Biomedical subjects

H C Chen

Publications and source records attributed to H C Chen.

At least 271 records · Page 15Linked to original sources

PubMed1996

Signal transduction in cell-matrix interactions.

Cell-matrix interactions play critical roles in a variety of physiological and pathological processes by regulating cell migration, proliferation, and differentiation. Integrin family cell-adhesion receptors have been identified as the major receptors for various extracellular matrix proteins and shown to be capable of transducing biochemical signals across the plasma membrane to regulate cellular functions. Protein tyrosine phosphorylation has been implicated in a central role in integrin-initiated signal transduction. Other signaling pathways have also been shown to be activated by cell-matrix interactions. This chapter reviews our current understanding of the key molecular components and their interactions in the integrin-signaling pathways and examines the role of various signaling pathways and molecular players in the context of cellular functions regulated by cell-matrix interactions.

Animals↗

PubMed1996

Effects of HI-6 on muscle acetylcholine receptor: analysis on minimal reaction model.

AIM: To study the action of 1-(2-hydroxyiminomethyl-1-pyridino)-3-(4-carbamoyl-1-pyridino)-2-oxapropane dichloride (HI-6) on skeletal muscle acetylcholine receptor (N2-ChR). METHODS: N2-ChR was expressed in Xenopus laevis oocyte after injection with mRNA extracted from denervated rat leg muscles. The inward membrane currents induced by various concentrations of carbamylcholine and effects of HI-6 or d-tubocurarine on the currents were measured with voltage clamp technique by fast cell flow of agents. The actions of HI-6 and d-tubocurarine on N2-ChR were analyzed by using the minimal reaction model. RESULTS: K of 40.05, 156.00, and 334.67 mumol.L-1 for HI-6, K of 0.02, 0.10, and 0.18 mumol.L-1 for d-tubocurarine were obtained by using the competing for single acetylcholine (ACh)-binding site model, the competing for two ACh-binding sites model, and the noncompetitive inhibition model, respectively. CONCLUSION: HI-6 and d-tubocurarine competed for two ACh-binding sites of N2-ChR with equal affinity to antagonize the effects of the agonist on N2-ChR. The N2-ChR inhibition by HI-6 is much weaker than that by d-tubocurarine.

Animals↗

PubMed1995

Identification of a rapidly dephosphorylating 95-kDa protein as elongation factor 2 during 8-Br-cAMP treatment of N1E115 neuroblastoma cells.

Treatment of 8-Br-cAMP promotes neurite outgrowth and neuronal differentiation in N1E115 mouse neuroblastoma cells. Prior or simultaneous treatment of PMA blocks 8-Br-cAMP-mediated neurite outgrowth. Phosphorylation of cellular proteins during these treatments was examined in a permeabilized cell system. While PMA promotes phosphorylation of the heat-stable protein kinase C substrates MARCKS and neuromodulin, 8-Br-cAMP hastens the dephosphorylation of a protein of M(r)95k (p95). Extensively purified, N-terminal sequenced, and judged from its phosphorylation properties, p95 was identified as the eukaryotic elongation factor-2 (eEF-2), whose dephosphorylation has been reported to be related to an increase in protein synthesis. It is likely 8-Br-cAMP stimulates dephosphorylation of eEf-2, promotes protein synthesis that eventually leads to neuronal differentiation in N1E115 cells.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1995

The site and stoichiometry of the N-phenylmaleimide reaction with myosin when weakly-binding crossbridges are formed in skinned rabbit psoas fibers.

Treatment of relaxed skinned rabbit psoas muscle fibers with 0.1 mM N-phenylmaleimide (NPM) for 1 h locks all of the crossbridges in a weakly-binding state resembling that of the myosin.ATP crossbridge. Under these conditions, NPM reacts mainly with myosin heavy chain (Barnett et al. (1992) Biophys. J. 61, 358-367). Here the specific sites for that reaction are explored. Small bundles of rabbit psoas muscle fibers were treated with Triton X-100 to make the fiber sarcolemmas permeable. The bundles were treated with 0.1 mM [14C]NPM for 1 h, and homogenized for SDS-PAGE. 43 +/- 2.2% of the muscle fiber protein ran in the myosin heavy chain band, the same as for untreated fibers. An alkylating stoichiometry of 2.2 +/- 0.33 moles NPM per mole myosin heavy chain was determined. Exhaustive trypsin digestion followed by two-dimensional thin-layer chromatography and reverse-phase HPLC revealed two major sites on myosin heavy chain for NPM binding. The sites contained about the same amount of linked NPM, suggesting that the reaction stoichiometry of each site under the conditions studied is approx. 1 mol NPM/mol myosin heavy chain. Comparison of the labeled tryptic peptides with NPM-reacted synthetic SH1 and SH2 tryptic peptides and analysis of the treated fiber bundles' ATPase activity suggested that the sites for NPM reaction on myosin heavy chain when it locks crossbridges in a weakly-binding state are Cys-697 (SH2) and Cys-707 (SH1).

Animals↗

PubMed1995

Inhibition of the integrase of human immunodeficiency virus (HIV) type 1 by anti-HIV plant proteins MAP30 and GAP31.

MAP30 (Momordica anti-HIV protein of 30 kDa) and GAP31 (Gelonium anti-HIV protein of 31 kDa) are anti-HIV plant proteins that we have identified, purified, and cloned from the medicinal plants Momordica charantia and Gelonium multiflorum. These antiviral agents are capable of inhibiting infection of HIV type 1 (HIV-1) in T lymphocytes and monocytes as well as replication of the virus in already-infected cells. They are not toxic to normal uninfected cells because they are unable to enter healthy cells. MAP30 and GAP31 also possess an N-glycosidase activity on 28S ribosomal RNA and a topological activity on plasmid and viral DNAs including HIV-1 long terminal repeats (LTRs). LTRs are essential sites for integration of viral DNA into the host genome by viral integrase. We therefore investigated the effect of MAP30 and GAP31 on HIV-1 integrase. We report that both of these antiviral agents exhibit dose-dependent inhibition of HIV-1 integrase. Inhibition was observed in all of the three specific reactions catalyzed by the integrase, namely, 3' processing (specific cleavage of the dinucleotide GT from the viral substrate), strand transfer (integration), and "disintegration" (the reversal of strand transfer). Inhibition was studied by using oligonucleotide substrates with sequences corresponding to the U3 and U5 regions of HIV LTR. In the presence of 20 ng of viral substrate, 50 ng of target substrate, and 4 microM integrase, total inhibition was achieved at equimolar concentrations of the integrase and the antiviral proteins, with EC50 values of about 1 microM. Integration of viral DNA into the host chromosome is a vital step in the replicative cycle of retroviruses, including the AIDS virus. The inhibition of HIV-1 integrase by MAP30 and GAP31 suggests that impediment of viral DNA integration may play a key role in the anti-HIV activity of these plant proteins.

Antiviral Agents↗

PubMed1995

Anti-HIV and anti-tumor activities of recombinant MAP30 from bitter melon.

MAP30 is an anti-HIV plant protein that we have identified and purified to homogeneity from bitter melon (Momordica charantia). It is capable of acting against multiple stages of the viral life cycle, on acute infection as well as replication in chronically infected cells. In addition to antiviral action, MAP30 also possesses anti-tumor activity, topological inactivation of viral DNA, inhibition of viral integrase and cell-free ribosome-inactivation activities. We have cloned and expressed the MAP30 gene. The objective of this study is to characterize recombinant MAP30 (re-MAP30) and to determine its anti-HIV, anti-tumor and other activities. We report here that re-MAP30 inhibits HIV-1 and certain human tumors to the same extent as its native counterpart, natural MAP30 (nMAP30). The anti-HIV activity was measured by quantitative focal syncytium formation on CEM-ss cell monolayers, viral core protein p24 expression and viral-associated reverse transcriptase activity in HIV-1-infected H9 cells. The anti-tumor activity was measured by metabolic labeling of protein synthesis in tumor cells. In the dose range of the assay, re-MAP30 exhibits little toxicity to the uninfected viral target cells and other normal human cells. Identical to nMAP30, re-MAP30 is also active in topological inactivation of viral DNA, inhibition of viral DNA integration and cell-free ribosome inactivation. The cloning and expression of the gene encoding biologically active re-MAP30 provides an abundant source of homogeneous material for clinical investigations, as well as structure-function studies of this novel antiviral and anti-tumor agent.

Amino Acid Sequence↗

PubMed1995

Interaction of focal adhesion kinase with cytoskeletal protein talin.

The interaction of cells with extracellular matrix proteins plays a critical role in a variety of biological processes. Recent studies suggest that cell-matrix interactions mediated by integrins can transduce biochemical signals to the cell interior that regulate cell proliferation and differentiation. These studies have placed the focal adhesion kinase (FAK), an intracellular protein tyrosine kinase, in a central position in integrin-initiated signal transduction pathways (Zachary, I., and Rozengurt, E. (1992) Cell 71, 891-894; Schaller, M., and Parsons, J. T. (1993) Trends Cell Biol. 3, 258-262). Here, we report data suggesting a possible association of FAK with the cytoskeletal protein talin in NIH 3T3 cells. We have identified a 48-amino acid sequence in the carboxyl-terminal domain of FAK necessary for talin binding in vitro. Furthermore, we have correlated the ability of integrin to induce FAK phosphorylation with its ability to bind talin using a mutant integrin lacking the carboxyl-terminal 13 amino acids. These studies suggest talin may be a mediator for FAK activation in signaling initiated by integrins and may provide an explanation for the dependence on the integrity of actin-cytoskeleton of multiple intracellular signaling pathways converging to FAK activation and autophosphorylation.

3T3 Cells↗

PubMed1995

Cloning and promoter mapping of mouse kappa opioid receptor gene.

We have isolated mouse kappa opioid receptor (KOR) genomic clones containing the complete 5'-untranslated region, a portion of the 3'-untranslated region, and its coding region. The mouse KOR gene spans at least 16 kb and contains at least 4 exons. Exon I encodes the major portion of 5'-untranslated sequence and transcription initiation is determined, by primer extension experiments, to reside at 3 major sites, located at 334, 340, and 716 nucleotides upstream from the translation initiation codon. No typical TATA or CAAT boxes can be identified within an approximately 1-kb upstream region. Exon II contains 271 nucleotides, including 14 nucleotides of 5'-untranslated sequence. Exon III contains 353 nucleotides and exon IV begins at amino acid position #204 and encodes the rest of the 3'-end sequence of the cloned cDNA. Based upon Northern blot analysis, KOR transcript is estimated to be approximately 6 kb in length.

Amino Acid Sequence↗

PubMed1995

Dephosphorylation of protein kinase C substrates, neurogranin, neuromodulin, and MARCKS, by calcineurin and protein phosphatases 1 and 2A.

Neurogranin, neuromodulin, and MARCKS are among the most prominent substrates of protein kinase C (PKC) in the mammalian brain. These phosphoproteins were dephosphorylated by three isoforms of rat brain calcineurin, also known as calmodulin (CaM)-dependent protein phosphatase (CaMPP). The three CaMPP isozymes dephosphorylate neurogranin, the most favorable substrate among the three tested, with subtle differences in their responses to divalent metal ions, Mn2+ and Ni2+. Dephosphorylation of neurogranin by all three CaMPP isozymes, CaMPP-1, -2, and -3, were stimulated to a higher extent by Mn2+ than by Ni2+ in the presence of CaM and Ca2+. The Km values of neurogranin in the presence of Mn2+ were lower than those in the presence of Ni2+ for CaMPP-1 and -2, but that for CaMPP-3 was comparable with either divalent metal ion. The Vmax values were higher in the presence of Mn2+ than those of Ni2+ for all three isozymes. Neurogranin and neuromodulin, both phosphorylated by PKC at a single site, were dephosphorylated completely by CaMPP; however, MARCKS, phosphorylated by PKC at three sites, was partially dephosphorylated by this phosphatase. A higher extent of dephosphorylation of MARCKS could be achieved by the combination of CaMPP and protein phosphatase 2A and a complete dephosphorylation of this protein was observed with protein phosphatase 1. Protein phosphatase 1 and 2A were also effective in a complete dephosphorylation of neurogranin and neuromodulin. Amino acid sequence analysis of the tryptic phosphopeptides derived from MARCKS dephosphorylated by CaMPP and protein phosphatase 2A revealed that the former preferentially dephosphorylated Ser155 and the latter Ser162 of rat brain MARCKS. Both phosphatases dephosphorylated poorly of Ser151. Because of the high concentration of CaMPP in the brain and the colocalization of this phosphatase with major PKC substrates in the various brain regions, it is likely that CaMPP is a phosphatase with potential to reverse the action of PKC.

Amino Acid Sequence↗

PubMed1995

The versatility of free rectus femoris muscle flap: an alternative flap.

In the 10-year period from June 1985 to December 1994, 54 free rectus femoris muscle or musculocutaneous flaps were performed at our hospital. It has been one of the most frequently used free muscle flaps in our institution and forms 2% of all free tissue transfers (total, 2,769 cases). In 26 patients, it was used for large wound coverage following debridement or tumor ablation, and in 27 patients, as a functioning free muscle transplantation for brachial plexus palsy or traumatic muscle loss. In one patient the components of the deep aponeurosis, muscle, and overlying skin were used for reconstruction of an abdominal wall defect after neurilemmoma excision. There were two complete failures, one due to diabetic foot infection and one due to venous occlusion. Four had superficial marginal skin necrosis. No significant disability of the donor limb was encountered. Easy approach, rapid harvest, large and reliable overlying skin flap, a single dominant neurovascular pedicle (with large vessel diameter and long motor nerve), easy primary closure of the donor site, and minimal donor site morbidity all make the rectus femoris flap a good alternative flap for free tissue transfer, in addition to the gracilis, rectus abdominis, and latissimus dorsi muscle flap.

Adult↗

PubMed1995

Expression of the CMS-associated urfS sequence in transgenic petunia and tobacco.

The expression of a 25 kDa protein, encoded by the fused mitochondrial pcf gene, is associated with cytoplasmic male sterility (CMS) in petunia. To investigate the role of the 25 kDa protein in CMS we have transformed petunia and tobacco plants with constructs expressing a portion of the urfS sequence of the pcf cDNA which encodes the 25 kDa protein. The urfS sequence was fused with two different mitochondrial targeting sequences. The chimeric gene coding region was placed under the control of the CaMV 35S promoter or a tapetum-specific promoter. Expression of the PCF protein was obtained in mitochondria of transgenic petunia and tobacco plants, yet fertility of the plants was not affected. Analysis of the location of the urfS-encoded protein revealed that it fractionates primarily into the soluble fraction in the transgenic plants whereas the genuine 25 kDa protein is found primarily in the soluble fraction but also in the membrane portion of immature buds from CMS petunia plants. Fertile transgenic plants were obtained which expressed the 25 kDa protein in the tapetal layer of post-meiotic anthers, while CMS plants express the endogenous 25 kDa protein in both the tapetal layer and sporogenous tissue of pre-meiotic anthers.

Amino Acid Sequence↗

PubMed1995

Active compounds from Saussurea lappa Clarks that suppress hepatitis B virus surface antigen gene expression in human hepatoma cells.

We have examined the antiviral activity of the crude extract prepared from the root of Saussurea lappa Clarks, a Chinese medicinal herb which is widely used for many illnesses including cancer. Two active components, costunolide and dehydrocostus lactone, were identified which show strong suppressive effect on the expression of the hepatitis B surface antigen (HBsAg) in human hepatoma Hep3B cells, but have little effect on the viability of the cells. Both costunolide and dehydrocostus lactone suppress the HBsAg production by Hep3B cells in a dose-dependent manner with IC50s of 1.0 and 2.0 microM, respectively. Northern blotting analysis shows that the suppression of HBsAg gene expression by both costunolide and dehydrocostus lactone were mainly at the mRNA level. Furthermore, the suppressive effect of costunolide and dehydrocostus lactone on HBsAg and hepatitis B e antigen (HBeAg), a marker for hepatitis B viral genome replication in human liver cells, was also observed in another human hepatoma cell line HepA2 which was derived from HepG2 cells by transfecting a tandemly repeat hepatitis B virus (HBV) DNA. Similarly, the mRNA of HBsAg in HepA2 cells was also suppressed by these two compounds. Our findings suggest that costunolide and dehydrocostus lactone may have potential to develop as specific anti-HBV drugs in the future.

Antiviral Agents↗

PubMed1995

Microflora and tetrodotoxin-producing bacteria in a gastropod, Niotha clathrata.

Shellfish (Niotha clathrata) were collected in both July and November from three locations in Taiwan (Pingtung, Kaohsiung and Chiai Prefecture) and assayed for anatomical distribution of tetrodotoxin (TTX) and aerobic heterotrophic bacteria. Pingtung specimens showed higher toxicity than those from Kaohsiung and Chiai, and did not show much seasonal variation. At each site, the total aerobic bacterial counts in November samples were higher than in July. The predominant genera were Vibrio, Pseudomonas, Pasteurella, Aeromonas and Plesiomonas. Vibrio comprised more than 35% of the genera, with V. alginolyticus as the major species. The viable counts of Vibrio species were higher in November than in July. However, the results did not suggest any relationship between the total count or viable count and the toxicity of the shellfish. HPLC, UV and gas chromatographic-mass spectrometric analyses demonstrated that some of the bacteria isolated, such as V. alginolyticus, V. parahaemolyticus, Pseudomonas spp. Plesiomonas sp. and Aeromonas sp., produced TTX and/or related substances.

Animals↗

PubMed1995

Polymicrobial bacterial pericarditis with mediastinitis after endotracheal intubation.

Polymicrobial bacterial pericarditis with mediastinitis after traumatic endotracheal intubation is an unusual condition. We report a 54-year-old man, a victim of nasopharyngeal carcinoma, who developed pericardial effusion with tamponade after traumatic endotracheal intubation. The diagnosis of polymicrobial bacterial pericarditis was made when pericardiocentesis revealed purulent fluid that grew a mixed culture of anaerobic and aerobic bacteria, reflecting the normal upper airway flora. By bronchoscopic study, a laceration over the lower trachea was detected. The patient received prolonged aggressive antibiotic therapy and pericardial drainage. The infection improved, but the patient later died from another episode of nosocomial infection.

Anti-Bacterial Agents↗

PubMed1995

The zeta protein kinase C isoform from the testis of the grey mullet Mugil cephalus with a specific reaction protein of M(r) 48,000 on oolemma.

The zeta protein kinase C isoform (PKC-zeta) was purified from the testis of the grey mullet Mugil cephalus and has relative masses (M(r)) of 65,000 and 63,000. The subunits of PKC-zeta from spermatozoa degenerated to M(r) 58,000 and 53,000 after continuous freezing and thawing. Proteins of M(r) 48,000 on the oolemma of the grey mullet Mugil cephalus were found to be the reaction proteins of the PKC-zeta from spermatozoa.

Amino Acid Sequence↗

PubMed1995

Metabolism of cyclic carotenoids: a model for the alteration of this biosynthetic pathway in Capsicum annuum chromoplasts.

The biosynthetic pathway of cyclic carotenoid is known to be quantitatively and qualitatively different in the non-green plastids of Capsicum annuum fruits compared with chloroplasts. Here, the cloning is described of a novel cDNA from this organism, which encodes an enzyme catalyzing the cyclization of lycopene to beta-carotene when expressed in Escherichia coli. The corresponding gene is constitutively expressed during fruit development. Significant amino acid sequence identity was observed between this enzyme and capsanthin/capsorubin synthase which is involved in the synthesis of the species-specific red carotenoids of C. annuum fruits. The latter enzyme was found also to possess a lycopene beta-cyclase activity when expressed in E. coli. A model is proposed for the origin of the capsanthin/capsorubin synthase gene and the role of this enzyme, together with the newly cloned lycopene cyclase, in the specific re-channeling of linear carotenoids into beta-cyclic carotenoids in C. annuum ripening fruits.

Amino Acid Sequence↗

PubMed1995

Neuropsychological predictors of complex obstacle avoidance in healthy older adults.

Global cognitive impairment in older adults has been associated with a greater risk of falling, and tripping has been implicated as an important factor in a large percentage of these falls. In order to evaluate the role of specific cognitive domains in tripping and falling, 23 healthy older adults completed basic and complex obstacle avoidance tasks, as well as a battery of neuropsychological tests. Using multiple regression analysis, a select pattern of neuropsychological measures was found to predict the decrement in performance evident as avoidance task complexity increased. Whereas measures of problem solving, response inhibition, general anxiety, and variability in attention were found to be significant predictors (in that order) of the relative decline in successful obstacle avoidance, measures of visuo-spatial discrimination and memory did not.

Accidental Falls↗

PubMed1995

Impact of decreased serum transaminase levels on the evaluation of viral hepatitis in hemodialysis patients.

The value of serum transaminases (ST) in evaluating hepatitis B (HBV) and C (HCV) infection was studied in 217 hemodialysis (HD) patients and 804 normal controls. Mean serum aspartate aminotransferase (AST) was 22.3 (22.0-22.7) and 22.6 (21.6-23.6) IU/l or 0.371 (0.366-0.378) and 0.376 (0.36-0.393) mu kat/l in controls and HD patients, respectively (nonsignificant), while mean serum alanine aminotransferase (ALT) was 20.3 (19.9-20.7) and 16.3 (15.3-17.3) IU/l or 0.338 (0.331-0.345) and 0.271 (0.255-0.288) mu kat/l in these two groups (p < 0.001). However, both AST and ALT became significantly depressed in HD patients after adjusting for age, gender, HBV surface antigen (HBsAg) and anti-HCV. The usual practice of regarding AST and ALT as being 'abnormal' in evaluating viral hepatitis when they exceeded the upper reference range (40 and 46 IU/l or 0.666 and 0.766 mu kat/l in our laboratory) was then critically assessed by the receiver operating characteristic (ROC) curve. ROC analysis showed that ST was useless in detecting HBsAg, while the best cutoff point for detecting the presence of anti-HCV was 18 IU/l (0.3 mu kat/l) for AST and 16 IU/l (0.266 mu kat/l) for ALT in HD patients, respectively. These are considerably lower than the conventional criteria for an 'abnormal' ST. We conclude that ST are decreased in HD patients and that the cutoff value of ST for detecting HCV should be set at lower levels to enhance their diagnostic yield.

Alanine Transaminase↗