Biomedical subjects

H Wang

Publications and source records attributed to H Wang.

At least 685 records · Page 38Linked to original sources

PubMed2000

Vibrio harveyi NADPH-FMN oxidoreductase arg203 as a critical residue for NADPH recognition and binding.

Luminous bacteria contain three types of NAD(P)H-FMN oxidoreductases (flavin reductases) with different pyridine nucleotide specificities. Among them, the NADPH-specific flavin reductase from Vibrio harveyi exhibits a uniquely high preference for NADPH. In comparing the substrate specificity, crystal structure, and primary sequence of this flavin reductase with other structurally related proteins, we hypothesize that the conserved Arg203 residue of this reductase is critical to the specific recognition of NADPH. The mutation of this residue to an alanine resulted in only small changes in the binding and reduction potential of the FMN cofactor, the K(m) for the FMN substrate, and the k(cat). In contrast, the K(m) for NADPH was increased 36-fold by such a mutation. The characteristic perturbation of the FMN cofactor absorption spectrum upon NADP(+) binding by the wild-type reductase was abolished by the same mutation. While the k(cat)/K(m,NADPH) was reduced from 1990 x 10(5) to 46 x 10(5) M(-1) min(-1) by the mutation, the mutated variant showed a k(cat)/K(m,NADH) of 4 x 10(5) M(-1) min(-1), closely resembling that of the wild-type reductase. The deuterium isotope effects (D)V and (D)(V/K) for (4R)-[4-(2)H]-NADPH were 1.7 and 1.4, respectively, for the wild-type reductase but were increased to 3.8 and 4.0, respectively, for the mutated variant. Such a finding indicates that the rates of NADPH and NADP(+) dissociation in relation to the isotope-sensitive redox steps were both increased as a result of the mutation. These results all provide support to the critical role of the Arg203 in the specific recognition and binding of NADPH.

Amino Acid Sequence↗

PubMed2000

SMA-3 smad has specific and critical functions in DBL-1/SMA-6 TGFbeta-related signaling.

A TGFbeta signal transduction cascade controls body size and male tail morphogenesis in the nematode Caenorhabditis elegans. We have analyzed the function of the sma-3 Smad gene, one of three Smad genes that function in this pathway. Null mutations in sma-3 are at least as severe as null mutations in the ligand and type I receptor genes, dbl-1 and sma-6, indicating that the other Smads do not function in the absence of SMA-3. Furthermore, null mutations in sma-3 do not cause defects in egg laying or in regulation of the developmentally arrested dauer larva stage, indicating no overlapping function with another C. elegans TGFbeta signaling pathway. The sma-3 gene is widely expressed at all developmental stages in hermaphrodites and males. The molecular lesions associated with eight sma-3 alleles of varying severity have been determined. The missense mutations cluster in two previously identified regions important for Smad function.

Alleles↗

PubMed2000

Self-assessed health-related quality of life in men who have completed radiotherapy for prostate cancer: instrument validation and its relation to patient-assessed bother of symptoms.

The purpose of this study was to develop a psychometrically reliable and valid questionnaire to assess the disease-specific dimensions of health-related quality of life (HRQOL) in the urinary function (UF), bowel function (BF), and sexual function (SF) domains of prostate cancer (PCa) patients treated with radiation therapy. Patients were given a six-page questionnaire using Likert-type questions assessing three HRQOL dimensions during their follow-up visits after completing radiotherapy. Scales created from an earlier study were utilized and tested for reliability and validity. In addition, we assessed the relationship between these dimensions and the degree to which a decreased HRQOL increases the degree to which patients feel bothered about their symptoms. There are two scales within each dimension: BF, Urgency and Daily Living; UF, Urgency and Weakness of Stream; SF, Interest/Satisfaction and Impotence. Internal-consistency reliability coefficients (Cronbach's alpha) for the proposed scales range from 0.48 to 0.92, and all item-scale correlations and divergence correlations validate the use of the scales, ranging from 0.49 to 0.89. The validity of these scales is also confirmed by the rising median scores with rising reported levels of patient-perceived "bother." The different dimensions have differing quantitative influences on patients. We have developed a prostate-specific HRQOL instrument that is an adequate and suitable tool for measuring HRQOL along three distinct dimensions for patients who have completed radiotherapy for PCa. Psychometric standards for reliability and validity were met for the proposed scales. Moreover, positive correlations were found between these dimensions and how bothered patients were by their symptoms, suggesting important relationships that should be followed in PCa patients after radiotherapy. Certain scales have strong influences on patient-perceived "bothersomeness" of symptoms, such as loss of control of BF, urgency of BF, urgency of urination, and level of interest/satisfaction in sex. Compared to our earlier study on patients being treated with radiotherapy for PCa, this study produced very similar results. With some modification, the same questionnaire could be used for both groups of patients. Int. J. Cancer (Radiat. Oncol. Invest.) 90, 163-172 (2000).

Adult↗

PubMed2000

Hepatocyte growth factor enhances MMP activity in human endothelial cells.

Scatter factor (SF) or hepatocyte growth factor (HGF) has been identified as an angiogenic factor. Angiogenesis requires not only tube formation but also invasion of pericytes and extracellular matrix (ECM) remodeling to promote new vessel stabilization. In the current study, the effect of SF/HGF on endothelial cell (EC) production of matrix metalloproteinases (MMPs) was explored. We showed that SF/HGF enhanced MT1-MMP synthesis and induced MMP-2 activation in two human EC lines: dermal microvessel EC and coronary arterial EC. Furthermore, SF/HGF accelerated EC invasion into matrix, an activity that could be inhibited by a MMP inhibitor. We also demonstrated that the MAP kinase cascade is critical in signal transduction pathway from SF/HGF stimulation to MT1-MMP up-regulation. The current study indicates that MMP activation is a novel effect of SF/HGF on ECs.

Cell Line↗

PubMed2000

Three telomerases with completely non-telomeric template replacements are catalytically active.

Telomerase is a reverse transcriptase minimally composed of a reverse transcriptase protein subunit and an internal RNA component that contains the templating region. Point mutations of template RNA bases can cause loss of enzymatic activity, reduced processivity and misincorporation in vitro. Here we report the first complete replacement of the nine base TETRAHYMENA: thermophila telomerase templating region in vivo with non-telomeric sequences. Rather than ablating telomerase activity, three such replaced telomerases (U9, AUN and AU4) were effective in polymerization in vitro. In vivo, the AU4 and AUN genes caused telomere shortening. We demonstrated the fidelity of the AUN and U9 telomerases in vitro and utilized AUN telomerase to demonstrate that 5' end primer recognition by telomerase is independent of template base pairing. However, the mutant AUN template telomerase catalyzed an abnormal DNA cleavage reaction. For these U-only and AU- substituted templates, we conclude that base-specific interactions between the telomerase template and protein (or distant parts of the RNA) are not absolutely required for the minimal core telomerase functions of nucleotide addition and base discrimination.

Animals↗

PubMed2000

Inability to induce hypertension in normotensive rat expressing AT(1) receptor antisense.

Our previous studies have shown that neonatal delivery of angiotensin type 1 receptor antisense (AT(1)R-AS) in a retroviral vector prevents spontaneously hypertensive rats from developing hypertension for life but has no effect on blood pressure (BP) in normotensive animals. Based on these results, we hypothesized that AT(1)R-AS transduction in normotensive rats would protect them from developing experimental hypertension. The present study was designed to evaluate this hypothesis. A single intracardiac administration of AT(1)R-AS by a retroviral-mediated delivery system (LNSV-AT(1)R-AS) in 5-day-old normotensive Sprague-Dawley rats resulted in long-term expression of the AT(1)R-AS without an effect on basal BP. However, angiotensin II (Ang II)-induced BP, dipsogenic responses, and renovascular contractility were significantly attenuated in the LNSV-AT(1)R-AS-treated rats. Chronic infusion of low-dose Ang II (55 ng. kg(-)(1). min(-)(1)) in LNSV-alone-treated rats caused a modest increase in BP, profound increase in cardiac hypertrophy, and increased vascular contractility. In contrast, the LNSV-AT(1)R-AS-treated rats were protected from developing these changes after Ang II infusion. These data establish that LNSV-AT(1)R-AS pretreatment protects healthy rats from developing Ang II-dependent hypertension.

Angiotensin II↗

PubMed2000

Electron acceleration and the propagation of ultrashort high-intensity laser pulses in plasmas

Reported are interactions of high-intensity laser pulses ( lambda = 810 nm and I</=3x10(18) W/cm(2)) with plasmas in a new parameter regime, in which the pulse duration ( tau = 29 fs) corresponds to 0. 6-2.6 plasma periods. Relativistic filamentation is observed to cause laser-beam breakup and scattering of the beam out of the vacuum propagation angle. A beam of megaelectronvolt electrons with divergence angle as small as 1 degrees is generated in the forward direction, which is correlated to the growth of the relativistic filamentation. Raman scattering, however, is found to be much less than previous long-pulse results.

Journal Article↗

PubMed2000

Rostrocaudal variation in targeting of N-methyl-D-aspartate and mu-opioid receptors in the rat medial nucleus of the solitary tract.

N-methyl-D-aspartate (NMDA) receptors and mu-opioid receptors (MOR) have been implicated in gustatory and cardiorespiratory visceral reflexes, respectively involving second order sensory neurons in rostral and intermediate portions of the medial nucleus of the solitary tract (mNTS). To determine whether there are cellular sites suggesting functional interaction involving NMDA receptors and MOR in these regions, we examined their ultrastructural immunocytochemical localization by using antisera recognizing the functional subunit of NMDA receptors (NR1) or MOR in rat brain. In both mNTS subdivisions, NR1 labeling was prominently seen along membranes of cytoplasmic organelles in somata and large dendrites, as well as on asymmetric postsynaptic densities in small dendrites and dendritic spines. Many of these profiles also contained MOR immunoreactivity that was mainly distributed along extrasynaptic plasma membranes. Quantitative regional comparison showed that dendrites composed 64% (167 of 261) and 35% (137 of 390) of the dually labeled structures in the rostral and intermediate mNTS, respectively. In contrast, only 11% (28 of 261) of the total dually labeled profiles in the rostral, but 46% (180 of 390) of those in the intermediate mNTS were axon terminals. Many of the terminals containing NR1 and/or MOR were large and formed asymmetric synapses with multiple targets, resembling those features of known visceral afferents. Our results suggest that opioids, active at MOR in mNTS, modulate excitatory visceral reflexes involving mainly postsynaptic NMDA receptors in the rostral region. In addition, they suggest that similar mechanisms exist in the intermediate mNTS, where both NMDA receptors and MOR may differentially regulate the presynaptic release of glutamate from the visceral afferents.

Animals↗

PubMed2000

The SCAN domain mediates selective oligomerization.

The SCAN domain is described as a highly conserved, leucine-rich motif of approximately 60 amino acids found at the amino-terminal end of zinc finger transcription factors. Although no specific biological function has been attributed to the SCAN domain, its predicted amphipathic secondary structure led to the suggestion that this domain may mediate protein-protein associations. A yeast two-hybrid screen identified members of two SCAN domain protein families that interact with the SCAN domain of the zinc finger protein ZNF202. The interacting ZNF191 protein represents the family of SCAN domain-containing zinc finger proteins, whereas the novel SDP1 protein establishes a new family of genes that encode an isolated SCAN domain. Isolated SCAN domain proteins may form asymmetric homodimers in solution. Biochemical binding studies confirmed the associations of ZNF191 and SDP1 with ZNF202 and established the SCAN domain as a selective hetero- and homotypic oligomerization domain. SCAN mediated protein associations might therefore represent a new regulatory mechanism of transcriptional activity.

Amino Acid Sequence↗

PubMed2000

Newly isolated archaerhodopsin from a strain of Chinese halobacteria and its proton pumping behavior.

A strain of extremely salt-loving halobacteria Halobacterium species xz515 from a salt lake in Tibet was isolated. SDS-polyacrylamide gel electrophoresis shows that there is only one protein on claret membrane, which is the same membrane fraction as purple membrane from Halobacterium salinarum, with a molecular weight close to bacteriorhodopsin (br). The purified retinal containing protein from xz515 has an absorption peak at around 550 nm. These facts indicate that it is a br-like protein. The partial sequence determination [H. Wang et al., Chin. Sci. Bull., 45 (2000)] shows that this br-like protein belongs to the archaerhodopsin family. The measurements of light-induced medium pH change in intact cells and cell envelope vesicles of xz515 suggest that this type of archaerhodopsin has a proton pumping function. However, the study about the dynamics of pumped protons across the membrane reveal that the proton release and proton uptake is in reverse order compared to br. The probable reason, attributing to regulating the rate of proton release is discussed.

Archaeal Proteins↗

PubMed2000

Structural and functional characteristics and tissue distribution pattern of rat OCTN1, an organic cation transporter, cloned from placenta.

This report describes the structure, function, and tissue distribution pattern of rat OCTN1 (novel organic cation transporter 1). The rat OCTN1 cDNA was isolated from a rat placental cDNA library. The cDNA is 2258 bp long and codes for a protein of 553 amino acids. Its amino acid sequence bears high homology to human OCTN1 (85% identity) and rat OCTN2 (74% identity). When expressed heterologously in mammalian cells, rat OCTN1 mediates Na(+)-independent and pH-dependent transport of the prototypical organic cation tetraethylammonium. The transporter interacts with a variety of structurally diverse organic cations such as desipramine, dimethylamiloride, cimetidine, procainamide, and verapamil. Carnitine, a zwitterion, interacts with rat OCTN1 with a very low affinity. However, the transport of carnitine via rat OCTN1 is not evident in the presence or absence of Na(+). We conclude that rat OCTN1 is a multispecific organic cation transporter. OCTN1-specific mRNA transcripts are present in a wide variety of tissues in the rat, principally in the liver, intestine, kidney, brain, heart and placenta. In situ hybridization shows the distribution pattern of the transcripts in the brain (cerebellum, hippocampus and cortex), kidney (cortex and medulla with relatively more abundance in the cortical-medullary junction), heart (myocardium and valves) and placenta (labyrinthine zone).

Amino Acid Sequence↗

PubMed2000

Reverse engineering a protein: the mechanochemistry of ATP synthase.

ATP synthase comprises two rotary motors in one. The F(1) motor can generate a mechanical torque using the hydrolysis energy of ATP. The F(o) motor generates a rotary torque in the opposite direction, but it employs a transmembrane proton motive force. Each motor can be reversed: The F(o) motor can drive the F(1) motor in reverse to synthesize ATP, and the F(1) motor can drive the F(o) motor in reverse to pump protons. Thus ATP synthase exhibits two of the major energy transduction pathways employed by the cell to convert chemical energy into mechanical force. Here we show how a physical analysis of the F(1) and F(o) motors can provide a unified view of the mechanochemical principles underlying these energy transducers.

Adenosine Triphosphate↗

PubMed2000

[Effect of calcitonin on the proliferation and membrane fluidity of human osteoblast-like cells].

The effects of calcitonin (CT) on the proliferation and membrane fluidity of human osteoblast-like cells line OS-732 were compared with the effect of 1,25(OH)2D3. The rate of DNA synthesis, cell proliferation, cell cycle and membrane fluidity of OS-732 cells were observed. Results showed that 1IU/ml CT stimulated cell proliferation and DNA synthesis. The percentage of cell in the G2 + M phase was increased and the percentage of cell in the synthetic phase was decreased. The membrane fluidity of OS-732 cells was increased after 0.1 mumol/L and 0.01 mumol/L 1,25(OH)2D3 treatment and was decreased after 1IU/ml CT treatment. It is concluded that calcitonin might increase the cell proliferation and decrease the membrane fluidity.

Bone Neoplasms↗

PubMed2000

[Mutation of p53 and Ki-ras gene in human fetal lung fibroblast cells in vitro by sterigmatocystin].

To explore the carcinogenic effects of sterigmatocystin(ST), one of the predominant contaminating mycotoxins in high risk areas of cancer in China, mutation of tumor suppressor gene p53 and oncogene Ki-ras in human fetal lung cells in vitro induced by ST was studied using cell culture and silver-staining PCR-SSCP methods. The results showed that, within 22 weeks of ST treatment, no abnormalities were found for both p53 and Ki-ras gene in electrophoresis. Abnormal electrophoretic migration bands were seen at exon 8 of p53 gene and Ki-ras gene in ST-treated human lung fibroblast cells 22 weeks after ST treatment. Thus, the results further confirmed the carcinogenic effects of ST on human fetal lung tissue.

Carcinogens↗

PubMed2000

[Recent progress in the study of common fusarium mycotoxins on apoptosis of cells].

Fumonisin B1, T-2 toxin, Fusarenon-X and Deoxynivalenol, which are all mycotoxins produced by fusarium, could induce apoptosis of liver cell, kidney cell, gastrointestinal epithelial cell, immunological cells as well as several cell lines of human and animals. The possible mechanisms of apoptosis induction are complicated. Sphingol, p21 gene, protein kinase, CAMP signal system and intracellular Ca2+ level, might be involved in the cellular apoptosis induction by Fusarium mycotoxins.

Animals↗

PubMed2000

Effects of the renin-angiotensin system on the current I(to) in epicardial and endocardial ventricular myocytes from the canine heart.

The Ca(2+)-independent portion of transient outward K(+) current (I(to)) exhibits a transmural gradient in ventricle. To investigate control mechanisms for this gradient, we studied canine epicardial and endocardial ventricular myocytes with use of the whole-cell patch-clamp technique. I(to) was larger in amplitude, had a more negative voltage threshold for activation, and had a more negative midpoint of inactivation in epicardium. Recovery from inactivation was >10-fold slower in endocardium. Incubation of epicardial myocytes with angiotensin II for 2 to 52 hours altered I(to) to resemble unincubated endocardium and reduced the amplitude of the phase 1 notch of the action potential. In contrast, incubation of endocardial myocytes with losartan for 2 to 52 hours altered I(to) to resemble unincubated epicardium and induced a phase 1 notch in the action potential. With RNase protection assays, we determined that incubations with angiotensin II or losartan did not alter mRNA levels for either Kv4.3 or Kv1.4; thus, a change in the alpha subunit for I(to) is unlikely to be responsible. To test whether posttranslational modification produced the effects of angiotensin II, we coexpressed Kv4.3 and the angiotensin II type 1a receptor in Xenopus oocytes. Incubation with angiotensin II increased the time constant for recovery from inactivation of the expressed current by 2-fold with an incubation time constant of 3.7 hours. No effect on activation or inactivation voltage dependence was observed. These results demonstrate that the properties of I(to) in endocardium and epicardium are plastic and likely under the tonic-differing influence of the renin-angiotensin system.

Action Potentials↗

PubMed2000

Vagus nerve stimulation attenuates the systemic inflammatory response to endotoxin.

Vertebrates achieve internal homeostasis during infection or injury by balancing the activities of proinflammatory and anti-inflammatory pathways. Endotoxin (lipopolysaccharide), produced by all gram-negative bacteria, activates macrophages to release cytokines that are potentially lethal. The central nervous system regulates systemic inflammatory responses to endotoxin through humoral mechanisms. Activation of afferent vagus nerve fibres by endotoxin or cytokines stimulates hypothalamic-pituitary-adrenal anti-inflammatory responses. However, comparatively little is known about the role of efferent vagus nerve signalling in modulating inflammation. Here, we describe a previously unrecognized, parasympathetic anti-inflammatory pathway by which the brain modulates systemic inflammatory responses to endotoxin. Acetylcholine, the principle vagal neurotransmitter, significantly attenuated the release of cytokines (tumour necrosis factor (TNF), interleukin (IL)-1beta, IL-6 and IL-18), but not the anti-inflammatory cytokine IL-10, in lipopolysaccharide-stimulated human macrophage cultures. Direct electrical stimulation of the peripheral vagus nerve in vivo during lethal endotoxaemia in rats inhibited TNF synthesis in liver, attenuated peak serum TNF amounts, and prevented the development of shock.

Acetylcholine↗

PubMed2000

N-hydroxysuccinimidyl fluorescein-O-acetate as a fluorescent derivatizing reagent for catecholamines in liquid chromatography.

A new amine-reactive derivatizing reagent, N-hydroxysuccinimidyl fluorescein-O-acetate (SIFA), was developed for catecholamine (CA) analysis in liquid chromatography. The reactivity of this reagent with the CAs norepinephrine (NE), epinephrine (E), and dopamine (DA) was investigated in detail. In aqueous methanol containing 32 mmol/L pH 9.0 H3BO3-Na2B4O7 buffer, SIFA reacted with NE, E, and DA under mild conditions. The derivatives were separated in 20 min on a C18 column with a mobile phase of methanol/water (38:62, v/v) containing 10 mmol/L pH 5.0 H3cit-Na2HPO4 buffer. At lambda(ex)/lambda(em) = 490/516 nm, the detection limits were 3.2, 12, and 56 fmol, respectively, with a signal-to-noise ratio of 3, which were comparable to those using 1,2-diphenylethylenediamine as the derivatizing reagent for CA analysis. Amino acids, aliphatic amines, and alcohols had no obvious interference with the determination. The proposed method has been applied to the determination of CAs in human urine, with recoveries of 95.3-103.9%.

Calibration↗