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W C Chang

Publications and source records attributed to W C Chang.

At least 19 recordsLinked to original sources
PubMed1995

Down-regulation of epidermal growth factor-induced 12-lipoxygenase expression by glucocorticoids in human epidermoid carcinoma A431 cells.

The effect of glucocorticoids on epidermal growth factor (EGF)-induced expression of 12-lipoxygenase in human epidermoid carcinoma A431 cells was studied. A significant suppression of the EGF-induced expression of 12-lipoxygenase was observed in cells pretreated with 1 microM dexamethasone for 2 hr. The same pretreatment for 8 hr resulted in 55 and 54% inhibition of EGF-induced 12-lipoxygenase activity and mRNA expression, respectively. Cortisol, but not sex and mineral steroids, had a similar inhibitory effect. The glucocorticoid antagonist RU486 completely blocked the inhibitory effect of dexamethasone, suggesting that the action of dexamethasone was mediated through the ligation of glucocorticoid receptors. The results indicated that pretreatment of A431 cells with glucocorticoids resulted in a down-regulation of the EGF-induced expression of 12-lipoxygenase at the mRNA and enzyme activity level, which was mediated through glucocorticoid receptor activation.

Arachidonate 12-Lipoxygenase↗
PubMed1995

Changes of protein kinase C subspecies in staphylococcal enterotoxin-B-induced thymocyte apoptosis.

Our previous studies demonstrated that intravenous administration of staphylococcal enterotoxin B (SEB) to BALB/c mice resulted in thymocyte apoptosis. In the present study, we evaluated the role of protein kinase C (PKC) in thymocyte apoptosis induced by SEB. Our results showed that the level of protein phosphorylation in the thymocytes was reduced after the in vivo SEB treatment for 24 h. The activity of classical PKC subspecies was decreased in both cytosolic and membrane fractions of thymocytes following SEB administration. The lowest level of PKC activity was reached by 24 and 48 h, then was recovered gradually after 72 h. Furthermore, the mRNA expression of PKC-beta and, to a much less extent, PKC-alpha, but not PKC-sigma, in thymocytes was reduced by SEB. The decrease of mRNA level of PKC-beta showed good correlation with the pattern of PKC activity. These results provide direct evidence showing the changes in PKC subspecies mRNA expression during the process of apoptosis.

Animals↗
PubMed1995

Spontaneous dissociation-association of monomers of the human-stem-cell-factor dimer.

In its native state, recombinant human-stem-cell-factor (SCF) dimer can spontaneously and rapidly undergo hybridization when two different SCF dimer species are incubated together. SCF species differing in molecular charge, e.g., a wild-type SCF form and a variant with Asp at position 10 instead of Asn, were used in the hybridization studies; the original species and newly formed dimer hybrid can be separated and quantified by cationic-exchange h.p.l.c. The hybridization reaches an equilibrium where the ratio of hybrid dimer to each of the original species is 2. Kinetic studies of the initial rate of hybridization enable a rate constant for monomer dissociation to be determined. This rate constant is influenced by pH, temperature and salt concentration. The pH and salt effects suggest that salt bridges between charged amino acids at the monomer-monomer interface may be present. From the temperature effects, the activation energy for monomer dissociation was determined to be 85.6 kJ/mol, which is typical for oligomeric proteins. Heavily glycosylated recombinant SCF from Chinese-hamster ovary cells exchanged equally well with the bacterially derived non-glycosylated SCF, indicating that the attached carbohydrate moieties had no effect on monomer exchange.

Amino Acid Sequence↗
PubMed1995

Rasch analysis for outcomes measures: some methodological considerations.

OBJECTIVE: To assess the four approaches to applying Rasch analysis to ordinal, repeated measures: (1) to perform separate Rasch analyses; (2) to perform a single analysis by treating the subjects measured on different occasions as distinct ones; (3) to perform a single analysis by treating items used on different occasions as distinct ones; and (4) to construct a 3-facet model by adding occasions as the third facet. DESIGN: Various Rasch and 3-facet models applied to retrospective, cohort data. SETTING: A stroke program at a rehabilitation hospital. PATIENTS: 99 of the 107 consecutive patients admitted between March and October, 1990, average age 69 years, 40% of whom were women, with an average stay of 54 days. INTERVENTION: Medical inpatient rehabilitation. MAIN OUTCOME MEASURE: The Functional Independence Measure (FIM), administered within 1 week of admission and discharge, and approximately 6 weeks after discharge. RESULTS: The four approaches resulted in various occasion-specific and generalized measures of item difficulty, patient ability, and occasion severity. Although these measures turned out to be congruent with each other as they should, they generated different sources of information needed for formative and summative evaluation of health care outcomes. All four approaches were found to be useful, although each had its own limitations. A four-step strategy based on these approaches was therefore designed and recommended. CONCLUSION: Rasch analysis of ordinal, repeated measures poses a special challenge to those interested in health care outcomes. A multi-method strategy incorporating various approaches is therefore needed to optimize its use in health care research.

Activities of Daily Living↗
PubMed1995

Long-term expression of the biologically active growth hormone in genetically modified fibroblasts after implantation into a hypophysectomized rat.

We employed the hypophysectomized rats as an animal model to explore the feasibility of using genetically engineered fibroblast cells for growth hormone gene therapy. An internal ribosome entry site (IRES)-directed bicistronic retroviral vector, PSN, which contained a porcine growth hormone (pGH) cDNA at the first cistron and a Neo(r) gene at the second cistron was used to infect primary rat embryo fibroblast (REF) cells. The infected cells (5 x 10(6) cells/rat) were injected directly into the peritoneum of syngeneic hypophysectomized rats. We demonstrate that the implanted PSN-infected REF cells could secrete biologically active pGH in vivo, leading to significant growth of the tibia at day 15 and day 57 post-implantation. We also treated the PSN-infected REF cells with collagen to form a tissue-like structure. The skin-like discs were grafted underneath the skin on the back of rats and cells were retrieved at different times. Using two criteria, semiquantitative reverse transcription-polymerase chain reaction on the pGH RNA extracted from the explants and G418 resistance conferred from the explanted cells, we demonstrate that pGH was expressed in the implanted fibroblasts up to 70 days. Despite the fact that the total pGH RNA level was reduced in the explants of long-time post-implantation, which was probably due to the reduction of transduced cells retained in the explants, the specific efficiencies of pGH RNA expression from these explants were maintained as high as the primary PSN-infected REF prior implantation. These results suggest that fibroblast cells are capable of expressing the foreign genes persistently in vivo.

3T3 Cells↗
PubMed1995

Characterization of gamma-crystallin from a catfish: structural characterization of one major isoform with high methionine by cDNA sequencing.

gamma-Crystallin is the major and most abundant lens protein present in the eye lens of most teleostean fishes. To facilitate structural characterization of gamma-crystallins isolated from the lens of the catfishes (Clarias fuscus), a cDNA mixture was synthesized from the poly(A)+mRNA isolated from fresh eye lenses, and amplification by polymerase chain reaction (PCR) was adopted to obtain cDNAs encoding various gamma-crystallins. Plasmids of transformed E. coli strain JM109 containing amplified gamma-crystallin cDNAs were purified and prepared for nucleotide sequencing by the dideoxynucleotide chain-termination method. Sequencing more than five clones containing DNA inserts of 0.52 kb revealed the presence of one major isoform with a complete reading frame of 534 base pairs, covering a gamma-crystallin (gamma M1) with a deduced protein sequence of 177 amino acids excluding the initiating methionine. It was of interest to find that this crystallin of pI 9.1 contains a high-methionine content of 15.3% in contrast to those gamma-crystallins of low-methionine content from most mammalian lenses. Sequence comparisons of catfish gamma M1-crystallin with those published sequences of gamma-crystallins from carp, bovine and mouse lenses indicate that there is approx. an 82% sequence homology between the catfish and the carp species of piscine class whereas only 51-58% homology is found between mammals and the catfish. Moreover the differences in the hydropathy profiles for these two groups of gamma-crystallins, i.e. one with a high-methionine content from teleostean fishes and the other with a low-methionine content from mammalian species, reflect a distinct variance in the polarity distributions of surface amino acids in these crystallins.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1995

Double pylorus: report of a longitudinal follow-up in two refractory cases with underlying diseases.

Double pylorus is either a congenital abnormality or an acquired complication of peptic ulcer disease. We had followed two patients for 3 and 5 yr, respectively, to observe the processes of formation and the prognosis of double pylorus. Initially, duodenal ulcer was found in one patient with diabetes mellitus and chronic renal failure, and gastric ulcer was found in the other with chronic obstructive pulmonary disease. Both developed double pylorus with refractory courses. In spite of intensive medical treatment, both of them had persistent ulcers in the fistulous tract and failed to develop reepithelization. Helicobacter pylori was found in all of the specimens of gastroduodenal biopsies in both cases. Therefore, we believe that the refractory courses of double pylorus may be related to the underlying diseases and/or the presence of H. pylori. Antibacterial treatment of H. pylori or surgical intervention should be considered for patients with this condition.

Aged↗
PubMed1994

Molecular cloning, genomic organization, and chromosomal localization of an additional human aldehyde dehydrogenase gene, ALDH6.

Aldehyde dehydrogenase isozymes have been suggested to play a major role in the detoxification of aldehydes generated by alcohol metabolism and lipid peroxidation. We previously cloned and characterized four human nonallelic ALDH genes encoding different isozymes. The existence of an unique ALDH isozyme in human saliva and its polymorphism has been demonstrated previously. In this paper, we describe the cloning, characterization, and chromosomal mapping of an aldehyde dehydrogenase gene (ALDH6) expressed in the human salivary gland. The cloned ALDH6 cDNA is 3457 bp in length and contains an open reading frame encoding 512 amino acid residues. The deduced amino acid sequence showed that ALDH6 is larger than the human liver ALDH1 by 11 amino acid residues at the N-terminal, and the degree of identity between the two isozymes is 70% with an alignment of 500 amino acid residues. The human ALDH6 gene spans about 37 kb and consists of 13 exons. The putative TATA and CCAAT boxes and Sp1 binding sites are found in the 5' upstream region of the gene. Northern blot analysis demonstrated that the ALDH6 gene is expressed at low levels in many tissues and at higher levels in salivary gland, stomach, and kidney. The ALDH6 gene was assigned to chromosome 15q26 using fluorescence in situ hybridization.

Aldehyde Dehydrogenase↗
PubMed1994

Sequence analysis and expression of phospholipase A2 from Taiwan cobra.

Polymerase chain reaction (PCR) was employed to amplify cDNAs constructed from the poly(A)+RNA of venom glands in Taiwan cobras to facilitate the cloning and sequencing of phospholipase A2 (PLA2) gene. The PCR product was then subcloned into pUC18 vector and transformed in E. coli strain JM109. Plasmids purified from the positive clones were prepared for nucleotide sequencing by dideoxynucleotide chain-termination method. Sequencing several clones containing about 0.5 kb DNA inserts constructed a complete and unambiguous full-length reading frame of 468 base pairs covering a precursor for phospholipase A2 with a deduced mature protein sequence of 119 amino acids and a 27 amino-acid segment of signal peptide. The sequenced major PLA2 with pI 4.991 shows a high degree of sequence homology to those PLA2 of the same or closely-related genus. The deduced protein sequence allows us to correct and resolve some discrepancy between the sequences determined by conventional protein sequencing (Toxicon, 19, 141(1981)) and X-ray crystallography (Science, 250, 1560(1990)). Expression of PLA2 in E. coli vector generated a polypeptide which can cross-react with the antiserum against the native and purified PLA2 from the same cobra venom albeit with a much lower activity.

Amino Acid Sequence↗
PubMed1994

cDNA and protein sequences coding for the precursor of phospholipase A2 from Taiwan cobra, Naja naja atra.

The cDNA sequence encoding phospholipase A2 (PLA2) was determined by analysis of polymerase-chain-reaction (PCR) product amplified from total cDNA mixture which had been constructed from the poly(A)+RNA of venom glands obtained from Taiwan cobras. Two oligonucleotide segments corresponding to the 5'- and 3'-noncoding regions of sea-snake PLA2 gene were used as primers for PCR-amplified reaction. Plasmids of transformed E. coli strain JM109 containing amplified PLA2 cDNA were purified and prepared for nucleotide sequencing by dideoxynucleotide chain-termination method. Sequencing more than five clones containing about 0.5 kb DNA inserts revealed two isoforms with complete reading frames of 468 base pairs each covering a precursor for phospholipase A2 with a deduced mature protein sequence of 119 amino acids and a 27 amino-acid signal peptide. These two enzymes of Group I PLA2 differ in six nucleotide residues at the gene level and three amino acids along the whole polypeptide chain, each consisting of 14 cysteine residues similar to all reported PLA2 of different snake venoms. The signal peptides and hydropathy profiles of Group I PLA2 reported here are distinctly different from those of Group II PLA2 in viperid snakes.

Amino Acid Sequence↗
PubMed1994

Induction of 12-lipoxygenase expression by epidermal growth factor is mediated by protein kinase C in A431 cells.

Epidermal growth factor (EGF) increases 12-lipoxygenase mRNA by about 2-fold with a lag period of 4 to 8 hr, which precedes the increase in 12-lipoxygenase activity by 2 to 4 hr in human epidermoid carcinoma A431 cells. Induction of 12-lipoxygenase expression in human erythroleukemia cells by phorbol 12-myristate 13-acetate (PMA) has been reported previously. The present report describes a study of the involvement of protein kinase C (PKC) in EGF-induced 12-lipoxygenase expression in A431 cells. EGF-induced 12-lipoxygenase expression was inhibited by methyl 2,5-dihydroxycinnamate, a tyrosine kinase inhibitor. Staurosporine and calphostin C, which are two PKC inhibitors, inhibited EGF-induced enzyme activity and mRNA expression of 12-lipoxygenase. 1,2-Dioctanoyl-sn-glycerol (a membrane-permeant diacylglycerol) and PMA significantly induced enzyme activity and mRNA expression. Simultaneous treatment of cells with EGF and PMA did not exhibit an additive effect, suggesting that EGF and PMA share a common biochemical pathway in 12-lipoxygenase induction. Expression of mRNA for PKC alpha, delta and zeta was detected in A431 cells, whereas no mRNA expression for PKC beta 1, gamma and epsilon was observed. Taken together, these results suggest that EGF-induced 12-lipoxygenase expression is at least in part mediated by the PKC signal transduction pathway.

Alkaloids↗
PubMed1992

Plasmin and the regulation of tissue-type plasminogen activator biosynthesis in human endothelial cells.

Plasmin inhibited the biosynthesis of tissue-type plasminogen activator (tPA) antigen by human umbilical vein endothelial cells (HUVEC) in a dose-dependent manner. The amount of tPA antigen found in the 24-h conditioned medium of cells treated with 100 nM plasmin for 1 h was 20-30% of that in the control group. However, in contrast to tPA, such treatment led to a 3-fold increase in plasminogen activator inhibitor (PAI) activity, whereas the amount of PAI type 1 antigen was unchanged. The effects of plasmin on HUVEC were binding- and catalytic activity-dependent and were specifically blocked by epsilon-aminocaproic acid. Microplasmin, which has no kringle domains, was less effective in reducing tPA antigen biosynthesis or enhancing PAI activity in HUVEC. Kringle domains of plasmin affected neither tPA antigen nor PAI activity of the cells. Other proteases including chymotrypsin, trypsin, and collagenase at comparable concentrations did not have a significant effect on the biosynthesis of tPA antigen or PAI activity of HUVEC. Thrombin stimulated the biosynthesis of tPA and PAI-1 antigens by HUVEC. Thrombin also stimulated an increase in the protein kinase activity in HUVEC, whereas plasmin inhibited the protein kinase activity of the cells. It is possible that plasmin regulates the biosynthesis of tPA in HUVEC through the signal transduction pathway involving protein kinase.

Aminocaproic Acid↗
PubMed1992

Insulin-like growth factor I stimulates transcription of the c-jun proto-oncogene in Balb/C 3T3 cells.

Treatment of quiescent Balb/c 3T3 cells with insulin-like growth factor I (IGF I) resulted in the stimulation of proto-oncogene c-jun transcription. Cells exposed to cycloheximide and IGF I together showed super-induction of c-jun transcripts. Nuclear run-off assay revealed that IGF I up-regulated c-jun only while cycloheximide was present. The stability of c-jun mRNA was markedly increased in the cells treated with IGF I. These results suggest that IGF I controls the expression of c-jun by increasing the transcriptional activity and stabilizing the existing transcripts.

3T3 Cells↗
PubMed1992

Epidermal growth factor enhances a microsomal 12-lipoxygenase activity in A431 cells.

12-Hydroxyeicosatetraenoic acid (12-HETE) is formed from arachidonic acid either by 12-lipoxygenase or by a cytochrome P450 monooxygenase. 12-Lipoxygenase is generally localized in the soluble cytosolic fraction, and the cytochrome P450 monooxygenase is a microsomal enzyme. In this study, 12-HETE biosynthesis and the regulation of 12-HETE biosynthesis by epidermal growth factor (EGF) in A431 cells were investigated. 12-HETE was biosynthesized from arachidonic acid by the microsomal fraction of A431 cells, but not by the cytosolic fraction. The formation of 12-HETE was inhibited by 5,8,11,14-eicosatetraynoic acid, nordihydroguaiaretic acid, and caffeic acid. Nordihydroguaiaretic acid at 10(-4) M and 5,8,11,14-eicosatetraynoic acid at 10(-5) M almost completely inhibited its formation. However, the formation of 12-HETE was not affected by the presence of an NADPH-generating system, carbon monoxide, or SKF 525A. The biosynthetic 12-HETE was analyzed by chiral stationary phase high performance liquid chromatography and was highly enriched in (12S)-HETE. We therefore concluded that the enzyme responsible for the formation of (12S)-HETE in the microsomes of A431 cells is a 12-lipoxygenase. The microsomal 12-lipoxygenase of A431 cells belongs to the "leukocyte-type" enzyme as determined by substrate specificity and enzyme kinetics studies. The microsomal 12-lipoxygenase oxygenated linoleic acid much faster than the cytosolic platelet 12-lipoxygenase and is a "self-catalyzed inactivation" enzyme. Treatment of cells with 50 ng/ml EGF significantly induced microsomal 12-lipoxygenase activity. The lag period for the expression of the stimulatory effect of EGF on 12-lipoxygenase activity was approximately 10 h. The stimulatory effect of EGF on 12-lipoxygenase activity was completely blocked by treatment with 35 microM cycloheximide, indicating a requirement for de novo protein biosynthesis. Furthermore, the presence of the endogenous inhibitor of 12-lipoxygenase (which masked (12S)-HETE biosynthesis in intact cells) was identified in the cytosolic fraction of A431 cells. The putative inhibitor was enzyme-selective. It inhibited the leukocyte-type 12-lipoxygenase, but not the "platelet-type" enzyme.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1992

Human stomach aldehyde dehydrogenase cDNA and genomic cloning, primary structure, and expression in Escherichia coli.

An aldehyde dehydrogenase isozyme, ALDH3, which is strongly expressed in the stomach, may play a role in the oxidation of toxic aldehydes. Using reverse genetic approach, we cloned and characterized the cDNA and the gene for the ALDH3. The full length cDNA is 1624 base pairs (bp) in length and contains an open reading frame encoding 453 amino acid residues. The deduced amino acid sequence shows a high degree of resemblance to that of rat hepatocarcinoma ALDH. The human ALDH3 gene spans about 8 kb in length and consists of 10 exons. The putative TATA and CCAAT boxes are located in the consensus upstream distance from the transcription initiation site. Southern blot analysis of total genomic DNA argues against the proposed two-gene model for the ALDH3 isozymes (Yin, S.-J., Cheng, T.-C., Chang, C.-P., Chen, Y.-J., Chao, Y.-C., Tang, H.-S., Chang, T.-M., and Wu, C.-W. (1988) Biochem. Genet. 26, 343-360). Northern blot hybridization and analysis of PCR amplification products of cellular RNA demonstrated the existence of a high level of ALDH3 mRNA in human stomach and hepatoma cells, but a very low level in the normal liver. Expression of ALDH3 cDNA in Escherichia coli yielded a protein of 55 kDa, which exhibited kinetic properties similar to that found in ALDH3 isozyme purified from human stomach and liver, and was hybridizable with rabbit anti-human-hepatoma ALDH serum.

Aldehyde Dehydrogenase↗
PubMed1992

Effects of retinoids on endothelial cell proliferation, prostacyclin production and platelet aggregation.

In addition to their anti-inflammatory and anti-cancer activities, retinoids have been shown to affect angiogenesis, endothelial proliferation and the process of wound healing. While peripheral vascular occlusion has not been observed as an adverse effect clinically, the effects of retinoids on prostacyclin production in endothelial cells and platelet aggregation are not known. We examined the effects of tretinoin, isotretinoin and etretinate (3.3 x 10(-8) to 3.3 x 10(-5) M) on cytotoxicity by 51Cr-release assay, growth and prostacyclin in bovine carotid endothelial cell cultures, and the aggregation of human platelets induced by ADP. All retinoids showed either no or only small effects on cytotoxicity and human platelet aggregation. Prostacyclin production was not significantly affected except for tretinoin and isotretinoin at 3.3 x 10(-5) M. Endothelial proliferation was affected by all three retinoids in a dose-dependent fashion; for tretinoin and isotretinoin an inhibitory trend was noted as the concentration increased but the reverse was true for etretinate. Retinoids at 3.3 x 10(-5) M induced alterations of typical endothelial morphology; the cells became fibroblastoid. The results of prostacyclin production and platelet aggregation in the present study are consistent with the absence of peripheral vascular occlusion as a side effect clinically.

Animals↗
PubMed1992

Cytoprotective effect of reduced glutathione in hydrogen peroxide-induced endothelial cell injury.

The kinetic effects of hydrogen peroxide (H2O2) on cultured endothelial cells isolated from bovine carotid artery were studied. The cytoprotective effects of glutathione (GSH) on H2O2-induced cell injury were also investigated. H2O2-induced a dose- and time-dependent cell injury in cultured endothelial cells. H2O2-induced cell injury was blocked by simultaneous treatment by catalase, but not by superoxide dismutase. H2O2 also induced endogenous PGI2 biosynthesis, and the maximum PGI2 production was reached after 1 h treatment. Stimulation of PGI2 production was parallel with arachidonate release from H2O2-treated cells. However the prostaglandin biosynthesis enzyme activity in cells was inhibited by H2O2 treatment. When the cells were treated with GSH, the intracellular GSH reached a plateau after 3 h treatment. Both H2O2-induced cell injury and PGI2 production were significantly inhibited by the 3 h pretreatment with GSH. The cytoprotective effect of GSH was completely inhibited by buthionine sulfoximine which is a specific inhibitor of gamma-glutamylcysteine synthetase. The results indicate that the cytoprotective effect of GSH on H2O2-induced cell injury in cultured bovine carotid artery endothelial cells depends on the increase in intracellular GSH content.

Animals↗