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C Chang

Publications and source records attributed to C Chang.

At least 325 records · Page 18Linked to original sources
PubMed1993

A point mutation in the second zinc finger of the DNA-binding domain of the androgen receptor gene causes complete androgen insensitivity in two siblings with receptor-positive androgen resistance.

We have analyzed the nucleotide sequence of complementary and genomic DNAs of the human androgen receptor (AR) gene in two siblings (patients 9006 and 9030) with receptor-positive complete androgen insensitivity (Rec(+)-CAI). Northern analysis indicated that mRNA of the AR was normal in size. However, its expression was relatively reduced in both patients. Consistent with the normal androgen-binding capacity (496 and 552 fmol/mg DNA for patients 9006 and 9030, respectively) but decreased DNA-binding ability (168 fmol/mg DNA) measured in genital skin fibroblasts, no mutation was found in both N-terminal and ligand-binding domains of the AR. However, a single base substitution (G-->A) was found in the second zinc finger of the DNA-binding domain at nucleotide 2372 of the AR cDNA in both cases. This resulted in the replacement of a highly conserved arginine residue (amino acid 614) by a histidine. When the mutated receptor plasmid was cotransfected into PC-3 cells together with the reporter chloramphenicol acetyltransferase gene, chloramphenicol acetyltransferase activity was not induced by 5 alpha-dihydrotestosterone treatment, confirming that the mutation renders the AR nonfunctional and can, therefore, be held responsible for the clinical features in these patients. These results highlight the importance of Arginine-614 in the second zinc finger of the DNA-binding domain of the AR in the protein-DNA interaction.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗
PubMed1993

Proliferative and cytotoxic T-cell clones recognize endogenously synthesized HBcAg in an asymptomatic HBsAg carrier.

The characterization of immune responses to hepatitis B virus is crucial for the understanding of hepatitis B virus-caused liver disease. However, lack of a suitable autologous effector-target cell system makes a precise study of the pathogenesis of hepatitis B difficult. In this study we established a model system by using autologous HBcAg-expressing Epstein-Barr virus-immortalized lymphoblastoid cell lines as stimulator/target cells. T-cell cultures were established by repetitive stimulation with recombinant HBcAg or autologous HBcAg-expressing lymphoblastoid cell lines. Both proliferative and cytotoxic T-cell clones were obtained from the peripheral blood of an asymptomatic HBsAg carrier. Clones T12 (CD8+) and T2B (CD4+) were cytotoxic clones specific against autologous lymphoblastoid cell lines expressing endogenously synthesized HBcAg, whereas five CD4+ T-cell clones proliferated in response to lymphoblastoid cell lines incubated with exogenous recombinant HBcAg and autologous HBcAg-expressing lymphoblastoid cell lines. These results indicate that autologous HBcAg-expressing lymphoblastoid cell lines are appropriate stimulator/target cells for the study of HBcAg-specific T lymphocytes. By using this approach, we have demonstrated that both proliferative and cytotoxic T lymphocytes recognizing endogenously synthesized HBcAg are induced during chronic hepatitis B virus infection.

Carrier State↗
PubMed1992

A point mutation in the MyoD basic domain imparts c-Myc-like properties.

MyoD and c-Myc, members of the large "basic-helix-loop-helix" family of proteins, regulate diverse aspects of both normal and neoplastic growth and specific gene regulation. These two proteins differ at 9 of the 14 amino acids that comprise the basic domains necessary for DNA binding and transcriptional control. Individual amino acids in the MyoD basic domain were mutated to those found at the analogous positions in c-Myc. Four classes of mutants were obtained: (i) those with no effects on MyoD-site binding or activation of MyoD-responsive genes, (ii) those with no effect on MyoD-site binding but with a loss of activation potential, (iii) those with a loss of both DNA binding and activation potential, and (iv) one mutant (mut 9, Leu122----Arg) that left MyoD-site binding unaffected but imparted a new c-Myc-site binding capability. mut 9 competed with wild-type protein for the activation of MyoD-responsive reporter genes but could, like c-Myc, also suppress the adenovirus major-late promoter, which contains a c-Myc binding site. Our studies thus identify specific amino acid residues in the MyoD basic domain that are important for its activity as a DNA-binding transcriptional activator. Most significantly, our results with mut 9 indicate that Leu122 of MyoD is a critical determinant of specific DNA binding and that mutation at this residue can alter this specificity.

Amino Acid Sequence↗
PubMed1992

Expression of cytoplasmic CD3 epsilon proteins in activated human adult natural killer (NK) cells and CD3 gamma, delta, epsilon complexes in fetal NK cells. Implications for the relationship of NK and T lymphocytes.

NK cells have been defined as CD3-, CD16+, and/or CD56+ lymphocytes that mediate MHC-unrestricted cytotoxicity against certain tumors and virus-infected cells. Although CD3 epsilon transcripts have been detected in some NK clones, it has generally been thought that NK cells do not express CD3 proteins other than zeta which is associated with CD16 (Fc gamma RIII). We demonstrate that adult peripheral blood NK cell lines and clones express cytoplasmic CD3 epsilon proteins, but not CD3 delta or gamma. CD3 epsilon proteins were detected by immunoprecipitation, Western blot analysis, and immunofluorescence using antiserum directed against the cytoplasmic domain of CD3 epsilon. Although resting, adult peripheral blood NK cells have essentially undetectable levels of CD3 epsilon protein, expression was increased substantially after activation. In contrast to adult NK cells, NK cell clones established from human fetal liver express CD3 gamma, delta, and epsilon protein subunits that associate and form CD3 epsilon, gamma and CD3 epsilon, delta complexes in the cytoplasm, but are apparently unable to be transported to the cell surface. These results indicate that expression of CD3 gamma delta epsilon subunits is not restricted to T lymphocytes and supports the possibility that NK and T cells may be derived from common origins.

Age Factors↗
PubMed1992

Characterization of human androgen receptor overexpressed in the baculovirus system.

An essential step in the process of understanding the structure and function of the human androgen receptor (hAR) involves the production of large quantities of the hAR. For this purpose, the full-length hAR has been overproduced in insect cells by using a baculovirus genetic expression system. The recombinant hAR is produced in Sf21 insect cells at approximately 7 pmol/mg of protein (1 x 10(6) AR molecules per cell), which is 70-150 times greater than levels detected in androgen target tissues. Androgen can bind to the baculovirus-expressed hAR with high affinity (Kd = 0.46 nM), and the specificity of hormone binding in baculovirus-expressed hAR is essentially identical to that of bona fide hAR. An anti-AR monoclonal antibody can recognize the baculovirus-expressed hAR at approximately 100 kDa upon Western blot analysis. Sucrose gradient analysis shows that baculovirus-expressed hAR complexes sediment at 4 S in a high salt medium and these complexes can interact with anti-AR monoclonal antibody to form complexes that sediment at 8-10 S. Therefore, production of recombinant hAR from the baculovirus expression system will provide an alternative source of biologically active hAR for studies on the molecular mechanisms of androgen action.

Androgens↗
PubMed1992

Cloning and nucleotide sequence of a full-length cDNA for human liver gamma-glutamylcysteine synthetase.

We have cloned and sequenced a full-length cDNA for human liver gamma-glutamylcysteine synthetase (GCS), the rate-limiting enzyme in glutathione biosynthesis. The cDNA consists of 2634 bp containing an open reading frame encoding a protein of 367 amino acids and having a calculated M(r) = 72,773. The nucleotide sequence of the cDNA for human liver GCS shares an 84% overall similarity with the composite rat GCS sequence deduced from three overlapping partial cDNAs (Yan and Meister, JBC 265: 1588-1593, 1990). The deduced amino acid sequences are 94% similar. Comparison of Northern blots of total RNA isolated from rat kidney or liver with that from human kidney revealed the GCS mRNA to be larger in the human tissue (approximately 4.0 kb vs. approximately 3.7 kb). (The sequence for the human liver GCS cDNA has been assigned accession number M90656 in GenBank/EMBL databases.

Amino Acid Sequence↗
PubMed1992

High-resolution MR imaging of water diffusion in the rabbit lens.

Movement of water in the lens, specifically water diffusion, was examined with diffusion imaging and H217O-enhanced proton imaging. Diffusion imaging showed restricted water self-diffusion in the cortex which was not concurrent with the change in proton density; nevertheless it was dependent on the gradient direction and might be related to lens fiber/protein organization. The difference in water diffusion patterns between control and galactosemic cataract was subtle but distinguishable. On the other hand, intracameral H217O injection allowed visualization of water movement across the lens. In galactosemic cataracts, water diffused from the anterior cortex towards posterior cortex and into the vitreous. This pattern was not seen in the normal lens.

Animals↗
PubMed1992

Magnetic resonance microscopy of rabbit eyes.

Magnetic resonance (MR) micro-imaging was performed on enucleated eyes from rabbits previously injected with perfluoropropane gas (C3F8), with or without the surgical creation of retinal detachment. Condensed vitreous, which exhibited shortened longitudinal relaxation time (T1), could be differentiated with proton-density and T1-weighted imaging. Gradient-echo imaging could in addition detect vitreo-retinal tractions. The detached retina itself was also seen. Further, proton-density but not T1-weighted imaging showed lens opacities appearing as high-intensity regions. MR microscopy is a convenient method for gross morphological examination of intact eyes.

Animals↗
PubMed1992

An estrogen receptor positive MCF-7 clone that is resistant to antiestrogens and estradiol.

The antiestrogen tamoxifen has been successfully used to control estrogen receptor (ER) and progesterone receptor positive breast cancer. However, the development of antiestrogen resistance is frequently observed in patients following long term treatment. We have studied the development of antiestrogen resistance in vitro and established an antiestrogen resistant variant of MCF-7 cells (clone 5C) after long term culture in estrogen free medium. The growth of clone 5C cells was not altered by either estradiol-17 beta or the antiestrogens 4-hydroxytamoxifen and ICI 164,384. Estrogen-stimulated progesterone receptor and reporter gene expression were markedly reduced in 5C cells compared to wild type MCF-7 cells. Only minor alteration in the levels of ER and no alteration in the affinity of ER for ligand were found in 5C cells. No mutation of ER cDNA in 5C cells was detected by polymerase chain reaction and DNA sequencing. This study demonstrates that change(s) in ER-mediated gene expression rather than the amino acid sequence of the ER itself may be associated with the development of at least one form of antiestrogen resistance.

Breast Neoplasms↗
PubMed1992

Nuclear localization of androgen receptor in heterogeneous samples of normal, hyperplastic and neoplastic human prostate.

To facilitate an understanding of how androgens participate in the genesis of human benign hyperplasia and carcinoma we assayed androgen receptor in the epithelium and stroma of human prostatic tissue from 57 patients. Immunohistochemical staining of human androgen receptor was performed on 106 sections of normal prostate, benign prostatic hyperplasia (BPH) and prostate cancer. To determine variability of androgen receptor staining sections taken from different portions of the gland were studied. Frozen tissue sections were incubated with monoclonal antiandrogen receptor antibodies and staining was completed by the indirect avidin-biotin peroxidase method. Antibody staining was found mainly in the nucleus of prostatic epithelial cells, although some stromal cells also showed positive staining. Unlike normal prostate, there was a heterogeneous distribution of androgen receptor in BPH and prostate cancer. The androgen receptor content in well differentiated adenocarcinoma epithelium was significantly higher compared to moderately (p less than 0.05) and poorly (p less than 0.05) differentiated adenocarcinoma. Regardless of the origin of stromal tissue, some staining was observed. In each specimen studied the androgen receptor staining was consistent qualitatively and quantitatively for each pathological component throughout the specimen. These data confirm that androgen receptor is a nuclear receptor protein. Furthermore, they show the ability of monoclonal antibodies to reveal cellular/subcellular distribution of androgen receptor, and demonstrate a correlation between the degree of tumor differentiation and androgen receptor content in epithelial but not in stromal cells. These observations may have important implications for understanding the variable tumor response to hormone therapy.

Adenocarcinoma↗
PubMed1992

Prolongation of survival in metastatic melanoma after active specific immunotherapy with a new polyvalent melanoma vaccine.

A new polyvalent melanoma cell vaccine (MCV) was administered to 136 stage IIIA and IV (American Joint Committee on Cancer) melanoma patients. Induction of cell-mediated and humoral immune responses to common melanoma-associated antigens present on autologous melanoma cells was observed in patients receiving the new MCV. This was accompanied by increased activation of tumor-infiltrating lymphocytes. Survival correlated significantly with delayed cutaneous hypersensitivity (p = 0.0066) and antibody responses to MCV (p = 0.0117). Of 40 patients with evaluable disease, nine (23%) had regressions (three complete). From our historical database of 126 stage IIIA and 1275 stage IV melanoma patients, there were no significant changes in the natural history of metastatic melanoma during the past 20 years. Univariate and multivariate analyses demonstrated prognostic significance for site of metastases (p = 0.0001) and immunotherapy with the new MCV (p = 0.0001). Overall our new MCV increased the median and 5-year survival of stage IIIA melanoma patients with regional soft tissue metastases twofold (p = 0.00024), and stage IV patients threefold (p = 0.0001) compared with previous immunotherapy and other treatments.

Adult↗
PubMed1992

The TMK1 gene from Arabidopsis codes for a protein with structural and biochemical characteristics of a receptor protein kinase.

Genomic and cDNA clones that code for a protein with structural and biochemical properties similar to the receptor protein kinases from animals were obtained from Arabidopsis. Structural features of the predicted polypeptide include an amino-terminal membrane targeting signal sequence, a region containing blocks of leucine-rich repeat elements, a single putative membrane spanning domain, and a characteristic serine/threonine-specific protein kinase domain. The gene coding for this receptor-like transmembrane kinase was designated TMK1. Portions of the TMK1 gene were expressed in Escherichia coli, and antibodies were raised against the recombinant polypeptides. These antibodies immunodecorated a 120-kD polypeptide present in crude extracts and membrane preparations. The immunodetectable band was present in extracts from leaf, stem, root, and floral tissues. The kinase domain of TMK1 was expressed as a fusion protein in E. coli, and the purified fusion protein was found capable of autophosphorylation on serine and threonine residues. The possible role of the TMK1 gene product in transmembrane signaling is discussed.

Amino Acid Sequence↗
PubMed1992

Global changes in gene expression related to antibiotic synthesis in Streptomyces hygroscopicus.

Two-dimensional gel electrophoresis was used to follow changes in gene expression associated with antibiotic (bialaphos) biosynthesis in Streptomyces hygroscopicus. Cultures were pulse-labelled with [35S]-methionine before, during, and after the switch from primary to secondary metabolism in order to compare kinetic profiles of bialaphos (antibiotic) production (bap) genes during this metabolic transition. Separation of gene products on two-dimensional gels revealed that 27 were dependent on brpA for optimal expression and were activated as the culture approached stationary phase. Genes which encoded 10 brpA-dependent proteins were mapped to a 10 kb SstI fragment of the 35 kb bap gene cluster by expressing them in Streptomyces lividans using the thiostrepton-inducible tipA promoter. N-terminal amino acid sequences of two brpA-dependent proteins, obtained by direct microsequencing of protein spots excised from two-dimensional gels, identified them as gene products mapping to the same region and involved in secondary metabolic conversions of the bap pathway. The kinetics of synthesis of 16 brpA-dependent gene products were characterized using QUEST computer software. Cluster analysis performed on the kinetics of synthesis of 346 of the most highly expressed gene products of HP5-29, including 16 brpA-dependent ones, identified 75 families having distinct patterns of expression. Many brpA-dependent proteins were clustered together; 10 were found in one kinetic family. These kinetic families also included brpA-independent gene products perhaps subject to similar regulatory mechanisms and thus possibly involved in bialaphos biosynthesis. The activation/derepression of bap expression took place as cultures approached stationary phase and was temporally related to synthesis of ppGpp.

Amino Acid Sequence↗
PubMed1992

An epidemiological study of temporomandibular disorders in university students of Taiwan.

A total of 2033 university students were surveyed for the prevalence of TMD in Taiwan. It was found that 42.9% of the students had one or more signs related to TMD, and females were slightly more frequently found. Clicking of the joint was the most common sign. Occlusal factors such as balancing contact, dental restoration, molar guidance in eccentric movement, etc., were more often found in the TMD group. Scores on stress, general anxiety, emotion and anger were higher in the TMD group while tension, ambition, depression, competition and test anxiety were not as significant. Although the prevalence of TMD in this population is high, the severity and treatment demands are low.

Adolescent↗
PubMed1992

Properties of a 72-kilodalton mosquitocidal protein from Bacillus thuringiensis subsp. morrisoni PG-14 expressed in B. thuringiensis subsp. kurstaki by using the shuttle vector pHT3101.

The mosquitocidal properties of Bacillus thuringiensis subsp. israelensis and B. thuringiensis subsp. morrisoni PG-14 are attributable to protein inclusions grouped together within a parasporal body. In both of these strains, the mosquitocidal activity resides in proteins with molecular masses of 27, 72, 128, and 135 kDa. In an attempt to determine the toxicity of each protein, the shuttle vector pHT3101 was used to express the cryIVD gene (encoding the 72-kDa CryIVD protein) from B. thuringiensis subsp. morrisoni in an acrystalliferous mutant of B. thuringiensis subsp. kurstaki. With this system, parasporal inclusions of the 72-kDa protein were obtained that were comparable in size, shape, and toxicity to those produced by parental B. thuringiensis subsp. morrisoni. The inclusions were bar shaped, measured 500 by 300 by 150 nm, and were easily visible with phase-contrast microscopy by 16 h of cell growth. A 50% lethal concentration of 64 ng/ml for these inclusions was determined in bioassays against fourth instars of Culex quinquefasciatus, which was similar to the 50% lethal concentration of 55 ng/ml obtained for the 72-kDa inclusion from B. thuringiensis subsp. israelensis. In contrast, expression of the cryIVD gene in Escherichia coli was very low and only detectable by immunoblot analysis. These results demonstrate that the pHT3101-B. thuringiensis expression system can be used to express the CryIVD protein in quantities and with properties comparable to that obtained with the natural host. This system may prove useful for the expression of other B. thuringiensis proteins and, in particular, for reconstitution experiments with inclusions produced by the mosquitocidal subspecies of B. thuringiensis.

Animals↗
PubMed1992

Characterization of the rate-dependent effects of ethmozine on conduction, in vivo, and on the sodium current, in vitro, in the newborn heart.

We evaluated the effects of ethmozine on resting conduction intervals, myocardial refractory periods and His-Purkinje and intraventricular conduction in vivo in 11 neonatal dogs aged 7 to 16 days. Ethmozine produced significant prolongation in resting sinus cycle length (P < .05), in the atrioventricular nodal (P < .001) and His-Purkinje conduction time (P < .01) intervals and in the QRS duration (P < .01). Ventricular, but not atrial refractory periods were significantly prolonged (P < .05). Rate-dependent changes in His-Purkinje and intraventricular conduction were demonstrated after ethmozine by direct pacing of the bundle of His and right ventricular pacing. The development of steady-state conduction delay at a paced cycle length of 200 msec was characterized by a time constant (tau on) of 23.5 beats. The time constant of diastolic recovery (tau off) from rate-dependent conduction delay, determined during His bundle extrastimulation, was 171 msec. Ethmozine was highly proarrhythmic. A total of 7 arrhythmias were induced in 6 out of 12 neonates after administration of ethmozine. We also characterized ethmozine block of cardiac sodium channels in isolated neonatal canine ventricular myocytes using the whole cell variation of the patch clamp technique. Ethmozine (1.3-40 microM) produced a use-dependent block of cardiac Na channels that was dependent upon both drug concentration and pulse duration. Drug binding to inactivated channel states accounted for the observed use-dependent block. The time constant of recovery from use-dependent block ranged between 10 and 30 sec at 16 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗