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Biomedical subjects

Q Huang

Publications and source records attributed to Q Huang.

At least 271 records · Page 15Linked to original sources
PubMed1994

Association of peroxisome proliferator-activated receptor and Hsp72.

In an effort to understand the relationship between a 72-kDa heat shock protein (Hsp72) and peroxisome proliferator-activated receptors (PPARs), we have characterized their interaction using clofibric acid-Sepharose chromatography and co-immunoprecipitation with antisera raised against either rat PPAR (rPPAR) or Hsp72. First, we observed that both rPPAR and Hsp72 elute in a clofibrate-dependent manner from the clofibric acid-Sepharose matrix. Second, we found that immunoprecipitation of either protein from solution resulted in the precipitation of the other. This result was obtained from rat liver cytosol, from Spodoptera frugiperda (Sf9) insect cells expressing rPPAR, and from reticulocyte lysate rPPAR expression systems. These results suggest that Hsp72 and rPPAR form a complex in vivo and that Hsp72 may play a role in the folding, subcellular localization, and/or signaling pathway of PPARs.

Animals↗
PubMed1994

Retrotrapezoid nucleus muscarinic receptor subtypes localized by autoradiography.

Microinjection of muscarinic receptor subtype antagonists into the region of the cat retrotrapezoid nucleus (RTN) decreases blood pressure (greatest efficacy; M2 subtype) and both baseline phrenic activity and CO2 sensitivity (greatest efficacy; M3/M1 subtype). Here we examine, in cat medullary sections at the level of the RTN, the effects of the same antagonists on binding of the high affinity muscarinic agonist quinuclidinyl benzilate (QNB). 3H-QNB binding was saturated and highly specific at 1 nM concentration and stable over 30 to 120 min (Kd, 0.49 nM; Bmax, 136 fm/mg protein). Studied biochemically, we found IC50 values for whole sections of 4.9 x 10(-6) M (M1 antagonist pirenzepine); 1.0 x 10(-6) M (M2 antagonist AFDX); and 0.64 x 10(-7) M (M3 antagonist DAMP; P < 0.03 vs PZ). Densitometric analysis of whole medullary cross section autoradiograms resulted in similar IC50 values as in the biochemical approach. Specific analysis of the RTN region demonstrated the presence of 3H-QNB binding and similar competition by the antagonists. Average IC50 values determined by densitometry were 14 x 10(-6) M (pirenzepine); 1.3 x 10(-6) M (AFDX; P < 0.01 vs PZ); and 0.53 x 10(-7) M (DAMP; P < 0.01 vs PZ). All three subtypes of muscarinic receptors identifiable via pharmacological antagonists appear to be present in the RTN region but we could not distinguish a subtype-specific pattern of receptor distribution.

Adamantane↗
PubMed1994

Studies on an artificial trypsin inhibitor peptide derived from the mung bean trypsin inhibitor: chemical synthesis, refolding, and crystallographic analysis of its complex with trypsin.

The active fragment with Lys at the reactive site of mung bean trypsin inhibitor (MBILF) is composed of two peptide chains, A1 of 26 residues and A2 of 9 residues linked via two disulfide bonds. In the present study, a peptide of 22 residue comprising the sequence of chain A1 from position 3 to 24 was synthesized by the solid-phase method. This synthetic peptide with six Cys residues contains a reactive site at position Lys11I-Ser12I (I denotes an inhibitor residue). Air oxidation and HPLC purification resulted in two antitrypsin active components, SPC1 and SPC2. Neither SPC1 nor SPC2 can stoichiometrically inhibit trypsin. The Ki values of SPC1 and SPC2 are 1.2 x 10(-7) and 4.0 x 10(-8) M, respectively. The complexes of SPC1 and SPC2 with bovine beta-trypsin (BTRY) were crystallized by ammonium sulphate precipitation at pH 6.4 and 6.0, respectively. The two crystals have the same crystal form with space group P2(1)2(1)2(1) and cell dimension of a = 63.2(2) A, b = 63.5(6) A, and c = 69.8(4) A. The crystal structure of one complex, SPC1-BTRY, was determined and refined at 2.2 A resolution to a final R-value of 19.2%. From the resulting electron density map, 9 residues of SPC1, from position 9I to 17I, were identified clearly and three-dimension atomic model of the 9-residue reactive loop formed by a disulfide bridge, Cys9I-Cys17I, was built. No electron density corresponding to the other 13 residues was observed in the present map.(ABSTRACT TRUNCATED AT 250 WORDS)

Binding Sites↗
PubMed1994

Infectious transcripts from PCR-amplified broad bean mottle bromovirus cDNA clones and variable nature of leader regions in RNA 3 segment.

The genome of broad bean mottle bromovirus (BBMV) contains three positive-sense ssRNA segments, each capped with m7GpppA. Full-length transcribable cDNA clones for four strains of BBMV were constructed by employing reverse transcriptase-PCR (RT-PCR) and a high fidelity Vent DNA polymerase. The transcribed BBMV RNAs contained a 5' non-viral G residue and, although delayed, produced symptoms similar to those observed in plants infected with authentic virion RNAs. The transcripts replicated inefficiently in protoplasts. In contrast, transcript-derived progeny BBMV RNAs had the repaired termini, were as infectious as the authentic BBMV RNAs and replicated to high levels in protoplasts. In vitro translation of synthetic RNAs confirmed the previously proposed gene expression strategy for BBMV. Sequencing of virion RNAs from the Bawden strain revealed two forms of BBMV RNA3 components, the longer form containing 21 5' extra nucleotides derived by the duplication of two short 5' leader regions. The relative concentration of the two RNA 3 forms was found to be host-dependent, with the longer form prevailing in broad bean and Nictiana clevelandii infections and the shorter form in bean infections.

Base Sequence↗
PubMed1994

[Study on soluble proteins in human fetal lens].

Soluble proteins from corter of human fetal clear lenses were separated by column chromatography with Sepharyl S-300 (superfine) and determined by disc-PAGE and SDS-PAGE. The results documented from 18W, 21W (2 cases) and 24W fetal lenses suggested that a negative correlation tended to be presented between the age of fetus and fraction of alpha, beta 1 crystallins (r = -0.9542; r = -0.9674) and a positive correlation between the age of fetus and fraction of beta 2, gamma crystallins (r = 0.9666; r = 0.9970). There was no change in the alpha, beta 1, beta 2 crystallins on disc-PAGE. Among the fetuses of different ages, the blurred band of gamma crystallin seen in 18W and 21W fetal lenses was thickened in the 24W fetal lens. The SDS-PAGE showed that there was no significant difference among the polypeptides in each of alpha, beta 1, beta 2, gamma crystallins in the studied fetal lenses of different age groups. It is presumed that the normal lens is proportionally composed of four fractions of crystallin. This is hypothetically contributed to the maintenance of normal structure of the lens, why the proper proportion of four fractions of crystallins is depended either on proteins synthesis or on protein molecular assembly is not yet clear.

Chromatography, Gel↗
PubMed1993

Cloning of a new member of the peroxisome proliferator-activated receptor gene family from mouse liver.

Peroxisome proliferators are postulated to elicit predictable pleiotropic responses in the liver by activating a peroxisome proliferator-activated receptor (PPAR). PPARs from mouse liver (mPPAR), rat liver (rPPAR), and Xenopus liver (xPPAR gamma) have been cloned recently. We now report the cloning of a new member from mouse liver which we designate mPPAR gamma. mPPAR gamma cDNA contained an open reading frame encoding a 475-amino acid protein exhibiting 75% amino acid similarity to xPPAR gamma, while it showed only 55% identity with mPPAR. The ligand-binding and DNA-binding domains are best conserved between mPPAR gamma, mPPAR, and xPPAR gamma. Like rPPAR, mPPAR gamma is able to impart peroxisome proliferator responsiveness to the promoter of peroxisomal bifunctional gene, which encodes the second enzyme of the peroxisomal fatty acid beta-oxidation system. Northern blot analysis revealed high expression of mPPAR gamma gene in mouse liver, kidney, and heart and low expression in the lung, testis, brain, skeletal muscle, and spleen. In mice treated with ciprofibrate, a peroxisome proliferator, a 2-fold increase in mPPAR gamma mRNA was observed in the liver and kidney. The presence of two PPARs in the mouse liver suggests the possibility of multiple signaling pathways for the peroxisome proliferator-induced pleiotropic responses.

Amino Acid Sequence↗
PubMed1993

Hydroxylation and dealkylation of methyl-n-butylnitrosamine and role of certain cytochrome P-450 isozymes in these reactions.

We studied the metabolism of methyl-n-butyl-nitrosamine (MBN), a carcinogen for the rat esophagus and liver. The 2-, 3- and 4-hydroxy derivatives were identified as new metabolites of MBN. In studies on tissue slices freshly removed from MRC-Wistar rats, MBN metabolism resembled that of the previously studied methyl-amylnitrosamine in the esophagus catalyzed 2- and 3- hydroxylation; liver, omega-1 hydroxylation; and lung, omega-hydroxylation of both nitrosamines. Liver microsomes from Sprague-Dawley rats catalyzed 2-, 3- and 4-hydroxylation of MBN, as well as the previously studied activating reactions of demethylation and debutylation. Phenobarbital induced all five reactions of MBN bh rat liver microsomes, especially debutylation; 3-methylhol-anthrene induced 3-hydroxylation and debutylation and isoniazid induced demethylation and debutylation. Monoclonal antibodies that inhibit specific cytochrome P-450 isozymes were used to identify the isozymes involved in each reaction. Antibody 4-7-1 appeared more specific than the previously used antibody 2-66-3 for inhibiting P-450 2B1 and/or 2B2. For the metabolism of both MBN and methylamylnitrosamine by rat liver microsomes, the antibody results indicated that P-450 2C11 mainly catalyzed demethylation and omega-1 hydroxylation, P-450 1A1 or 1A2 catalyzed 3-hydroxylation and debutylation or depentylation, P-450 2E1 produced demethylation and P-450 2B1 or 2B2 produced omega-1 hydroxylation, demethylation and debutylation or depentylation.

Animals↗
PubMed1993

Conversion of lytic to persistent alphavirus infection by the bcl-2 cellular oncogene.

Little is known about virus-host cell interactions that regulate the lytic potential of viruses during productive replication. Sindbis virus (SV), a single-stranded positive-sense RNA virus in the alphavirus genus (family Togaviridae), results in lytic infection in most vertebrate cell lines, but persistent productive infection in post-mitotic neurons. The cellular oncogene bcl-2, which encodes an inner mitochondrial membrane protein of M(r) 26,000 (ref. 2), blocks programmed cell death (apoptosis) in neurons. We therefore investigated whether SV infection induces programmed cell death in non-neuronal cells, and if so, whether virus-induced programmed cell death can be blocked by transfection with bcl-2. We demonstrate that SV infection of baby hamster kidney (BHK-2), mouse neuroblastoma (N18), and rat prostatic adenocarcinoma (AT-3) cells results in programmed cell death, whereas SV infection of bcl-2-transfected AT-3 cells results in long-term persistent productive infection. Thus cellular bcl-2 oncogene expression plays a role in the establishment of persistent viral infection by blocking virus-induced programmed cell death.

Adenocarcinoma↗
PubMed1993

Refined 1.6 A resolution crystal structure of the complex formed between porcine beta-trypsin and MCTI-A, a trypsin inhibitor of the squash family. Detailed comparison with bovine beta-trypsin and its complex.

The crystal structure of the complex formed by porcine beta-trypsin with the MCTI-A inhibitor (Momordica charantia, Linn. Cucurbitaceae) has been determined at 1.6 A resolution using the molecular replacement method. The sequence of MCTI-A was determined by recognizing the electron density, and shows that MCTI-A is a member of the squash family of trypsin inhibitors. We report the first high-resolution structure of porcine beta-trypsin. Detailed comparisons have been made on the overall structure, solvent structure and active-site geometries between this complex and bovine beta-trypsin and its complexes. On the basis of our results, we discuss the interaction patterns between inhibitor and trypsin. Unlike other complex structures formed by bovine trypsin with inhibitors, no out-of-plane distortion around the inhibitor's scissible peptide was observed. The role of the trypsin catalytic triad is also discussed on the basis of this structure.

Animals↗
PubMed1993

Characterization of defective interfering RNA components that increase symptom severity of broad bean mottle virus infections.

Several strains of the broad bean mottle virus (BBMV), an icosahedral tripartite plant RNA virus, which show distinct reactions on certain plant hosts have been described (K. M. Makkouk et al., Neth. J. Plant Pathol. 94, 195-212, 1988). Here we report defective interfering (DI) RNAs encapsidated in two BBMV strains from Morocco and Tunisia. While not effective in some plants, these DI RNAs exacerbated the severity of symptoms in others. The most dramatic, lethal effect of DI RNAs has been found on pea (Pisum sativum, cv. Rondo) seedlings. Sequence analysis has revealed that the DI RNAs were derived by single in-frame central deletions of 448 to 537 nt in the corresponding genomic RNA2 components. A comparison of the intensities of full-length RNA2 bands from DI molecule-containing and DI molecule-deficient virion RNA preparations revealed that the DI RNAs decreased the level of RNA2 components in total RNA preparations. The differences between corresponding virion RNAs were much smaller. This suggests an interference with RNA replication. In vitro assays and an analysis of the polyribosomal RNA fractions confirmed the translational activity of DI RNAs. This paper reports the first description of natural DI RNAs in tripartite isometric plant RNA viruses.

Base Sequence↗
PubMed1993

Rat mesangial cell hypertrophy in response to transforming growth factor-beta 1.

Central features of progressive glomerular sclerosis are initial glomerular hypertrophy and subsequent accumulation of extracellular matrix proteins. Since TGF-beta 1 may play a key role in this glomerular response to injury, the present study sought to explore further TGF-beta 1 actions and regulated expression of its receptor in rat mesangial cells. The rat TGF-beta type II receptor (TGF-beta RII) homolog was cloned by screening a rat kidney cDNA library with a human TGF-beta RII cDNA probe, and sequenced. Expression of this receptor subtype in rat mesangial cells was then demonstrated by RNase protection assay, and by Northern blot analysis of poly (A)+ RNA, TGF-beta RII expression was down-regulated in cells treated with exogenous TGF-beta 1. Affinity cross linking studies demonstrated presence of this receptor on cell surface. Rat mesangial cells also expressed TGF-beta 1 and autoinduction by TGF-beta 1 was observed in the same cells, suggesting that this polypeptide may act in an autocrine fashion on mesangial cells, and that it may stimulate a positive autoamplification loop. TGF-beta 1 inhibited mesangial cell proliferation and stimulated significant overall protein and collagen production. Furthermore, mesangial cell size increased in response to chronic TGF-beta 1 treatment. These findings demonstrate that rat mesangial cells express key components of the TGF-beta system and raise the intriguing possibility that in the glomerular mesangium, TGF-beta 1 may not only induce extracellular matrix synthesis, but may also participate in the process of glomerular hypertrophy in response to injury.

Amino Acid Sequence↗
PubMed1993

Metabolism of carcinogenic nitrosamines in the rat and human esophagus and induction of esophageal adenocarcinoma in rats.

The mechanism is discussed by which certain nitrosamines induce esophageal papillomas and squamous cancer in rats, and some evidence is presented for the view that nitrosamines also induce the same cancer in humans, especially in China and South Africa. Studies on the metabolism of nitrosamines by cytochrome P450 isozymes in rat and human esophagus, including the activation reactions of formaldehyde and pentaldehyde formation from methyl-n-amylnitrosamine (MNAN), are reviewed. These reactions are catalyzed by microsomes from the rat and human esophagus, probably because these microsomes contain specific cytochrome P450 isozymes. Evidence is reviewed for the occurrence of nitrosamines related to MNAN in fungus-infected corn. The incidence of esophageal adenocarcinoma is rising in Western countries. The precursor lesion, Barrett's esophagus, is associated with colon cancer, suggesting a role for bile salts in the induction of the esophageal tumor. Studies are described in which rats were subjected to esophago-duodenostomy (joining the duodenum to the esophagus) and then treated with nitrosamines that normally induce esophageal squamous cancer. Adenocarcinomas of the lower esophagus were induced as well as Barrett's esophagus (under one set of conditions). Feeding a high-fat diet with this system increased the incidence of esophageal adenocarcinoma. This tumor was not induced when the operation was changed to esophago-gastroplasty (widening the lower esophageal sphincter). These results support a role of reflux of duodenal contents (including bile and pancreatic juice) rather than of gastric contents in the etiology of human esophageal adenocarcinoma.

Adenocarcinoma↗
PubMed1993

Inhibition by phenylethyl and phenylhexyl isothiocyanate of metabolism of and DNA methylation by N-nitrosomethylamylamine in rats.

We investigated the effect of 2-phenylethyl and 6-phenylhexyl isothiocyanate (PEITC and PHITC) on the metabolism of the rat esophageal carcinogen, N-nitrosomethylamylamine (NMAA). PEITC was administered orally to MRC-Wistar rats as single doses of 0.1 or 1.0 mmol/kg, or by other regimens. When esophagi and liver slices from the treated rats were incubated with 23 microM NMAA, the formation of 2- to 5-hydroxy-NMAA was inhibited by 45-90% for esophagus and by 14-19% for liver slices. In contrast, when esophagi and liver slices from untreated MRC-Wistar rats were incubated in vitro with NMAA and 10 microM PEITC, the PEITC inhibited hydroxy-NMAA formation similarly (by 79-89%) in the two tissues. Also, PEITC inhibited the formation from NMAA of the hydroxy-NMAAs, formaldehyde and pentaldehyde by esophageal and liver microsomes to similar extents. In studies on DNA methylation by NMAA, 7- and O6-methylguanine (O6-MeG) were determined by HPLC with fluorimetric detection. Guanine methylation in esophageal and liver DNA was generally close to linear for doses of 5-50 mg NMAA/kg. With 50 mg NMAA/kg, guanine methylation in esophageal and liver DNA peaked after 5 h, and 8-11% of the peak O6-MeG persisted after 72 h. A single dose of 0.1 or 1.0 mmol PEITC/kg reduced the O6-MeG levels by 44-51% in the esophagus but by only 7-22% in the liver. Administration of the PEITC homolog, PHITC, inhibited NMAA metabolism by liver slices from the treated rats and the methylation of guanine in liver DNA, but had little effect in the esophagus, i.e. PHITC tended to have the opposite tissue specificity to PEITC. The finding that administration of PEITC specifically inhibited NMAA metabolism in the rat esophagus supports the view that PEITC may be a useful chemopreventive agent against esophageal carcinogenesis in humans.

Animals↗
PubMed1993

Extracellular matrix-stimulated phospholipase activation is mediated by beta 1-integrin.

beta 1-Integrins are major mediators of interactions between cells and extracellular matrix (ECM). Adhesion of rat glomerular epithelial cells (GEC) to collagen stimulated phospholipase C. As a result, 1,2-diacylglycerol (DAG) was increased, and inositol phospholipids were decreased in collagen-adherent cells, as compared with GEC adherent to plastic substrata. Adhesion to collagen also stimulated production of free arachidonic acid (the precursor for eicosanoids) due to metabolism of DAG through the DAG lipase pathway and due to phospholipase A2-induced hydrolysis of phospholipids. Phospholipase A2 appeared to be stimulated as a result of protein kinase C (PKC) activation, probably secondary to increased DAG. The collagen-induced increases in DAG and free arachidonic acid, as well as the decrease in inositol phospholipids, were partially inhibited by lowering extracellular Ca2+ concentration to 200 nM or less and by anti-beta 1-integrin antibody Fab. In contrast, anti-beta 1-integrin immunoglobulin G (IgG) enhanced collagen-mediated increases in DAG and arachidonic acid. Proliferation of GEC adherent to collagen was reduced in the presence of anti-beta 1-integrin IgG. The antiproliferative effect of anti-beta 1-IgG appeared to be mediated through PKC, since it was absent in PKC-depleted GEC. Immunoprecipitation with integrin subunit-specific antibodies demonstrated alpha 2 beta 1- and alpha 3 beta 1-integrins in GEC. Thus, in GEC, ECM induces activation of phospholipases C and A2, which is mediated, at least in part, by beta 1-integrins. Products of integrin-mediated phospholipase activation may modulate GEC proliferation.

Animals↗
PubMed1993

Cerebellar control of expiratory activities of medullary neurons and spinal nerves.

Expiratory-related activities of spinal nerves are augmented after stimulation of the infracerebellar nucleus of the cerebellum. These stimulations alter neither inspiratory spinal neural activities nor inspiratory and expiratory laryngeal neural activities. It was hypothesized that efferents from the infracerebellar nucleus impinge on spinal motoneurons by a pathway that bypasses the medulla. In decerebrate, paralyzed, and vagotomized cats, phrenic and expiratory triangularis sterni activities and activities of medullary respiratory neurons were recorded. During infracerebellar stimulation, activities increased for expiratory bulbospinal neurons and neurons with tonic discharge patterns. After unilateral ablation of the infracerebellar nucleus by kainic acid, triangularis sterni discharge was eliminated and activities of expiratory bulbospinal neurons continued at reduced frequencies. Stimulations of the anterior interposed nucleus caused both inspiratory and expiratory activities to increase, whereas no systematic changes followed stimulations of the vermis. Results establish that at least a portion of the changes in expiratory activities of spinal motoneurons after perturbations of the infracerebellar nucleus occurs secondarily to changes in activities of medullary neurons. The possibility of separate cerebellar projections to medullary and spinal neurons is discussed.

Animals↗