Biomedical subjects

Y Xu

Publications and source records attributed to Y Xu.

At least 343 records · Page 19Linked to original sources

PubMed2000

Troglitazone improves recovery of left ventricular function after regional ischemia in pigs.

BACKGROUND: This study determined whether treatment of normal (nondiabetic) pigs with the insulin-sensitizing agent troglitazone improves recovery of left ventricular (LV) function after acute ischemia and whether such effects are associated with altered myocardial substrate metabolism. METHODS AND RESULTS: Juvenile pigs (n=6) were treated with troglitazone (75 mg. kg(-1). d(-1) PO) for 8 weeks. Untreated pigs (n=8) served as controls. Under anesthetized, open-chest conditions, pigs underwent 90 minutes of moderate regional LV ischemia and 90 minutes of reperfusion. Regional LV function was assessed with subendocardial sonomicrometry crystals. Fasting plasma insulin and free fatty acid levels were lower in troglitazone-treated pigs than in untreated pigs, whereas blood glucose did not differ between groups. These findings suggest that treatment enhanced systemic insulin sensitivity. Baseline hemodynamics and regional LV function did not differ between groups. After ischemia and reperfusion, systolic function (external work) of the ischemic region recovered to 44+/-6% of baseline in troglitazone-treated pigs versus 18+/-6% of baseline in untreated pigs (P<0.05). Regional diastolic function (maximum rate of wall expansion) recovered to 78+/-7% of baseline in treated pigs versus 52+/-7% of baseline in untreated pigs (P<0.05). Recovery of global LV systolic and diastolic function was also significantly greater in treated pigs. Myocardial glucose uptake did not differ between groups under any condition; however, net myocardial lactate uptake after reperfusion was 7 times greater in troglitazone-treated pigs than in untreated pigs, suggesting that treatment enhanced myocardial carbohydrate oxidation after reperfusion. CONCLUSIONS: In nondiabetic pigs, chronic troglitazone treatment improves recovery of LV systolic and diastolic function after acute ischemia.

Animals↗

PubMed2000

The yeast mitochondrial citrate transport protein. Probing the roles of cysteines, Arg(181), and Arg(189) in transporter function.

Utilizing site-directed mutagenesis in combination with chemical modification of mutated residues, we have studied the roles of cysteine and arginine residues in the mitochondrial citrate transport protein (CTP) from Saccharomyces cerevisiae. Our strategy consisted of the sequential replacement of each of the four endogenous cysteine residues with Ser or in the case of Cys(73) with Val. Wild-type and mutated forms of the CTP were overexpressed in Escherichia coli, purified, and reconstituted in phospholipid vesicles. During the sequential replacement of each Cys, the effects of both hydrophilic and hydrophobic sulfhydryl reagents were examined. The data indicate that Cys(73) and Cys(256) are primarily responsible for inhibition of the wild-type CTP by hydrophilic sulfhydryl reagents. Experiments conducted with triple Cys replacement mutants (i.e. Cys(192) being the only remaining Cys) indicated that sulfhydryl reagents no longer inhibit but in fact stimulate CTP function 2-3-fold. Following the simultaneous replacement of all four endogenous Cys, the functional properties of the resulting Cys-less CTP were shown to be quite similar to those of the wild-type protein. Finally, utilizing the Cys-less CTP as a template, the roles of Arg(181) and Arg(189), two positively charged residues located within transmembrane domain IV, in CTP function were examined. Replacement of either residue with a Cys abolishes function, whereas replacement with a Lys or a Cys that is subsequently covalently modified with (2-aminoethyl)methanethiosulfonate hydrobromide, a reagent that restores positive charge at this site, supports CTP function. The results clearly show that positive charge at these two positions is essential for CTP function, although the chemistry of the guanidinium residue is not. Finally, these studies: (i) definitely demonstrate that Cys residues do not play an important role in the mechanism of the CTP; (ii) prove the utility of the Cys-less CTP for studying structure/function relationships within this metabolically important protein; and (iii) have led to the hypothesis that the polar face of alpha-helical transmembrane domain IV, within which Arg(181), Arg(189), and Cys(192) are located, constitutes an essential portion of the citrate translocation pathway through the membrane.

Amino Acid Sequence↗

PubMed2000

CpG DNA induces stronger immune responses with less toxicity than other adjuvants.

The ability to augment protective immune responses with minimal side effects is quintessential for a good adjuvant. This study has compared various adjuvants that are used in animal research (Freund's complete and incomplete adjuvants, Titermax Gold), are licensed for human use (alum), or are in clinical testing for humans (monophosphoryl lipid, CpG DNA), for their ability to augment humoral responses to a model antigen (hepatitis B surface antigen) and for the degree of damage they caused in the injected muscle. According to the data, the adjuvant combination CpG DNA+alum had the greatest potential to augment immune responses with minimal side effects at the injection site. Evaluation of antibody isotypes indicated Th2 responses (no IgG2a) with all adjuvants except monophosphoryl lipid and CpG DNA, which gave mixed Th1/Th2 responses (IgG1 and IgG2a). Strong Th1 responses (predominantly IgG2a) were obtained with combinations of CpG DNA with other adjuvants.

Adjuvants, Immunologic↗

PubMed2000

Salmonella pathogenicity island 2-dependent evasion of the phagocyte NADPH oxidase.

A type III protein secretion system encoded by Salmonella pathogenicity island 2 (SPI2) has been found to be required for virulence and survival within macrophages. Here, SPI2 was shown to allow Salmonella typhimurium to avoid NADPH oxidase-dependent killing by macrophages. The ability of SPI2-mutant bacteria to survive in macrophages and to cause lethal infection in mice was restored by abrogation of the NADPH oxidase-dependent respiratory burst. Ultrastructural and immunofluorescence microscopy demonstrated efficient localization of the NADPH oxidase in the proximity of vacuoles containing SPI2-mutant but not wild-type bacteria, suggesting that SPI2 interferes with trafficking of oxidase-containing vesicles to the phagosome.

Animals↗

PubMed2000

Effect of ADP and ionic strength on the kinetic and motile properties of recombinant mouse myosin V.

Mouse myosin V is a two-headed unconventional myosin with an extended neck that binds six calmodulins. Double-headed (heavy meromyosin-like) and single-headed (subfragment 1-like) fragments of mouse myosin V were expressed in Sf9 cells, and intact myosin V was purified from mouse brain. The actin-activated MgATPase of the tissue-purified myosin V, and its expressed fragments had a high V(max) and a low K(ATPase). Calcium regulated the MgATPase of intact myosin V but not of the fragments. Both the MgATPase activity and the in vitro motility were remarkably insensitive to ionic strength. Myosin V and its fragments translocated actin at very low myosin surface densities. ADP markedly inhibited the actin-activated MgATPase activity and the in vitro motility. ADP dissociated from myosin V subfragment 1 at a rate of about 11.5 s(-1) under conditions where the V(max) was 3.3 s(-1), indicating that, although not totally rate-limiting, ADP dissociation was close to the rate-limiting step. The high affinity for actin and the slow rate of ADP release helps the myosin head to remain attached to actin for a large fraction of each ATPase cycle and allows actin filaments to be moved by only a few myosin V molecules in vitro.

Actins↗

PubMed2000

Improvement of the enantioselectivity of lipase-catalyzed naproxen ester hydrolysis in organic solvent.

A method is presented to improve the enantioselectivity of lipase-catalyzed hydrolysis of naproxen methyl ester in water-saturated isooctane. It is shown that coupling of the enantioselective hydrolysis of Naproxen methyl ester with the photo-dissociation methanol leads to the photocatalytic conversion of methanol into water, by which the equilibrium constant (K) of the lipase-catalyzed hydrolysis was changed. The equilibrium yield and enantiomeric excess are increased. Because the lipase would not dissolve in the organic solvent, it was adsorbed on photocatalyst particles, which may facilitate the isolation of enzyme from reaction system.

Journal Article↗

PubMed2000

New structural motifs on the chymotrypsin fold and their potential roles in complement factor B.

Factor B and C2 are two central enzymes for complement activation. They are multidomain serine proteases and require cofactor binding for full expression of proteolytic activities. We present a 2.1 A crystal structure of the serine protease domain of factor B. It shows a number of structural motifs novel to the chymotrypsin fold, which by sequence homology are probably present in C2 as well. These motifs distribute characteristically on the protein surface. Six loops surround the active site, four of which shape substrate-binding pockets. Three loops next to the oxyanion hole, which typically mediate zymogen activation, are much shorter or absent. Three insertions including the linker to the preceding domain bulge from the side opposite to the active site. The catalytic triad and non-specific substrate-binding site display active conformations, but the oxyanion hole displays a zymogen-like conformation. The bottom of the S1 pocket has a negative charge at residue 226 instead of the typical 189 position. These unique structural features may play different roles in domain-domain interaction, cofactor binding and substrate binding.

Amino Acid Sequence↗

PubMed2000

Studies on the fluorescence reaction between nucleic acid and the complex of cobalt(II) with 5-(3-fluoro-4-chlorophenylazo)-8-sulfonamidoquinoline and its applications.

Based on the enhancement of the fluorescence intensity of the complex of 5-(3-fluoro-4-chlorophenylazo)-8-sulfonamidoquinoline (FCPBSQ) with Co(2+) by nucleic acid in the presence of Tween 80 and in the weakly basic medium, a fluorescence method for the determination of nucleic acid was proposed. The calibration graphs for the determination of denatured calf thymus DNA (ct DNA), fish sperm DNA (fs DNA), and yeast RNA (yt RNA) were obtained in concentration ranges 0.050-4.0, 0.10-3.0, and 0.050-3.0 microg/ml with limits of detection of 0.010, 0.030, and 0.020 microg/ml, respectively. The method has been satisfactorily used for the determination of DNA in wheat cell extraction and ct DNA, fs DNA, and yt RNA in synthetic samples. Investigations on the binding mode by the Scatchard plots method suggested that both intercalation and electrostatic binding modes existed in this system.

Animals↗

PubMed2000

Mutational analysis of the primary substrate specificity pocket of complement factor B. Asp(226) is a major structural determinant for p(1)-Arg binding.

Factor B is a serine protease, which despite its trypsin-like specificity has Asn instead of the typical Asp at the bottom of the S(1) pocket (position 189, chymotrypsinogen numbering). Asp residues are present at positions 187 and 226 and either one could conceivably provide the negative charge for binding the P(1)-Arg of the substrate. Determination of the crystal structure of the factor B serine protease domain has revealed that the side chain of Asp(226) is within the S(1) pocket, whereas Asp(187) is located outside the pocket. To investigate the possible role of these atypical structural features in substrate binding and catalysis, we constructed a panel of mutants of these residues. Replacement of Asp(187) caused moderate (50-60%) decrease in hemolytic activity, compared with wild type factor B, whereas replacement of Asn(189) resulted in more profound reductions (71-95%). Substitutions at these two positions did not significantly affect assembly of the alternative pathway C3 convertase. In contrast, elimination of the negative charge from Asp(226) completely abrogated hemolytic activity and also affected formation of the C3 convertase. Kinetic analyses of the hydrolysis of a P(1)-Arg containing thioester by selected mutants confirmed that residue Asp(226) is a primary structural determinant for P(1)-Arg binding and catalysis.

Amino Acid Sequence↗

PubMed2000

Rescue of defective T cell development and function in Atm-/- mice by a functional TCR alpha beta transgene.

The Atm-/- mice recapitulate most of the defects observed in ataxia-telangiectasia (A-T) patients, including a high incidence of lymphoid tumors and immune defects characterized by defective T cell differentiation, thymus hypoplasia, and defective T-dependent immune responses. To understand the basis of the T cell developmental defects in Atm-/- mice, a functional TCR alpha beta transgene was introduced into these mutant mice. Analysis of the Atm-/-TCR alpha beta+ mice indicated that the transgenic TCR alpha beta can rescue the defective T cell differentiation and partially rescue the thymus hypoplasia in Atm-/- mice, indicating that thymocyte positive selection is normal in the Atm-/- mice. In addition, cell cycle analysis of the thymocytes derived from Atm-/-TCR alpha beta+ and control mice suggested that Atm is involved in the thymocyte expansion. Finally, evaluation of the T-dependent immune responses in Atm-/-TCR alpha beta+ mice indicated that Atm is dispensable for normal T cell function. Therefore, the defective T-dependent immune responses in Atm-/- mice must be secondary to greatly reduced T cell numbers in these mutant mice.

Animals↗

PubMed2000

Asynchronous activation of 10 muscle-specific protein (MSP) genes during zebrafish somitogenesis.

In the present study, 10 zebrafish cDNA clones coding for muscle-specific proteins (MSPs) were characterized and most of them encode fast skeletal muscle isoforms. They are skeletal muscle alpha-actin (acta1), fast skeletal muscle a-tropomyosin (tpma), fast skeletal muscle troponin C (tnnc), fast skeletal muscle troponin T (tnnt), fast skeletal muscle myosin heavy chain (myhz1), fast skeletal muscle myosin light chain 2 (mylz2), fast skeletal muscle myosin light chain 3 (mylz3), muscle creatine kinase (ckm), parvalbumin (pvalb), and desmin (desm). Using these cDNA probes, their expression patterns in developing embryos and adults were compared by Northern blot hybridization and whole-mount in situ hybridization. All of the 10 genes are expressed in both embryos and adult fish, and the expression is highly abundant in skeletal muscle. Among them, acta1, tpma, tnnc, tnnt, myhz1, mylz2, mylz3 and pvalb, are expressed specifically in fast skeletal muscle while ckm and desm are expressed in both fast and slow skeletal muscles. In addition, tpma, ckm, and desm are also expressed in the heart. Ontogenetically, the onset of expression of these MSP genes in zebrafish skeletal muscle varies and the expression occurs rostral-caudally in developing somites. Shortly after the expression of myoD, desm is the first to be activated at approximately 9 hpf, followed by tpma (approximately 10 hpf), tnnc (approximately 12 hpf), acta1 (approximately 12 hpf), ckm (approximately 14 hpf), myhz1 (approximately 14 hpf), mylz2 (approximately 16 hpf), mylz3 (approximately 16.5 hpf), tnnt (approximately 16.5 hpf), and pvalb (approximately 16.5 hpf). At later stages (after 48 hpf), these MSP genes are also expressed in fin buds and head muscles including eye, jaw, and gill muscles. Thus, our experiment demonstrated the order of expression of the 10 MSP genes, which may reflect the sequence of muscle filament assembly. In spite of the asynchrony in activation of these MSP genes, the timing of expression for each individual MSP gene appears to be synchronous to somite development as each somite has an identical timetable to express the set of MSP genes.

Actins↗

PubMed2000

Reduced slab boundary artifact in multi-slab 3D fast spin-echo imaging.

The three-dimensional multi-slab fast spin-echo (3DFSE) sequence is a time-efficient technique for volume scanning which provides images with a good signal-to-noise ratio, adjustable contrast weighting, and high spatial resolution. Unfortunately, it suffers from slice-to-slice amplitude variation associated with imperfect slab definition. This slab boundary artifact becomes especially apparent when multiplanner reformatting is used to create alternate anatomical views. The shifted interleaved multi-volume acquisition (SIMVA) described here suppresses slab boundary artifact in image space. It displaces each slab (RF excitation) position incrementally along the slice (z) axis, in coordination with the primary phase encode step, so that the slab boundary artifact is converted into a correctable amplitude modulation in the primary phase encode direction (k(Y)-axis). After the Fourier transform in primary phase encoding, the slab boundary artifact is mapped into a different and less severe artifact on a different spatial axis. Preliminary measurements show that SIMVA reduces the slab boundary artifact by an order of magnitude in multiplanar reformatted views. Magn Reson Med 44:269-276, 2000.

Artifacts↗

PubMed2000

Real-time quantitative PCR for retrovirus-like particle quantification in CHO cell culture.

Chinese hamster ovary (CHO) cells have been widely used to manufacture recombinant proteins intended for human therapeutic uses. Retrovirus-like particles, which are apparently defective and non-infectious, have been detected in all CHO cells by electron microscopy (EM). To assure viral safety of CHO cell-derived biologicals, quantification of retrovirus-like particles in production cell culture and demonstration of sufficient elimination of such retrovirus-like particles by the down-stream purification process are required for product market registration worldwide. EM, with a detection limit of 1x10(6) particles/ml, is the standard retrovirus-like particle quantification method. The whole process, which requires a large amount of sample (3-6 litres), is labour intensive, time consuming, expensive, and subject to significant assay variability. In this paper, a novel real-time quantitative PCR assay (TaqMan assay) has been developed for the quantification of retrovirus-like particles. Each retrovirus particle contains two copies of the viral genomic particle RNA (pRNA) molecule. Therefore, quantification of retrovirus particles can be achieved by quantifying the pRNA copy number, i.e. every two copies of retroviral pRNA is equivalent to one retrovirus-like particle. The TaqMan assay takes advantage of the 5'-->3' exonuclease activity of Taq DNA polymerase and utilizes the PRISM 7700 Sequence Detection System of PE Applied Biosystems (Foster City, CA, U.S.A.) for automated pRNA quantification through a dual-labelled fluorogenic probe. The TaqMan quantification technique is highly comparable to the EM analysis. In addition, it offers significant advantages over the EM analysis, such as a higher sensitivity of less than 600 particles/ml, greater accuracy and reliability, higher sample throughput, more flexibility and lower cost. Therefore, the TaqMan assay should be used as a substitute for EM analysis for retrovirus-like particle quantification in CHO cell-based production system.

Animals↗

PubMed2000

Perception of dynamic acoustic patterns by an individual with unilateral verbal auditory agnosia.

Previous studies have found that subjects diagnosed with verbal auditory agnosia (VAA) from bilateral brain lesions may experience difficulties at the prephonemic level of acoustic processing. In this case study, we administered a series of speech and nonspeech discrimination tests to an individual with unilateral VAA as a result of left-temporal-lobe damage. The results indicated that the subject's ability to perceive steady-state acoustic stimuli was relatively intact but his ability to perceive dynamic stimuli was drastically reduced. We conclude that this particular aspect of acoustic processing may be a major contributing factor that disables speech perception in subjects with unilateral VAA.

Adult↗

PubMed2000

Distribution, transformation, and long-term accumulation of polychlorinated dibenzo-p-dioxins and dibenzofurans in different tissues of fish and piscivorous birds.

The present study monitored 10-year-old fish and piscivorous birds from sites contaminated for many years. The data reflected the results of actual, long-term environmental exposures. The results demonstrate that different tissues of fish have quite different concentrations of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/F). The concentration order of PCDD/F within fish is liver congruent with egg congruent with intestine>kidney congruent with heart>gill congruent with bladder>muscle>brain. The concentration order of PCDD/F within piscivorous birds was liver>egg congruent with heart>muscle congruent with stomach>brain. The results obtained also demonstrate that the accumulation patterns of piscivorous birds and fish are quite different. The tissues of fish and piscivorous birds have different capacities for bioaccumulation and biotransformation of PCDD/F; variable proportions of TEQs were also found throughout their bodies. In fish, toxic equivalency quotient (TEQ): PCDD/F ratios in various tissues ranged from 0.01 to 0.07, whereas in birds the ratios ranged from 0.07 to 0.43. If the concentrations are normalized with lipid content, the results vary less. The effect of different lipid properties is obvious in the case of brain tissue, which is richer in phospholipids.

Animals↗

PubMed2000

The change of interleukin-6 and tumor necrosis factor in patients with obstructive sleep apnea syndrome.

The levels of lipopolysaccharide (LPS)-induced interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) expression in culture of peripheral blood mononuclear cells (PBMC) and the plasma levels of IL-6 and TNF-alpha in the patients with obstructive sleep apnea syndrome (OSAS) were measured and the relationship between OSAS and IL-6 or TNF-alpha expression studied. Both IL-6 and TNF-alpha were detected by using ELISA in 22 patients with OSAS and 16 normal controls. The levels of LPS-induced IL-6 (787.82 +/- 151.97 pg/ml) and TNF-alpha (4165.45 +/- 1501.43 pg/ml) expression in the supernatant of the culture of PBMC and plasma level of IL-6 (50.67 +/- 4.70 pg/ml) and TNF-alpha (299.09 +/- 43.57 pg/ml) in the patients with OSAS were significantly higher than those in the normal controls (in the supernatant of the culture of PBMC: 562.69 +/- 197.54 pg/ml and 1596.25 +/- 403.08 pg/ml respectively; in the plasma; 12.69 +/- 2.75 pg/ml and 101.88 +/- 21.27 pg/ml respectively). There were significantly positive correlation between the levels of IL-6 and TNF-alpha and the percentage of time of apnea and hyponea, as well as the percentage of time spending at SaO2 below 90% in the total sleep time. It was concluded that LPS-induced IL-6 and TNF-alpha levels as well as plasma IL-6 and TNF-alpha levels in the patients with OSAS were up-regulated, which may be associated with the pathogenesis of OSAS.

Adult↗

PubMed2000

Effects of intrinsic nitric oxide on the expression of interleukin-4 and IFN-gamma mRNA in the bronchial and lung tissues of sensitized rats.

To investigate the effects of intrinsic nitric oxide (NO) on the expression of interleukin-4 (IL-4) mRNA and interferon-gamma (IFN-gamma) mRNA in the airway inflammation of asthma, the rat models of asthmatic inflammation were established by sensitizing and then challenging the animals with ovalbumin. The 24 animals were randomly divided into control group, sensitized group, sensitized and L-Arg-treated group as well as L-NAME-treated group equally. By using in situ hybridization combined with compute physiological quantitative imaging analysis techniques, the influence of intrinsic NO on the expression of IL-4 mRNA and IFN-gamma mRNA in the airway inflammatory cells was observed. In situ hybridization study demonstrated that IL-4 mRNA expression was obviously increased as compared with that in the control group, mainly distributed in the inflammatory cells in the submucous of airways in the sensitized group. The increase of intensity of IL-4 mRNA expression was positively correlated with the numbers of eosinophil (Eos) and lymphocyte (both with P < 0.05) in the sensitized group. There was no statistically difference in IFN-gamma expression between the control group and the sensitized group. Imaging analysis showed that L-NAME could inhibit the expression of IL-4 mRNA (P < 0.05) and increase the expression of IFN-gamma mRNA (P < 0.05), while L-Arg could increase the expression of IL-4 mRNA in inflammatory cells (P < 0.05). It was indicated that a suitable levels of intrinsic NO can influence the expression of IL-4 mRNA of Th2 lymphocytes and the expression of IFN-gamma mRNA of Th1 lymphocytes and in turn, promote the development of asthmatic airway inflammation.

Animals↗

PubMed2000

The diagnosis value and its implication of impulse oscillometry in obstructive sleep apnea syndrome patients.

The change of measurements of impulse oscillometry (IOS) in obstructive sleep apnea syndrome (OSAS) patients and its mechanism were observed. The respiratory impedance was measured by using IOS technique and polysomnography (PSG) was monitored synchronously in 36 OSAS patients, 14 patients with chronic obstructive pulmonary disease (COPD) and 12 normal controls. Results showed that R20 in OSAS group was significantly higher than in COPD group and control group (P < 0.01). R5-R20 in OSAS group was lower than that in COPD group, but significantly higher than that in control group (P < 0.01). The levels of R20 and R5-R20 were positively correlated with severity degree of the disease. In addition, apnea-hyponea index (AHI) was positively correlated with R5 and R20 with the correlation index (r)being 0.66 and 0.86 respectively. The lowest SO2 was negatively correlated with R5 and R5-R20, with r being -0.66 and -0.79 respectively. The mean SO2 was negatively correlated with R5 and R5-R20 with r being -0.81 and -0.69 respectively. IOS technique could be used as a valuable tool for assessing the degree of upper airway obstruction in the patients with OSAS, and could help to explore its pathological mechanism.

Adult↗