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Biomedical subjects

H Shen

Publications and source records attributed to H Shen.

At least 307 records · Page 17Linked to original sources
PubMed1994

Expression of immunoreactive polysialylated neural cell adhesion molecule in the suprachiasmatic nucleus.

Light-microscopic and immunoblot immunochemical procedures were used to study the distribution and biochemical characteristics of neural cell adhesion molecule (NCAM) and its polysialylated form (PSA-NCAM) in the suprachiasmatic nuclei (SCN) of the adult Siberian hamster. In the adult brain PSA-NCAM is located in regions capable of undergoing morphological rearrangements and thus is generally considered to be an indicator of neural plasticity. Immunostaining for PSA in the Siberian hamster SCN (using a monoclonal antibody against the alpha 2,8-linked PSA of NCAM) was evident throughout the rostrocaudal axis of the SCN, with the most intense reaction in the ventrolateral region. Immunoreactivity was present in the neuropil, which delineated groups of cells with unstained cytoplasm. Many of the SCN cells were aggregated into cords or clusters. The optic chiasm was free from label, except for short processes apparently extending from the densely stained neuropil in the ventrolateral SCN. Immunoreactivity was abolished by preincubating sections in an endoneuraminidase (endo-N) or by preincubation of the primary antibody with PSA-NCAM. Immunostaining of the nonsialylated NCAM polypeptide was also limited to the neuropil, but this was more diffuse and less regionally specific than PSA staining. The ventral SCN exhibited the most intense labeling for NCAM. Immunoblot analyses revealed the immunoreactive PSA-NCAM as a broad band migrating between apparent molecular weights in the range of 150-300 kD, which was ablated by treatment of SCN tissue extract with endo-N. This electrophoretic migration pattern of PSA-NCAM from the SCN region was similar to that seen in several other brain regions.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

[A new concept and method of standardizing predictive value].

A new concept and method of standardizing predictive value was put forward. The theoretical predictive value which was not affected by prevalence rate or different component in normal and abnormal groups and was gotten under the appointed standardizing condition that made the cases in the two groups to be same with each other and to be called standardizing component was defined as standardizing predictive value. The value will change only with the change of sensitivity and specificity of diagnostic test. By the new concept and method, the value of diagnostic test can be explained, judged and expressed in unique standard.

False Negative Reactions↗
PubMed1993

Use of a DNA-based test for the mutation associated with porcine stress syndrome (malignant hyperthermia) in 10,000 breeding swine.

To test the hypothesis that the mutation associated with porcine stress syndrome (PSS; malignant hyperthermia) was present in a large proportion of North American and English swine, a simple and rapid laboratory protocol was used for cost-effective, large-scale diagnosis of susceptibility to PSS. This PSS test was applied to 10,245 breeding swine of various breeds from 129 farms in the United States, Canada, and England. Approximately 1 of 5 swine was a heterozygous carrier of the PSS mutation, with approximately 1% being homozygotes. Prevalence of the PSS mutation was 97% for 58 Pietrain, 35% for 1,962 Landrace, 15% for 718 Duroc, 19% for 720 Large White, 14% for 496 Hampshire, 19% for 1,727 Yorkshire, and 16% for 3,446 crossbred swine. The PPS gene frequencies for these breeds were 0.72, 0.19, 0.08, 0.10, 0.07, 0.10, and 0.09, respectively. In addition to these breeds, we have identified the PSS mutation in Poland China and Berkshire breeds. These gene frequencies were 30 to 75% lower in Canadian swine than in US swine, with the exception of Yorkshires, for which the gene frequency was threefold higher in Canadian swine. English swine were similarly, or more so, affected than were US swine. Accuracy was estimated at > 99%. Cost to perform the test was < $20/animal. Depending on the perceived net balance of deleterious and beneficial effects of the mutation, the PSS test could be used to eradicate the PSS mutation from herds, or for controlled expression of the mutation.

Animals↗
PubMed1993

Purification, characterization and binding interactions of the Chinese-cobra (Naja naja atra) serum antitoxic protein CSAP.

The characterization of the single-chain protein in Chinese-cobra (Naja naja atra) blood serum, which yields strong specific protection against the venom of the same snake, is reported. The protein, CSAP (cobra serum antitoxic protein), was purified to electrophoretic homogeneity. Over the pH range 5-9 it formed stable complexes with the neuro- and the cardio-toxin of the snake. The molecular size of the CSAP was estimated to be 70.3 +/- 0.3 kDa. Tryptic hydrolysis of CSAP yielded several peptides that were able to bind to the toxin. The native CSAP maximally bound 8 +/- 1 toxin molecules/molecule. Six tryptic fragments, containing 5-39 residues, were sequenced. The longest of these displayed sequence similarity to rat serum albumin. The protective effect of the CSAP was demonstrated in vivo on mice and in vitro by measurement of the rate of haemolysis. Kinetic and thermodynamic parameters of the binding interactions of the neurotoxin and the CSAP were determined from the rates of displacement of 125I-labelled toxin from its complexes with the CSAP by unlabelled toxin by using a DEAE-cellulose filter assay for CSAP-toxin complexes. The toxin molecules rapidly dissociated from one type of site and slowly from a second. The binding capacity and concentration of the CSAP suffice to explain the protective effect of the latter against the toxin.

Amino Acid Sequence↗
PubMed1993

Characterization of vascular permeability factor/vascular endothelial growth factor receptors on mononuclear phagocytes.

Vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) is a polypeptide mediator, elaborated by certain tumors and other cell types, that exerts multiple effects on endothelium via interaction with a class of high-affinity binding sites. In this report, the interaction of VPF/VEGF with human mononuclear phagocytes (MPs) is characterized. Radioligand binding studies at 4 degrees C showed the presence of a single class of binding sites, kd approximately 300 to 500 pmol/L (approximately 20 times lower affinity than the high-affinity binding site on endothelial cells [ECs]), the occupancy of which correlated with VPF/VEGF-induced MP migration and expression of tissue factor. These binding results were paralleled by functional experiments which indicated that the same VPF/VEGF preparations were about an order of magnitude less effective in stimulating MP chemotaxis than in inducing EC proliferation. When MPs with surface-bound 125I-VPF/VEGF were warmed to 37 degrees C, endocytosis and degradation occurred. Occupancy of VPF/VEGF binding site resulted in subsequent activation of intracellular signal transduction mechanisms, as shown by an increase in MP intracellular calcium concentration. Cross-linking studies with 125I-VPF/VEGF showed a new high-molecular weight band (corresponding to putative 125I-VPF/VEGF-receptor complex), the appearance of which was blocked by excess unlabeled VPF/VEGF. Consistent with these results, immunoprecipitation of 32PO4-labeled MPs exposed to VPF/VEGF showed a single band of similar mobility, not seen in untreated controls. These results demonstrate that the interaction of VPF/VEGF with MPs, though of lower affinity than that observed with ECs, also results from interaction of the polypeptide with a specific cell-surface protein and leads to activation of intracellular transduction mechanisms.

Cell Division↗
PubMed1993

Identification of cysteine residues involved in disulfide formation in the inactivation of glutathione transferase P-form by hydrogen peroxide.

We previously reported that rat glutathione transferase P-form (GST-P) is inactivated by hydrogen peroxide (H2O2). This involves formation of intra- or intersubunit disulfides, at least three extra bands with molecular masses of 21.5, 18, and 37 kDa being exhibited in addition to the native subunit band of 23.5 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under nonreducing conditions. In the present study, GST-P mutants whose cysteine residues were independently substituted with alanine (C14A, C47A, C101A, and C169A) by site-directed mutagenesis were used to identify the cysteine residues responsible for the disulfide bond formation. C14A and C169A were much more inactivated than native GST-P by 1 mM H2O2, whereas C47A and, especially, C101A appeared insensitive to H2O2. On SDS-PAGE, the 21.5-kDa band was not detected in either C47A or C101A. Hydrogen peroxide treatment of mouse GST II, highly homologous to rat GST-P but possessing glycine instead of cysteine at the 101st residue, did not result in generation of the 21.5-kDa band and was also associated with less inactivation. This band was therefore considered to be due to an intrasubunit disulfide bond between Cys-47 and Cys-101. The 37-kDa band was suggested to be due to the formation of intersubunit disulfide bonds between Cys-47 residues in different subunits. Thus the Cys-47 residue together with Cys-101 may be located in an important region for GSH binding, disulfide bond formation between these residues resulting in steric hindrance.

Animals↗
PubMed1993

Localized gene transfer into organotypic hippocampal slice cultures and acute hippocampal slices.

Viral vectors derived from herpes simplex virus, type-1 (HSV), can transfer and express genes into fully differentiated, post-mitotic neurons. These vectors also transduce cells effectively in organotypic hippocampal slice cultures. Nanoliter quantities of a virus stock of HSVlac, an HSV vector that directs expression of E. coli beta-galactosidase (beta-gal), were microapplied into stratum pyramidale or stratum granulosum of slice cultures. Twenty-four hours later, a cluster of transduced cells expressing beta-gal was observed at the microapplication site. Gene transfer by microapplication was both effective and rapid. The titer of the HSVlac stocks was determined on NIH3T3 cells. Eighty-three percent of the beta-gal forming units successfully transduced beta-gal after microapplication to slice cultures. beta-Gal expression was detected as rapidly as 4 h after transduction into cultures of fibroblasts or hippocampal slices. The rapid expression of beta-gal by HSVlac allowed efficient transduction of acute hippocampal slices. Many genes have been transduced and expressed using HSV vectors; therefore, this microapplication method can be applied to many neurobiological questions.

Animals↗
PubMed1993

Comparative mammal model of chronic rate overload: relationship of myocardial Ca-cycling to heart, metabolic and lipoperoxidation rates.

1. The cardiac cycle is generally believed to be timed by a sarcoplasmic Ca-cycle and powered by a sarcoplasmic ATP-cycle that uses fatty acids as fuel and generates toxic free radicals as a side-product. 2. This study used a comparative mammal approach to test this model and the hypothesis that these cycles were closely coupled and correlated with fatty acid oxidation and peroxidative injury. 3. Fatty acid oxidation and ATP-cycling rates correlated to log heart rate whereas Ca-cycling rate was directly coupled to heart rate in a one-to-one relationship. 4. Both Ca-pump and Ca-channel activities coordinately increased as heart rate increased across species. 5. Thus, Ca-cycling activity is constant across mammals, when normalized to heart rate. 6. Comparison of Ca-ATPase and Ca-flux rates indicated that sarcoplasmic volume was inversely correlated with log heart rate. Basal lipoperoxidation of myocardium and susceptibility of SR to lipoperoxidation correlated with metabolic rate. 7. We identified that the horse is a metabolic outlier amongst mammals, with abnormally high fatty acid oxidation and ATP-synthetase activity compared to its heart rate.

Animals↗
PubMed1993

Production and properties of a factor X-cellulose-binding domain fusion protein.

A fusion protein, FX-CBDCex, which comprises factor X with a cellulose-binding domain (CBDCex) fused to its C-terminus, was produced in BHK cells. It was purified from the culture medium by affinity chromatography on cellulose. FX-CBDCex could be activated to FXa-CBDCex with Russell viper venom. FXa-CBDCex was as active as FXa against a chromogenic substrate and against proteins containing the Ile-Glu-Gly-Arg sequence hydrolysed by FXa. FXa-CBDCex retained its activity when adsorbed to cellulose.

Amino Acid Sequence↗
PubMed1993

Characterization of enzymatic reduction of hexavalent chromium by Escherichia coli ATCC 33456.

Chromium reduction by Escherichia coli ATCC 33456 quantitatively transferred hexavalent chromium, Cr(VI), to trivalent chromium, Cr(III). The reduced chromium was predominantly present in the external medium. Supernatant fluids of cell extract, obtained by centrifugation at 12,000 and 150,000 x g, showed almost the same Cr(VI) reduction activity, indicating that Cr(VI) reduction by E. coli ATCC 33456 was a largely soluble reductase activity. In studies with respiratory inhibitors, no inhibitory effects on aerobic and anaerobic Cr(VI) reduction were demonstrated by addition of cyanide, azide, and rotenone into both intact cell cultures and supernatant fluids of E. coli ATCC 33456. Although cytochromes b and d were identified in the membrane fraction of cell extracts, Cr(VI) was not reduced by the membrane fraction alone. The cytochrome difference spectra analysis also indicated that these cytochromes of the respiratory chain require the presence of the soluble Cr(VI) reductase to mediate electron transport to Cr(VI). Stimulation of Cr(VI) reduction by an uncoupler, 2,4-dinitrophenol, indicated that the respiratory-chain-linked electron transport to Cr(VI) was limited by the rate of dissipation of the proton motive force.

Aerobiosis↗
PubMed1993

Characterization of the promoter of avirulence gene D from Pseudomonas syringae pv. tomato.

The avirulence gene D (avrD) from Pseudomonas syringae pv. tomato comprises the first open reading frame (ORF) of a putative operon consisting of at least five tandem ORFs. The promoter of the avrD operon was localized to a 150-bp DNA fragment occurring 5' to the avrD gene by using the Tn7-lux and gus reporter systems. The avrD promoter in P. syringae pv. tomato and P. syringae pv. glycinea was poorly expressed when bacteria were grown in complex culture media but was activated during bacterial growth in plants. The timing and level of induction were similar in compatible and incompatible plant-pathogen interactions. When bacteria were grown in minimal culture medium, promoter activity was repressed by certain carbon sources, high concentrations of nitrogen compounds, and pH values above 6.5. Primer extension experiments on RNA from bacteria grown in minimal medium identified two transcription initiation sites 87 and 41 nucleotides upstream from the translational start site. Only the -41 transcriptional start site was identified in bacteria grown in soybean leaves. A sigma 54 promoter consensus sequence (GG-10 bp-GC) occurred 14 bp upstream of the -41 transcriptional start, and 3' deletions into this region completely abolished promoter activity. Little expression was observed when a gus fusion with the avrD promoter was introduced into an ntrA mutant strain of P. syringae pv. phaseolicola deficient in the sigma 54 cofactor. Expression from the avrD promoter also required the hrp regulatory genes, hrpS and hrpL. Deletions from the 5' end of the promoter region and base substitution analyses also identified two upstream elements important for expression. Sequence comparison of these elements with other cloned avirulence genes revealed the presence of a conserved consensus sequence elements with other cloned avirulence genes revealed the presence of a conserved consensus sequence (GGAACC-N15/16-CCAC) in the promoters of nine different avirulence genes from P. syringae pathovers.

Amino Acid Sequence↗
PubMed1993

[Pharyngeal cross-sectional area and pharyngeal compliance in normal Japanese males and females].

It is reported that obstructive sleep apnea syndrome (OSAS) has a marked elderly male predominance and that patients with OSAS have reduced pharyngeal cross-sectional area and increased pharyngeal collapsibility. These reports suggest that the size and mechanical properties of the pharynx have sex- and age-related differences even in normal subjects. However, there are only a few reports on normal values of pharyngeal cross-sectional area and compliance. The purpose of this study was to determine the normal values of the pharyngeal cross-sectional area and compliance in Japanese healthy subjects and to determine sex-related differences and the effect of age, body size, and body posture on these parameters. We examined 181 subjects with an age range spanning 21-69 years, with normal range of body mass index and without enlarged tonsils or jaw retrusion. No subject complained of symptoms suggestive of OSAS. We measured the pharyngeal cross-sectional area in three body positions (sitting, lateral and supine) using the acoustic reflection technique (AAAR). Fifty airway area versus distance functions with a rate of 3 times per second were measured and averaged in each body position during normal breathing of room air. After identifying the fauces and the glottis on the averaged area versus distance function, we defined a region between 2 cm distal to the fauces and 2 cm proximal to the glottis as a pharyngeal segment and computed the mean area of that segment (mean pharyngeal cross-sectional area).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1993

Soft X-ray microimages of immunogold-labelled tumor cell surface antigens.

After successfully introducing the colloidal immunogold labelling technique to soft X-ray contact microscopy (SXCM), SXCM replica microimages of immunogold labelled tumor cell HLA-A2 surface antigens have been acquired for the first time. A biological sample preparation method using immunocytochemical staining techniques suitable for SXCM has also been established.

Antibodies, Monoclonal↗