains and autophosphorylations of the PTKs p561ck/p59fyn. Moreover, the age-related defects in these early TCR/CD3-mediated phosphorylation events were readily detectable in both CD45RO+ memory and CD45RA+ naive T cells. Thus, these results suggest that defects in early TCR/CD3-mediated phosphorylation events among CD45RO+ memory and CD45RA+ naive T cells from certain elderly humans may con tribute to impaired induction of ZAP-70 catalytic activity.

Aged↗
PubMed1999

A transient, neuron-wide form of CREB-mediated long-term facilitation can be stabilized at specific synapses by local protein synthesis.

In a culture system where a bifurcated Aplysia sensory neuron makes synapses with two motor neurons, repeated application of serotonin (5-HT) to one synapse produces a CREB-mediated, synapse-specific, long-term facilitation, which can be captured at the opposite synapse by a single pulse of 5-HT. Repeated pulses of 5-HT applied to the cell body of the sensory neuron produce a CREB-dependent, cell-wide facilitation, which, unlike synapse-specific facilitation, is not associated with growth and does not persist beyond 48 hr. Persistent facilitation and synapse-specific growth can be induced by a single pulse of 5-HT applied to a peripheral synapse. Thus, the short-term process initiated by a single pulse of 5-HT serves not only to produce transient facilitation, but also to mark and stabilize any synapse of the neuron for long-term facilitation by means of a covalent mark and rapamycin-sensitive local protein synthesis.

Animals↗
PubMed1999

Analysis of variable region of T-cell receptor beta chain usage in the human anti-porcine xenoresponse.

BACKGROUND: The human xenoreactive T-cell receptor (TCR) repertoire is not well documented. The aim of this study was to analyze the TCR repertoire in human anti-porcine xenoresponses. METHODS: Peripheral blood lymphocytes (PBLs) from healthy donors were used to generate human T-cell lines against two different haplotypes of inbred Yucatan miniature swine (y/y and z/z). The variable region of TCR beta-chain (Vbeta) gene usage was determined by fluorescence CDR3 spectrotyping. RESULTS: TCR Vbeta usage of an established human antiporcine T-cell line analyzed at weeks 5, 7, and 9 showed a sequential increase in Vbeta 1, 2, 6.2, 11, and 19 as compared to unprimed peripheral blood lymphocytes, whereas the usage of other Vbetas decreased. The selection of limited Vbeta genes correlated with the sequential increase in the specific lysis of the T-cell line, suggesting a non-random clonal selection and expansion of T-cell clones that recognized porcine targets. Different Vbeta restriction was found using the same peripheral blood lymphocytes against a different haplotype of swine, indicating this selection of Vbeta gene was swine leukocyte antigen-dependent. CONCLUSIONS: There is restricted TCR Vbeta usage in the human anti-porcine response, suggesting that a limited number of xenogeneic epitopes are recognized by human T cells. The selection of particular TCR Vbeta clonotypes depends on the swine leukocyte antigen background.

Animals↗
PubMed1999

Endothelin-1-induced interleukin-8 production in human brain-derived endothelial cells is mediated by the protein kinase C and protein tyrosine kinase pathways.

We have previously demonstrated that endothelin-1 (Et-1) induces human central nervous system-derived endothelial cells (CNS-EC) to produce and secrete the chemokine interleukin 8 (IL-8). In the present study, we use specific inhibitors and activators to elucidate the signal transduction pathways involved in this process. Et-1-induced IL-8 production was blocked by ET(A) receptor antagonist BQ610, but not by ET(B) receptor antagonist BQ788, demonstrating that CNS-EC activation is initiated by Et-1 binding to the ET(A) receptor. IL-8 mRNA expression is blocked by the protein kinase C inhibitor bisindolylmaleimide or protein tyrosine kinase inhibitors, genestein and geldanamycin, establishing the involvement of the protein kinase C and protein tyrosine kinase pathways in the activation process. The transcription factor, NF-kappaB, is involved in Et-1 activation as determined by specific inhibitors of translocation and direct analysis of DNA-binding proteins. Neither inhibition nor activation of cAMP-dependent protein kinase affected IL-8 production in the absence or presence of Et-1. Similarly, no effect was observed upon inhibition of protein phosphatases by okadaic acid. Thus, the signal transduction process induced by Et-1 in CNS-EC, leading to increased mRNA IL-8 expression, is initiated by Et-1 binding to ET(A) receptor followed by subsequent activation of protein kinase C, protein tyrosine kinase, and NF-kappaB. Because increased expression of Et-1 is associated with hypertension and stroke and IL-8 is likely to be involved in the accumulation of neutrophils causing tissue damage in ischemic/reperfusion injury, identification of the mechanism involved in the Et-1-induced increase in IL-8 production may have significant therapeutic value.

Antihypertensive Agents↗
PubMed1999

Bullous systemic lupus erythematosus (type I) in a dog.

In human patients with systemic lupus erythematosus, cutaneous subepidermal blistering can occur because of the production of antibodies specific for basement membrane antigens. This condition is referred to as bullous systemic lupus erythematosus (BSLE). A dog was diagnosed with BSLE because it fulfilled the following criteria: (i) a diagnosis of systemic lupus erythematosus by standard methods; (ii) an acquired, vesicular, erosive and ulcerative eruption; (iii) microscopical subepidermal vesicles with neutrophil-predominant inflammation at the dermo-epidermal junction; (iv) deposition of IgG at the epidermal basement membrane zone; and (v) circulating IgG autoantibodies against type VII collagen. Anti-collagen VII type I-BSLE therefore needs to be considered as a possible differential diagnosis for canine autoimmune subepidermal blistering diseases.

Animals↗
PubMed1999

Alpha 2 beta 1 integrin mediates dermal fibroblast attachment to type VII collagen via a 158-amino-acid segment of the NC1 domain.

Dermal fibroblasts are in apposition to type VII (anchoring fibril) collagen in both unwounded and wounded skin. The NC1 domain of type VII collagen contains multiple submodules with homology to known adhesive molecules, including fibronectin type III-like repeats and a potential RGD cell attachment site. We previously reported the structure and matrix binding properties of authentic and recombinant NC1. In this study, we examined the interaction between dermal fibroblasts and the NC1 domain of type VII collagen. We found that both recombinant and authentic NC1 vigorously promoted human fibroblast attachment. Adhesion of fibroblasts to NC1 was dose dependent, saturable, and abolished by both polyclonal and monoclonal antibodies to NC1. Cell adhesion to NC1 was divalent cation dependent and specifically inhibited by a monoclonal antibody directed against the alpha2 or beta1 integrin subunits, but not by the presence of RGD peptides. Furthermore, the cell-binding activity of NC1 was not conformation dependent, since heat-denatured NC1 still promoted cell adhesion. Using a series of recombinant NC1 deletion mutant proteins, the cell binding site of NC1 was mapped to a 158-aa (residues 202-360) subdomain. We conclude that human dermal fibroblasts interact with the NC1 domain of type VII collagen and this cell-matrix interaction is mediated by the alpha2beta1 integrin and is RGD independent.

Binding Sites↗
PubMed1999

A post-stishovite SiO2 polymorph in the meteorite Shergotty: implications for impact events.

Transmission electron microscopy and electron diffraction show that the martian meteorite Shergotty, a shocked achondrite, contains a dense orthorhombic SiO2 phase similar to post-stishovite SiO2 with the alpha-PbO2 structure. If an SiO2 mineral exists in Earth's lower mantle, it would probably occur in a post-stishovite SiO2 structure. The presence of such a high-density polymorph in a shocked sample indicates that post-stishovite SiO2 structures may be used as indicators of extreme shock pressures.

Crystallography↗
PubMed1999

Characterization of the murine A1 adenosine receptor promoter, potent regulation by GATA-4 and Nkx2.5.

Adenosine acts via A1 adenosine receptors (A1ARs) in the heart and brain to potently influence mammalian physiology. A1ARs are expressed very early in embryonic development, and A1ARs are among the earliest expressed G protein coupled receptors in the heart and brain. To understand the biologic basis of A1AR expression, a genomic fragment containing the murine A1AR promoter was cloned. Reporter assay studies using DDT1 MF2 cells that express A1ARs revealed that 500 base pairs of the proximal A1AR promoter contained essential elements for A1AR gene expression. Transgenic mice with A1AR proximal promoter coupled with the beta-galactosidase reporter gene had heavy labeling of the brain and atria, consistent with normal patterns of A1AR expression. Within the proximal A1AR promoter, putative binding sites for cardiac transcription factors GATA and Nkx2.5 were identified. Co-expression studies revealed that GATA-4 and Nkx2.5 could individually drive A1AR promoter activity and act synergistically to activate A1AR expression. These observations suggest that embryonic A1AR expression involves activation of the A1AR promoter by GATA-4 and Nkx2.5.

Animals↗
PubMed1999

Topogenesis of cystic fibrosis transmembrane conductance regulator (CFTR): regulation by the amino terminal transmembrane sequences.

Cystic fibrosis transmembrane conductance regulator (CFTR) is a member of the ATP-binding cassette (ABC) transport superfamily. CFTR folding and assembly appear to involve several events occurred in the cytosol and ER. Misfolding of CFTR causes cystic fibrosis, and thus, understanding the folding mechanism of CFTR is extremely important. Recently, detailed study of membrane insertion process suggests that the first two transmembrane (TM) segments of CFTR have two distinct but independent mechanisms to ensure the correct membrane folding of its amino terminal end [Lu, Y., Xiong, X., Helm, A., Kimani, K., Bragin, A., Skach, W. R. (1998) J. Biol. Chem. 273, 568-576]. To understand how other TM segments are ensured to insert into membranes correctly, we investigated the topogenesis of TM3 and TM4 of CFTR in a cell-free expression system. We found that the correct membrane insertion of TM3 and TM4 of CFTR was ensured by their flanking amino acid sequences and controlled by the correct membrane insertion of their preceding TM1 and TM2. Thus, correct membrane insertion and folding of TM1 and TM2 play an essential role in the membrane insertion and folding of the subsequent TM segments of CFTR.

Amino Acid Sequence↗
PubMed1999

An improved method for routine preparation of intact artificial chromosome DNA (340-1000 kb) for transfection into human cells.

The transfer of high molecular weight (HMW) DNA into mammalian cells is an important strategy for assessing human gene expression and chromosome structure and function. However, using current methods, it is difficult to dependably prepare intact HMW DNA because of the susceptibility of the DNA to degradation and physical shearing. Here we describe a strategy whereby intact artificial chromosome DNA (as large as 1 Mb) can be routinely prepared from yeast. Strict adherence to this protocol has resulted in: (i) >90% of liquid DNA preparations containing largely intact DNA; (ii) transfection efficiencies for the development of stable human clonal cell lines ranging from 5 x 10(-7) to 8.8 x 10(-5); and (iii) the presence of markers from both YAC arms in 30-42% of the human fibrosarcoma cell HT1080 clones and 100% of the CF lung epithelial cell lines IB3-1 and CFT1 clones, suggesting that the HMW DNA is potentially intact in a substantial proportion of clones. Using this protocol for DNA preparation, successful transfection of functional 1 Mb human artificial chromosome DNA into human cells has also been achieved. This methodology should prove useful to those interested in using HMW human DNA for gene expression and functional analysis or for linear artificial chromosome construction, since integrity is absolutely critical for the success of these studies.

Cell Line↗