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Biomedical subjects

L Yang

Publications and source records attributed to L Yang.

At least 325 records · Page 18Linked to original sources
PubMed1999

Determination of dissociation constants of amino acids by capillary zone electrophoresis.

A method is proposed for the determination of dissociation constants of amino acids by capillary zone electrophoresis. According to the dissociation equilibrium of amino acids and the basic theory of electrophoresis, the nonlinear relationship between the pH value of the buffer and the effective electrophoretic mobilities of the analyte was obtained. The dissociation constants can be calculated from the pH values and the corresponding effective electrophoretic mobilities using the program written in C++. The dissociation constants, pKa1 and pKa2, of 11 kinds of amino acids were determined successfully by the proposed method. The determined dissociation constants were compared with values in the literature; the differences between them are in the range of -0.03 to 0.06. No significant differences were observed between the determined dissociation constants and the corresponding literature values.

Amino Acids↗
PubMed1999

Adhesion strength differential of human ligament fibroblasts to collagen types I and III.

Fibroblasts embedded in the amorphous healing tissue matrix of ligaments migrate into damaged sites during the inflammatory process that precedes the formation of new connective tissue after ligament injury. Cell motility involved in this migration is strongly influenced by cellular adhesion to proteins of the extracellular matrix. The adhesion mechanism of interest in this study is the attachment of fibroblasts from the anterior cruciate and medial collateral ligaments to types I and III collagen, two fibrillar collagens secreted by fibroblasts during tissue repair. Types I and III collagen constitute a major portion of these ligaments and are assembled by fibroblasts into long cable-like fibrils in the extracellular space. In this study, a micropipette aspiration technique was used to measure the force required to separate fibroblasts of the anterior cruciate and medial collateral ligaments from substrates composed of type I or III collagen, each at a concentration of 2 or 5 microg/ml. Approximately 1,200 fibroblasts from the anterior cruciate ligament and 1,600 from the medial collateral ligament were used, and the adhesion force and area of these cells were determined. Fibroblasts from the anterior cruciate ligament exhibited greater adhesion force than did those from the medial collateral ligament for all concentrations of types I and III collagen. In addition, the adhesiveness of fibroblasts from both ligaments was dependent on seeding time for all experimental conditions. To determine the adhesiveness per unit area, defined here as the adhesion strength, the adhesion force was normalized by the adhesion area. At early seeding times (15-45 minutes), fibroblasts from the anterior cruciate ligament exhibited greater adhesion strength on surfaces coated with type-I collagen than did those from the medial collateral ligament. However, for both collagen substrates, adhesion strength for fibroblasts from the anterior cruciate ligament decreased with seeding time whereas that for fibroblasts from the medial collateral ligament remained relatively constant for all seeding periods (15-75 minutes).

Adult↗
PubMed1999

Interphotoreceptor retinoid-binding protein (IRBP) is rapidly cleared from the Xenopus interphotoreceptor matrix.

The interphotoreceptor matrix (IPM) is a highly-organized extracellular matrix critical to retinal development and function. Although the concentrations of its components are carefully regulated, little is known about the mechanisms of this regulation. Interphotoreceptor retinoid-binding protein (IRBP) is the most abundant soluble protein component of the IPM. Although its rate of clearance is thought to be an important factor regulating the concentration of IRBP within the IPM, no study has measured the rate of its extracellular turnover. Here we determine the rate of turnover of matrix IRBP in Xenopus. The rate of IRBP turnover was estimated by measuring the loss of radioactivity from protein labeled by a single injection of a radiolabeled protein precursor. To provide an estimate of the rate of IRBP turnover, we have examined the following issues: (1) Quantitative extraction of IRBP from the IPM for biochemical analysis. (2) Routes of delivery of radiolabeled precursor to achieve a pulse label in vivo. (3) Selection of labeled precursor in order to minimize reutilization of radiolabel. Using Western blot analysis, immunoprecipitation and immuno-electron microscopy, we found that IRBP can be quantitatively extracted from the IPM by a simple saline wash. IRBP was radiolabeled by systemic or intravitreal injection of either [35S]methionine or carboxyl-terminal labeled [1-14C]leucine. The specific activity of matrix IRBP was determined by either phosphorimaging or fluorography of Coomassie blue-stained SDS-polyacrylamide gels. Intravitreal injection of tracer was more effective than systemic delivery in achieving a pulse of radiolabel to the retina. This may be due to intravitreal injection allowing the body to act as a 'sink' for radiolabeled amino acid. When radiolabeled precursor was delivered by intravitreal injection, the calculated half-life of matrix IRBP using [35S]methionine was 25. 6+/-0.82 hr; in contrast, it was 10.7+/-2.9 hr using [1-14C]leucine. The faster apparent IRBP turnover using [1-14C]leucine is interpreted in context of the early decarboxylation of leucine during its degradation. Our results demonstrate rapid turnover of IRBP in the Xenopus IPM in vivo and suggest that the IPM is a dynamic structure undergoing continuous renewal.

Animals↗
PubMed1999

Novel free radical spin traps protect against malonate and MPTP neurotoxicity.

Both malonate and 1-methyl-4-phenyl-1,2,5,6 tetrahydropyridine (MPTP) are neurotoxins which cause energy depletion, secondary excitotoxicity, and free radical generation. Malonate is a reversible inhibitor of succinate dehydrogenase, while MPTP is metabolized to 1-methyl-4-phenylpyridinium, an inhibitor of mitochondrial complex I. We examined the effects of pretreatment with the cyclic nitrone free radical spin trap MDL 101,002 on malonate and MPTP neurotoxicity. MDL 101,002 produced dose-dependent neuroprotection against malonate-induced striatal lesions. MDL 101, 002 produced significant protection against MPTP induced depletions of dopamine and its metabolites. MDL 101,002 also significantly attenuated MPTP-induced increases in striatal 3-nitrotyrosine concentrations. The free radical spin trap tempol also produced significant protection against MPTP neurotoxicity. These findings provide further evidence that free radical spin traps produce neuroprotective effects in vivo and suggest that they may be useful in the treatment of neurodegenerative diseases.

Animals↗
PubMed1999

Creatine and cyclocreatine attenuate MPTP neurotoxicity.

Systemic administration of 1-methyl-4-phenyl-1,2,3, 6-tetrahydropyridine (MPTP) produces parkinsonism in experimental animals by a mechanism involving impaired energy production. MPTP is converted by monoamine oxidase B to 1-methyl-4-phenylpyridinium (MPP+), which blocks complex I of the electron transport chain. Oral supplementation with creatine or cyclocreatine, which are substrates for creatine kinase, may increase phosphocreatine (PCr) or cyclophosphocreatine (PCCr) and buffer against ATP depletion and thereby exert neuroprotective effects. In the present study we found that oral supplementation with either creatine or cyclocreatine produced significant protection against MPTP-induced dopamine depletions in mice. Creatine protected against MPTP-induced loss of Nissl and tyrosine hydroxylase immunostained neurons in the substantia nigra. Creatine and cyclocreatine had no effects on the conversion of MPTP to MPP+ in vivo. These results further implicate metabolic dysfunction in MPTP neurotoxicity and suggest a novel therapeutic approach, which may have applicability for Parkinson's disease.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1999

Preliminary clinical application of contrast-enhanced MR portography.

The clinical application of contrast enhanced (CE) MR was evaluated. A total of 66 CE MR portograms were obtained by performing fast imaging with steady procession (FISP) technique on a 1.5-T Siemens magnetom vision. A maximum intensity projection algorithm was also employed to include all vessels in a single image. The patency of portal venous system, the presence and extent of varices were also evaluated. The results showed that all images had diagnostic quality. Main portal vein (MPV) and its 4th-6th level intrahepatic branches were visualized in 10 normal persons serving as control. The diameter of MPV, splenic vein and superior mesenteric vein was 1.02 +/- 0.21, 0.8 +/- 0.15, 0.8 +/- 0.26 cm respectively, which were significantly lower than that in portal hypertension patients (1.38 +/- 0.27, 1.26 +/- 0.18, 1.24 +/- 0.18 cm, respectively). In 23 preoperative cases of portal hypertension, dilated portal vein and tortuous enlarged splenic vein were found in 23 cases; esophageal and coronary varices in 12 and 19 cases, respectively. In 7 postoperative re-examined cases with portal hypertension, the flow velocity and flow of MPV were decreased in all cases and esophageal varices could still be observed in 3 cases. New vessels appeared in the great curvature of stomach in 2 cases. In 20 cases of liver carcinoma, occlusion of MPV or its intrahepatic branches were showed in 14, compression and dislocation of intrahepatic portal vein were found in 6. In other 6 cases, 2 were splenic venous thrombosis and 4 were tumors in the intestine or retro-peritoneum. It is concluded that three-dimensional CE MR portography is an accurate technique for evaluating the portal venous system. It is a reliable and noninvasive technique that can provide important information for the evaluation of patients' condition before TIPSS and liver transplantation.

Contrast Media↗
PubMed1999

Antigenic and genetic variations of the 15 kD nucleocapsid protein of porcine reproductive and respiratory syndrome virus isolates.

The antigenic variability of the 15 kD nucleocapsid protein of porcine reproductive and respiratory syndrome (PRRS) virus was characterized with a panel of 24 monoclonal antibodies (MAbs) raised against the American PRRS virus isolate ISU-P. Five continuous epitopes designated EpORF7-A through E were revealed by the reactivity pattern of these MAbs with 67 American field isolates, two modified-live vaccine viruses, and the European Lelystad virus as determined by the indirect immnofluorescence assay and Western immunoblotting and confirmed by additivity and blocking enzyme-linked immunosorbent assays. The reactivity pattern of isolates in the IFA permitted their subdivision into 4 American antigenic groups which represented 84.1, 11.6, 2.9 and 1.4% of viruses tested. The antigenic variation among isolates was correlated to single, group specific nucleotide substitutions and mediated by a combination of at least 4 of the 5 epitopes. EpORF7-A was conserved in all American isolates and the Lelystad virus which constituted a separate antigenic group. Consequently, monoclonal antibodies specific for EpORF7-A may prove useful as the antigenic basis for a universal diagnostic test for the PRRS virus. EpORF7-C, D and E were only present in the American isolates tested.

Amino Acid Sequence↗
PubMed1999

Active transforming growth factor-beta in wound repair: determination using a new assay.

Transforming growth factor (TGF)-beta regulates wound repair and scarring in an isoform-specific fashion. TGF-beta is produced in a latent form, and its activation is a critical regulatory step controlling the bioactivity of this growth factor. To date, it has been impossible to determine latent TGF-beta activation in vivo due to a lack of quantitative assays. We describe here a semiquantitative modification of the plasminogen activator inhibitor-1/luciferase bioassay (PAI/L assay) for TGF-beta, which we used to determine active and latent TGF-beta isoforms in frozen sections of rat wound tissue. We found that significant amounts of latent TGF-beta were rapidly activated upon wounding (38% of the total TGF-beta at 1 hour after wounding). A second peak of active TGF-beta (17% of total) occurred at 5 days after wounding. The predominant isoforms were TGF-beta1 and -2 with only minor amounts of TGF-beta3 present. This is the first TGF-beta bioassay allowing semiquantitative determination of active and latent isoforms present in vivo, and our results document the significance and temporal regulation of latent TGF-beta isoform activation in wound repair.

Animals↗
PubMed1999

Supramolecular structures of peptide assemblies in membranes by neutron off-plane scattering: method of analysis.

In a previous paper (Yang et al., Biophys. J. 75:641-645, 1998), we showed a simple, efficient method of recording the diffraction patterns of supramolecular peptide assemblies in membranes where the samples were prepared in the form of oriented multilayers. Here we develop a method of analysis based on the diffraction theory of two-dimensional liquids. Gramicidin was used as a prototype model because its pore structure in membrane in known. At full hydration, the diffraction patterns of alamethicin and magainin are similar to gramicidin except in the scale of q (the momentum transfer of scattering), clearly indicating that both alamethicin and magainin form pores in membranes but of different sizes. When the hydration of the multilayer samples was decreased while the bilayers were still fluid, the in-plane positions of the membrane pores became correlated from one bilayer to the next. We believe that this is a new manifestation of the hydration force. The effect is most prominent in magainin patterns, which are used to demonstrate the method of analysis. When magainin samples were further dehydrated or cooled, the liquid-like diffraction turned into crystal-like patterns. This discovery points to the possibility of investigating the supramolecular structures with high-order diffraction.

Cell Membrane↗
PubMed1999

Experimental evidence for hydrophobic matching and membrane-mediated interactions in lipid bilayers containing gramicidin.

Hydrophobic matching, in which transmembrane proteins cause the surrounding lipid bilayer to adjust its hydrocarbon thickness to match the length of the hydrophobic surface of the protein, is a commonly accepted idea in membrane biophysics. To test this idea, gramicidin (gD) was embedded in 1, 2-dilauroyl-sn-glycero-3-phosphocholine (DLPC) and 1, 2-myristoyl-sn-glycero-3-phosphocholine (DMPC) bilayers at the peptide/lipid molar ratio of 1:10. Circular dichroism (CD) was measured to ensure that the gramicidin was in the beta6.3 helix form. The bilayer thickness (the phosphate-to-phosphate distance, or PtP) was measured by x-ray lamellar diffraction. In the Lalpha phase near full hydration, PtP is 30.8 A for pure DLPC, 32.1 A for the DLPC/gD mixture, 35.3 A for pure DMPC, and 32.7 A for the DMPC/gD mixture. Gramicidin apparently stretches DLPC and thins DMPC toward a common thickness as expected by hydrophobic matching. Concurrently, gramicidin-gramicidin correlations were measured by x-ray in-plane scattering. In the fluid phase, the gramicidin-gramicidin nearest-neighbor separation is 26.8 A in DLPC, but shortens to 23.3 A in DMPC. These experiments confirm the conjecture that when proteins are embedded in a membrane, hydrophobic matching creates a strain field in the lipid bilayer that in turn gives rise to a membrane-mediated attractive potential between proteins.

Circular Dichroism↗
PubMed1999

Theoretical analysis of hydrophobic matching and membrane-mediated interactions in lipid bilayers containing gramicidin.

We present a quantitative analysis of the effects of hydrophobic matching and membrane-mediated protein-protein interactions exhibited by gramicidin embedded in dimyristoylphosphatidylcholine (DMPC) and dilauroylphosphatidylcholine (DLPC) bilayers (Harroun et al., 1999. Biophys. J. 76:937-945). Incorporating gramicidin, at 1:10 peptide/lipid molar ratio, decreases the phosphate-to-phosphate (PtP) peak separation in the DMPC bilayer from 35.3 A without gramicidin to 32.7 A. In contrast, the same molar ratio of gramicidin in DLPC increases the PtP from 30.8 A to 32.1 A. Concurrently, x-ray in-plane scattering showed that the most probable nearest-neighbor separation between gramicidin channels was 26.8 A in DLPC, but reduced to 23.3 A in DMPC. In this paper we review the idea of hydrophobic matching in which the lipid bilayer deforms to match the hydrophobic surface of the embedded proteins. We use a simple elasticity theory, including thickness compression, tension, and splay terms to describe the membrane deformation. The energy of membrane deformation is compared with the energy cost of hydrophobic mismatch. We discuss the boundary conditions between a gramicidin channel and the lipid bilayer. We used a numerical method to solve the problem of membrane deformation profile in the presence of a high density of gramicidin channels and ran computer simulations of 81 gramicidin channels to find the equilibrium distributions of the channels in the plane of the bilayer. The simulations contain four parameters: bilayer thickness compressibility 1/B, bilayer bending rigidity Kc, the channel-bilayer mismatch Do, and the slope of the interface at the lipid-protein boundary s. B, Kc, and Do were experimentally measured; the only free parameter is s. The value of s is determined by the requirement that the theory produces the experimental values of bilayer thinning by gramicidin and the shift in the peak position of the in-plane scattering due to membrane-mediated channel-channel interactions. We show that both hydrophobic matching and membrane-mediated interactions can be understood by the simple elasticity theory.

Biophysical Phenomena↗
PubMed1999

Does retrograde warm blood cardioplegic perfusion provide better protection of ischemic areas than antegrade warm blood cardioplegic perfusion? A magnetic resonance study in pig hearts.

OBJECTIVE: The purpose of this study was to determine whether retrograde continuous normothermic blood cardioplegic perfusion provides better protection to ischemic areas of the left and right ventricles than does antegrade continuous normothermic blood cardioplegic perfusion. Localized phosphorus 31 magnetic resonance spectroscopy was used to monitor the changes in energy metabolism and intracellular pH in the ventricles of pig hearts. METHODS: Ten isolated pig hearts received 20 minutes of antegrade continuous normothermic blood cardioplegic perfusion for collection of control (baseline) data, followed by 60 minutes of either antegrade continuous normothermic blood cardioplegic perfusion (n = 5) or retrograde continuous normothermic blood cardioplegic perfusion (n = 5) with occlusion of the left anterior descending and the right coronary arteries. The hearts were then subjected to antegrade continuous normothermic blood cardioplegic perfusion for 20 minutes. The perfusion pressures were maintained between 80 and 100 mm Hg and between 38 and 43 mm Hg during antegrade and retrograde continuous normothermic blood cardioplegic perfusions, respectively. Intracellular pH and creatine phosphate, inorganic phosphate, and adenosine triphosphate levels were measured continuously in each ventricle by means of localized phosphorus 31 magnetic resonance spectroscopy with 2 surface coils. RESULTS: Both antegrade and retrograde continuous normothermic blood cardioplegic perfusion resulted in a significant increase in inorganic phosphate level and decreases in creatine phosphate level, adenosine triphosphate level, and intracellular pH. No significant differences in these changes were observed between the two groups. The creatine phosphate and adenosine triphosphate levels were significantly lower in the right ventricle than in the left ventricle during retrograde continuous normothermic blood cardioplegic perfusion. On reperfusion, the inorganic phosphate level, creatine phosphate level, and intracellular pH recovered completely; however, no recovery in the adenosine triphosphate level was seen in the ventricles of either group. CONCLUSIONS: Retrograde continuous normothermic blood cardioplegic perfusion does not provide better protection to ischemic areas of the ventricles than does antegrade continuous normothermic blood cardioplegic perfusion under our experimental conditions.

Adenosine Triphosphate↗
PubMed1999

Human CD34+ hematopoietic cells transduced by retrovirus-mediated interferon alpha gene maintains regeneration capacity and engraftment in NOD/SCID mice.

To achieve long-term expression of human interferon alpha-5 (IFNalpha) gene in the bone marrow (BM) hematopoietic microenvironment, replication-deficient retroviral vector LSN-IFNalpha was used to deliver the IFNalpha gene into human BM CD34+ cells. After fibronectin-facilitated transduction, a fraction of CD34+ cells was plated in methylcellulose medium with or without G418 to assess transduction efficiency and the effect of IFNalpha gene transfer on colony formation. Colony-forming assay in the presence of G418 (400 microg/mL) revealed that 41% CFU-GM colonies are G418 resistant after infection with LSN-IFNalpha retrovirus. There was no significant difference in CFU-GM/BFU-E colony formation among IFNalpha gene-transduced CD34+ cells, control vector (LXSN) transduced-CD34+ cells and nontransduced CD34+ cells. Another portion of CD34+ cells was grown in liquid medium to measure IFNalpha production. RIA revealed that IFNalpha gene-transduced CD34+ cells produced 72.2 +/- 15.4 U/mL (10(6) cells/24 hours) of IFNalpha compared with 8.3 +/- 2.1 U/mL and 4.3 +/- 1.2 U/mL in LXSN-transduced or nontransduced CD34+ cells, respectively. The remaining portion of transduced CD34+ cells was transplanted into immunodeficient (NOD/SCID) mice to allow analysis of long-term expression of IFNalpha. Transplantation of 1x10(6) CD34+ cells into sublethally irradiated NOD/SCID mice showed that IFNalpha and neo(r) mRNA were detectable in engrafted mouse BM cells for up to 6 months. We conclude that continual local expression of IFNalpha in transduced CD34+ cells does not impair either CD34+ cell growth and differentiation or engraftment and long-term survival in NOD/SCID mice.

Animals↗
PubMed1999

A biomechanical study of median sternotomy closure techniques.

OBJECTIVE: Sternal dehiscence is a complication of median sternotomy incisions with high mortality and morbidity. Different techniques of sternal closure have been described. Rigid fixation of the sternum results in earlier union. We measured the rigidity of sternotomy fixation using a mechanical model in order to differentiate different techniques of sternal closure using their biomechanical characteristics. METHODS: We measured the force-displacement curves of six different fixation techniques using a metal sternal model using a computerized materials-testing machine. We tested straight wires (the most commonly used surgical technique), figure-of-8 wires, 'repair' technique (used when a wire breaks), Ethibond, Sterna-band and a 'multitwist' closure described for the first time. RESULTS: At 20 kg force, twisted wires used for sternotomy closures start to untwist. The most rigid closure was a multitwist closure that displaced only 0.37 mm at a force of 20 kg. Straight wires displaced 0.78 mm, figure-of-8 wires 1.20 mm, Sterna-band 1.37 mm, repair wires 5.08 mm, Ethibond 9.37 mm. The single factor Anova test for the rigidity of the different closures had P-values <0.0001. CONCLUSIONS: We applied a mathematical model to calculate chest wall forces during coughing, in order to determine the force placed upon a sternotomy closure. We conclude that severe coughing may cause wires to untwist. We discuss potential applications of different wire closures based on their characteristics.

Biomechanical Phenomena↗
PubMed1999

Inhibition of allergic inflammation in a murine model of asthma by expression of a dominant-negative mutant of GATA-3.

The cytokines IL-4, IL-5, and IL-13, secreted by Th2 cells, have distinct functions in the pathogenesis of asthma. We have previously shown that the transcription factor GATA-3 is expressed in Th2 but not Th1 cells. However, it was unclear whether GATA-3 controls the expression of all Th2 cytokines. Expression of a dominant-negative mutant of GATA-3 in mice in a T cell-specific fashion led to a reduction in the levels of all the Th2 cytokines IL-4, IL-5, and IL-13. Airway eosinophilia, mucus production, and IgE synthesis, all key features of asthma, were severely attenuated in the transgenic mice. Thus, targeting GATA-3 activity alone is sufficient to blunt Th2 responses in vivo, thereby establishing GATA-3 as a potential therapeutic target in the treatment of asthma and allergic diseases.

Aerosols↗
PubMed1999

Identification of the portal vein: wedge hepatic venography with CO2 or iodinated contrast medium.

RATIONALE AND OBJECTIVES: The purpose of this study was to evaluate the utility of CO2 versus iodinated contrast medium for wedge hepatic venography in identifying portal vein anatomy during transjugular intrahepatic portosystemic shunt (TIPS) procedures. MATERIALS AND METHODS: Wedge hepatic venograms obtained with CO2 or iodinated contrast medium and direct portograms of 43 patients undergoing TIPS procedures were analyzed retrospectively. Wedge venography was performed in 23 patients with CO2 and in 21 with iodinated contrast medium; direct portography was subsequently performed in 42 of 44 patients with iodinated contrast medium and in one with CO2. All cases were reviewed systematically to compare portal vein anatomy and completeness of anatomic identification between direct portography and wedge venography, and the results with CO2 were compared to those with iodinated contrast material. RESULTS: On the basis of opacification of the main portal trunk, branches, or both, the portal vein appearance (definition of the portal bifurcation) was good to excellent in 21 of 23 patients imaged with CO2 but in only two of 20 patients imaged with iodinated contrast medium. Wedge venograms agreed with direct portograms in 91% (21 of 23) of the CO2 cases and in 10% (two of 20) of the iodinated contrast medium cases. The two patients with poor opacification using CO2 had poor delineation of the main portal trunk, branches, and varices. TIPS could not be created in three patients, In two, abnormal morphology was identified at CO2 venography; in the third, wedge venography was not performed. CONCLUSION: Wedge hepatic venography with CO2 compared with iodinated contrast medium has a substantially higher likelihood of correctly and completely identifying the location and anatomy of the portal vein.

Adult↗