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Biomedical subjects

J C Shih

Publications and source records attributed to J C Shih.

At least 127 records · Page 7Linked to original sources
PubMed1990

The expression of human MAO-A and B genes.

Northern analysis using 32P-labeled subfragments of human liver MAO-A and B cDNA clones detected 5Kb and 3Kb transcripts, respectively in fetal tissues and adult brains. The tissue distribution of these transcripts was determined. Small intestine and placenta express, in addition to the MAO-A 5Kb transcript, a 2Kb transcript determined to lack a 3' flanking region. MAO-A appeared prior to MAO-B in the fetal brain, whereas both MAO-A and B were found in adult brain.

Blotting, Northern↗
PubMed1990

Effect of snake venom of Agkistrodon halys on atherosclerosis and blood characteristics in Japanese quail.

Extracts of snake venom have been widely used for the treatment of vascular thrombotic diseases, yet the therapeutic mechanism is not clear. The effect of snake venom fractions on atherosclerosis in Japanese quail was studied. The venom of Agkistrodon halys was fractionated by DEAE-cellulose chromatography and the pooled protein fractions that resulted were injected intravenously into the quail with aortic atherosclerosis induced by dietary cholesterol. After 7 weeks of injections on every other day, the quail were killed, blood clotting times and serum cholesterol levels were determined, and aortic atherosclerosis and fatty liver were scored. The results showed that while no regression of atherosclerosis was observed, the lowering of serum cholesterol, prolonged blood clotting time and reduced fatty liver were significantly affected by the injection of one of the pooled protein fractions. This venom fraction contained two major protein components, one of which had arginine esterase activity. From this study we conclude that snake venom has little effect on the regression of atherosclerosis, but it prolongs blood clotting and lowers serum cholesterol.

Animals↗
PubMed1990

Peptide displacement of [3H]5-hydroxytryptamine binding to bovine cortical membranes.

Chemical studies have demonstrated that peptides such as the encephalitogenic (EAE) peptide of myelin basic protein (MBP) and luteinizing hormone-releasing hormone (LHRH) can bind serotonin (5-hydroxytryptamine, 5-HT) in vitro. The present research was undertaken to determine whether such binding interferes with 5-HT binding to its 5-HT1 receptors on bovine cerebral cortical membranes. EAE peptide and LHRH displaced [3H]5-HT with IC50s of 4.0 x 10(-4) and 1.8 x 10(-3) M respectively. MBP itself also showed apparent displacing ability with an IC50 of 6.0 x 10(-5) M, though it also caused aggregation of cortical membranes that might have interfered with normal receptor binding. These results support previous suggestions that the tryptophan peptide region of MBP may act as a 5-HT receptor in the neural system. We also tested the effects of muramyl dipeptide (N-acetyl-muramyl-L-Ala-D-isoGln, MD), a bacterial cell-wall breakdown product that acts as a slow-wave sleep promoter, binds to LHRH and EAE peptide, and competes for 5-HT binding sites on macrophages. It showed no significant displacement of 5-HT binding to cortical membranes (IC50 greater than 10(-1) M), but its D-Ala analogue did (IC50 = 1.7 x 10(-3) M). Thus, it seems likely that the 5-HT-related effects of naturally occurring muramyl peptides are physiologically limited by receptor types.

Acetylmuramyl-Alanyl-Isoglutamine↗
PubMed1990

Tissue distribution of human monoamine oxidase A and B mRNA.

Monoamine oxidase (MAO) A and B play important roles in the metabolism of biogenic amines. Northern analysis using 32P-labeled subfragments of human liver MAO A and B cDNA clones detected a 5- and a 3-kb transcript, respectively, in most human tissues examined. However, fetal heart and thymus express minute amounts of MAO A transcript, whereas fetal brain, muscle, thymus, spleen, meninges, and placenta express minute amounts of MAO B transcript. Small intestine and placenta express, in addition to the MAO A 5-kb transcript, a 2-kb transcript, which may arise from an alternative polyadenylation site. MAO A and B transcripts are expressed in similar regions of adult human brain. The highest concentrations of these transcripts were located in frontal cortex and locus coeruleus. This study demonstrates the tissue-specific distribution of the MAO genes and will provide insight into the physiological functions of MAO A and B.

Brain↗
PubMed1990

Isolation, identification, and characterization of a feather-degrading bacterium.

A feather-degrading culture was enriched with isolates from a poultry waste digestor and adapted to grow with feathers as its primary source of carbon, sulfur, and energy. Subsequently, a feather-hydrolytic, endospore-forming, motile, rod-shaped bacterium was isolated from the feather-degrading culture. The organism was Gram stain variable and catalase positive and demonstrated facultative growth at thermophilic temperatures. The optimum rate of growth in nutrient broth occurred at 45 to 50 degrees C and at pH 7.5. Electron microscopy of the isolate showed internal crystals. The microorganism was identified as Bacillus licheniformis PWD-1. Growth on hammer-milled-feather medium of various substrate concentrations was determined by plate colony count. Maximum growth (approximately 10 cells per ml) at 50 degrees C occurred 5 days postinoculation on 1% feather substrate. Feather hydrolysis was evidenced as free amino acids produced in the medium. The most efficient conditions for feather fermentation occurred during the incubation of 1 part feathers to 2 parts B. licheniformis PWD-1 culture (10 cells per ml) for 6 days at 50 degrees C. These data indicate a potential biotechnique for degradation and utilization of feather keratin.

Journal Article↗
PubMed1989

Human monoamine oxidase A and B genes map to Xp 11.23 and are deleted in a patient with Norrie disease.

Monoamine oxidase A and B (MAO A and B) are the central enzymes that catalyze oxidative deamination of biogenic amines throughout the body. The regional locations of genes encoding MAO A and B on the X chromosome were determined by using full-length cDNA clones for human MAO A and B, respectively. Using somatic cell hybrids, in situ hybridization, and field-inversion gel electrophoresis as well as deletion mapping in a patient with Norrie disease, we concluded that these two genes are close to each other and to the DXS7 locus (Xp 11.3).

Animals↗
PubMed1989

Discovery of noninfectious viral genes complementary to Marek's disease herpes virus in quail susceptible to cholesterol-induced atherosclerosis.

Japanese quail genetically selected on the basis of atherosclerosis susceptibility were tested for infection by Marek's disease herpesvirus (MDV). Viral DNA was detected in the atherosclerotic aortas of susceptible (SUS) quail by the technique of DNA hybridization. Southern blot analysis demonstrated that restriction mapping of aortic DNA was specific and different from that of MDV. Screening of quail embryos by dot-blot hybridization detected that MDV DNA existed in 100% of SUS quail tested. Resistant (RES) quail were a mixed population, with 16% of embryos resembling the SUS group. Functional MDV was not found by a number of methods including virus isolation, serological test, and exposure of sentinel chicks to SUS quail. These results suggest that the SUS quail possess a portion of the MDV genome in the germline, and the viral genes have been coselected by their susceptibility to cholesterol-induced atherosclerosis.

Animals↗
PubMed1989

Expression of functional human monoamine oxidase A and B cDNAs in mammalian cells.

Monoamine oxidase (MAO) A and B are important enzymes that metabolize biogenic amines throughout the body. Previous studies had suggested that both MAO A and B consist of two subunits of molecular masses of 63 and 60 kilodaltons, respectively. The cDNAs encoding one subunit of human liver MAO A and B have been expressed in mammalian cells by transfection of the individual clones. The proteins expressed from these cDNAs are shown to be catalytically active. Similar to the endogenous enzymes, the expressed MAO A prefers serotonin as a substrate and is sensitive to the inhibitor clorgyline. In contrast, the expressed MAO B prefers phenylethylamine as a substrate and is sensitive to the inhibitor deprenyl. These results suggest that a single polypeptide of MAO A (or B), existing as either a monomer or homodimer, is enzymatically active. The ability to obtain functional MAO A and B from their respective cDNA clones allows us to study further the structure and function relationships of these important enzymes.

Cell Line↗
PubMed1988

Structure and functional expression of cloned rat serotonin 5HT-2 receptor.

A complementary DNA (cDNA) encoding a serotonin receptor with 51% sequence identity to the 5HT-1C subtype was isolated from a rat brain cDNA library by homology screening. Transient expression of the cloned cDNA in mammalian cells was used to establish the pharmacological profile of the encoded receptor polypeptide. Membranes from transfected cells showed high-affinity binding of the serotonin antagonists spiperone, ketanserin and mianserin, low affinity for haloperidol (a dopamine D2 receptor antagonist), 8-OH-DPAT as well as MDL-72222 and no detectable binding of [3H]serotonin. This profile is consonant with the 5HT-2 subtype of serotonin receptors. In agreement with this assignment, serotonin increased the intracellular Ca2+ concentration and activated phosphoinositide hydrolysis in transfected mammalian cells. The agonist also elicited a current flow, blocked by spiperone, in Xenopus oocytes injected with in vitro synthesized RNA containing the cloned nucleotide sequences.

Amino Acid Sequence↗
PubMed1988

Prevention of hypercholesterolemia and atherosclerosis in Japanese quail by high intake of soy protein.

This study was undertaken to determine the influence of levels of soy protein on cholesterol metabolism and the development of atherosclerosis in Japanese quail (Coturnix coturnix japonica). Quail were fed purified diets containing one of four levels (10, 20, 40 or 60%) of soy protein either with (atherogenic) or without (control) 0.5% cholesterol. Results showed that higher proportions of protein (40 and 60%) in atherogenic diets had a preventive effect on the development of atherosclerosis in adult male quail. For the atherogenic diets the higher protein levels resulted in significantly lower serum and aorta cholesterol concentrations than observed with the 10 or 20% protein levels and free of aortic atherosclerosis. A dual isotope technique was used to measure the absorption rates for cholesterol in quail fed with 20 and 40% soy protein. Absorption rates were not affected by the level of dietary protein but were influenced by the presence of dietary cholesterol. Relative rates were 40% in quail fed control diets and 30% in quail fed atherogenic diets. The excretion of neutral sterols, largely cholesterol, and bile acids increased with the high intake of soy protein. These results demonstrate that in quail the presence of higher dietary levels of soy protein has both a hypocholesterolemic action and a preventive effect on cholesterol-induced atherosclerosis and one of the possible mechanisms is through increased excretion of cholesterol.

Absorption↗
PubMed1988

cDNA cloning of human liver monoamine oxidase A and B: molecular basis of differences in enzymatic properties.

The monoamine oxidases play a vital role in the metabolism of biogenic amines in the central nervous system and in peripheral tissues. Using oligonucleotide probes derived from three sequenced peptide fragments, we have isolated cDNA clones that encode the A and B forms of monoamine oxidase and have determined the nucleotide sequences of these cDNAs. Comparison of the deduced amino acid sequences shows that the A and B forms have subunit molecular weights of 59,700 and 58,800, respectively, and have 70% sequence identity. Both sequences contain the pentapeptide Ser-Gly-Gly-Cys-Tyr, in which the obligatory cofactor FAD is covalently bound to cysteine. Based on differences in primary amino acid sequences and RNA gel blot analysis of mRNAs, the A and B forms of monoamine oxidase appear to be derived from separate genes.

Amino Acid Sequence↗
PubMed1988

Identification of 5-hydroxytryptamine1A receptor proteins in bovine frontal cortex.

5-Hydroxytryptamine1A (5-HT1A) receptor proteins were identified by a novel approach in which photoaffinity labeling technique was used in conjunction with affinity column chromatography. 5-HT1A receptors were solubilized from bovine frontal cortical membranes with 0.3% digitonin and 0.1% Nonidet P-40, and bound effectively to 1-[2-(4-aminophenyl)ethyl]-4-(3-trifluoromethylphenyl)piperazine (PAPP)-coupled Affi-Gel 10 in a time-dependent manner. PAPP was shown previously to be a selective ligand for the 5-HT1A receptor. Two protein bands with molecular masses of approximately 55,000 and 38,000 daltons revealed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis were eluted from the affinity column with either 1 mM 5-HT or 1 microM [3H]1-[2-(4-azidophenyl)ethyl]-4-(3-trifluoromethyl-phenyl)piperazine ([3H]p-azido-PAPP). [3H]p-Azido-PAPP is a selective photoaffinity labeling probe for the 5-HT1A receptor. The intensity of these two protein bands and the incorporation of [3H]p-azido-PAPP into these two proteins decreased significantly when the solubilized fraction was preincubated with excess 5-HT or PAPP (saturating all 5-HT1A receptors) prior to affinity column chromatography. These results suggest strongly that these two proteins are related to the 5-HT1A receptor protein. The isoelectric points of the photolabeled 5-HT1A receptor proteins were 6.0 and 6.5.

Affinity Labels↗
PubMed1988

Effect of anaerobic digestion on oocysts of the protozoan Eimeria tenella.

The effect of anaerobic digestion of poultry waste on oocysts of the protozoan Eimeria tenella, a common enteric pathogen that causes coccidiosis in poultry, was investigated in this study. Thermophilic (50 degrees C) and mesophilic (35 degrees C) anaerobic digestors, with poultry manure as the substrate, were inoculated with the oocysts. The oocysts were damaged during anaerobic digestion, as determined by morphological change and loss of their ability to sporulate. The recovered oocysts were tested for their infectivity in young chicks, as measured by body weight gain, mortality, and cecal lesions. Oocysts lost all their infectivity during thermophilic digestion, while oocysts subjected to mesophilic digestion remained moderately infective in comparison with untreated oocysts, which produced severe coccidiosis, high mortality, and low body weight gain in chicks. Oocysts were inactivated at 50 degrees C when they were suspended in digestor fluid or saline. Inactivation at 35 degrees C was significantly stronger in the digestor fluid than in the saline, which implied that factors other than temperature were involved in the lethal effect of anaerobic digestion on protozoan oocysts. In this study we demonstrated that the treatment of animal waste by anaerobic digestion, especially at a thermophilic temperature, has the benefits of pathogen control and protection of human and animal health in a farm environment.

Anaerobiosis↗
PubMed1988

Photoaffinity labeling of human placental monoamine oxidase-A by 4-fluoro-3-nitrophenyl azide.

Our previous work has shown that low concentrations of 4-fluoro-3-nitrophenyl azide (FNPA) (0.01-1 microM) photodependently inhibited only the type B monoamine oxidase in rat brain [Biochem. Pharmacol. 34:781-785 (1985)]. Evidence is presented in this paper indicating that higher concentrations of FNPA (15 microM) photodependently inhibit type A monoamine oxidase (MAO-A) from human placenta. FNPA acted as a competitive inhibitor for human placental MAO-A in the dark (Ki = 10 microM) when [14C]serotonin was used as the substrate. The inhibition of MAO-A activity by FNPA was concentration dependent and also irradiation time dependent. The specificity of the photodependent incorporation of FNPA to MAO-A was shown by the protective effect of serotonin during the irradiation. The kinetic analysis showed that the Vmax was decreased whereas the Km was not changed after FNPA was photolyzed with MAO-A. Furthermore, there was no recovery of MAO-A activity upon washing of the photolyzed FNPA-enzyme mixture. These results suggest that FNPA may be covalently bound to the substrate-binding site. Thus, under the present experimental conditions, FNPA is a suitable photoaffinity labeling probe for human placental MAO-A. This is the first photoaffinity label for MAO-A, which may be useful for characterizing the substrate-binding site of this enzyme.

Affinity Labels↗
PubMed1987

Photoaffinity labeling of beef liver monoamine oxidase-B by 4-fluoro-3-nitrophenyl azide.

4-Fluoro-3-nitrophenyl azide (FNPA) competitively inhibited beef liver monoamine oxidase-B (MAO-B) in the dark (Ki = 2.8 microM). Upon irradiation in the presence of FNPA, a concentration-dependent photoinactivation of MAO-B was observed. The kinetic analysis showed that the photoinactivation of MAO-B resulted in a decrease in Vmax but no change in Km. This result suggests that an irreversible linkage may be formed between the enzyme and the photolyzed FNPA. When [3H]FNPA was photoirradiated with the purified MAO-B, a single radioactive band associated with MAO-B was observed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The photo-dependent incorporation could be protected by phenylethylamine, the substrate for MAO-B, in a concentration-dependent manner. Complete tryptic-chymotryptic digestion of [3H]FNPA-labeled MAO-B resulted in three radioactive peaks on Sephadex G-25 column chromatography. With the same digestion and separation procedures, only one major radioactive peak was observed for the [3H]pargyline-labeled MAO-B, and its elution volume was different from that of [3H]FNPA-labeled peptides. These results suggest that, upon photolysis, FNPA may incorporate into a region in the active site of MAO-B which may be different from the pargyline binding site--the FAD prosthetic group of the enzyme.

Affinity Labels↗
PubMed1987

Detection of specific DNA segments of Marek's disease herpes virus in Japanese quail susceptible to atherosclerosis.

Genetically selected lines of Japanese quail, highly susceptible (SUS) and resistant (RES) to atherosclerosis, were used to study the possible involvement of Marek's disease herpes virus (MDV). An EcoRI gene library of MDV cloned in pBR328 was used to prepare the 32P-DNA probe in dot-blot and Southern blot hybridizations to detect the presence of MDV DNA sequence in the aorta, embryo and other tissue specimens. The viral DNA was found present in the aorta of SUS quail and it increased with the severity of the aortic lesion. For the DNA isolated from the atherosclerotic aorta, the endonuclease restriction map is specific but not identical to MDV genome. When screening the embryos of SUS and RES quail, it was found that all the SUS were positive with approximately 10 or more viral genome equivalents or virus copies per cell. The RES embryos were heterogeneous, 41% negative (less than 0.1 copy per cell), 43% intermediate (1-10 copies per cell) and 16% positive (10 or more copies per cell). The vaccination of SUS quail with the herpes virus of turkey vaccine did not prevent the disease. These results indicated that a part of MDV genome or another related herpes virus genome was integrated into the host DNA of SUS quail. The integrated viral gene or genes are believed to be important in atherogenesis, because they are genetically co-selected with the atherosclerosis-susceptibility.

Animals↗