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Biomedical subjects

D Song

Publications and source records attributed to D Song.

At least 109 records · Page 6Linked to original sources
PubMed1997

Primary structure and functional expression of heavy- and light-chain variable region genes of a monoclonal antibody specific for human fibrin.

The immunoglobulin heavy- and light-chain variable region (VH and VK) genes were isolated from 8E5 hybridoma cells, which secreted monoclonal antibody against human fibrin by RT-PCR. An expression vector pOPE51-8E5 was constructed for the recombinant VH-VK scFv expression. The primary sequence of the variable regions was determined. Expression product was found in the periplasmic space and inclusion bodies by SDS-PAGE and immunoblotting. It was a 30 KDa single chain fragment (scFv) with the antigen-binding specificity of the parental monoclonal antibody. A light chain shuffling with an unspecific VL did not result in a loss of fibrin binding specificity.

Amino Acid Sequence↗
PubMed1997

Optimal dose of nicardipine for maintenance of hemodynamic stability after tracheal intubation and skin incision.

UNLABELLED: To determine the optimal dose of nicardipine (N) for maintenance of hemodynamic stability during the postinduction period, we designed a randomized, double-blind, placebo-controlled, dose-ranging study using four different doses of N administered after a standardized anesthetic induction sequence. A total of 106 patients were assigned to one of the following treatment groups: saline (control), N 0.5 mg (N0.5), N 1 mg (N1), N 2 mg (N2), and N 4 mg (N4). The study medication was administered intravenously (I.V.) in 2.5 mL of saline over 30 s 2 min before laryngoscopy. Mean arterial pressure (MAP) and heart rate (HR) were recorded at 1-min intervals for 15 min after tracheal intubation and for 5 min after skin incision. After intubation, the peak MAP values differed from the preinduction baseline MAP values by 21% +/- 20%, 9% +/- 12%, 1% +/- 13%, -10% +/- 12%, and -15% +/- 13% (mean +/- SD) in the control, N0.5, N1, N2, and N4 groups, respectively. However, the percent change in the pre- to postintubation MAP values (37% to 47%) was similar in all five groups. The highest postintubation HR values were recorded in the N4 group (P < 0.05 versus the other groups). However, the increases in MAP values after skin incision were the least in the N4 group. In conclusion, N1 I.V., administered 2 min before laryngoscopy provides optimal control of arterial blood pressure during the postinduction period. IMPLICATIONS: Acute increases in blood pressure during anesthesia are undesirable in patients with preexisting cardiovascular diseases. This double-blind study found that the calcium-channel blocker, nicardipine, 1 mg intravenously 2 min before tracheal intubation maintained hemodynamic stability during the intraoperative period.

Adult↗
PubMed1997

Titration of volatile anesthetics using bispectral index facilitates recovery after ambulatory anesthesia.

BACKGROUND: The bispectral (BIS) index has previously been shown to be a quantifiable measure of the sedative and hypnotic effects of anesthetic drugs. This study was designed to assess the effect of BIS monitoring on the utilization of volatile anesthetics and their recovery profiles after ambulatory surgery. METHODS: Sixty consenting women undergoing outpatient laparoscopic tubal ligation procedures were randomly assigned to one of four treatment groups. After a standardized induction, anesthesia was maintained with either desflurane (Groups I and II) or sevoflurane (Groups III and IV) in combination with nitrous oxide, 65%, and fentanyl. In the control groups (Groups I and III), the anesthesiologists were blinded to the BIS value, and the volatile anesthetics were administered according to standard clinical practice. In Groups II and IV, the volatile anesthetics were titrated to maintain the BIS value at 60. The volatile anesthetic usage and the times from discontinuation of anesthesia to verbal response, orientation, and home-readiness were recorded. RESULTS: During the maintenance period, the BIS values were significantly lower in the control groups (mean, 42) compared with the BIS-titrated groups (mean, 60). The volatile anesthetic usage in the BIS-titrated groups was 30-38% lower (P < 0.05) compared with the control groups. Similarly, the times to verbal responsiveness were 30-55% shorter in the BIS-titrated (vs. control) groups. CONCLUSIONS: Titrating desflurane and sevoflurane using the BIS monitor decreased their utilization and contributed to a faster emergence from anesthesia in outpatients undergoing laparoscopic tubal ligation procedures.

Adult↗
PubMed1997

Timing of lymphocyte activation in neonates infected with human immunodeficiency virus.

Human immunodeficiency virus (HIV) infection in children is associated with qualitative and quantitative changes in the peripheral lymphocyte surface phenotype beyond the normal maturational changes. Neonates, however, have been reported to have a delayed immune response to HIV compared to HIV-infected adults. We prospectively performed immunophenotyping of T lymphocytes by three-color immunofluorescent labeling and laser flow cytometry to determine the timing of phenotypic alterations in 112 neonates born to HIV-infected mothers. Serial testing was performed at birth (cord blood) and at 2, 6, and 12 weeks of age. Data were divided retrospectively for analysis into those for HIV-infected (n = 14) infants and those for exposed, uninfected infants. Our results show that both infected and uninfected infants had a decline in the percentages and numbers of CD4 cells beginning at 2 weeks of age but that the decline was greater in the HIV-infected group. The activation and differentiation of CD8 T cells in HIV+ infants were shown by a significant increase in CD45RA- CD45RO+ CD8+ cells by 6 weeks of age and by increases in CD8+ S6F1+ CD3+ cells and HLA-DR+ CD38+ CD8+ cells by 2 weeks of age. These results indicate that HIV-infected neonates show alterations in T-cell phenotype reflecting those reported for older HIV-infected children. Most importantly, neonatal T cells are able to respond to HIV within the first weeks of life.

Antigens, CD↗
PubMed1997

[Cloning, expression and purification of the chaperonin GroESL in Escherichia coli].

The DNA fragment encoding the molecular chaperion GroESL was subcloned into high-expression vector pKC220, and the GroESL were high expressed in the E. coli strain harboring the recombinant plasmid by high temperature induction. The amount of the expressed GroEL and GroES protein were about 40% and 15% of the total cellular proteins, respectively. These two subunits were both purified from E. coli by (NH4)2SO4 salt out DEAE-52 chromography and Sephadex G-50 chromography.

Bacterial Proteins↗
PubMed1997

[Cloning and expression of heavy- and light-chain variable region genes of monoclonal antibody specific for human fibrin].

OBJECTIVE: To obtain the minimum molecule having activity to bind human fibrin. METHODS: The immunoglobulin heavy- and light-chain variable region (VH and Vkappa) genes were isolated from 8E5 hybridoma cells, which secreted a monoclonal antibody against human fibrin, by RT-PCR. An expression vector pOPE51-8E5 was constructed for the recombinant VH-Vkappa expression. The transformed E. coli JM 109 cells were propagated and induced by IPTG. RESULTS AND CONCLUSION: Expression product was found in the periplasmic space and inclusion bodies by SDS-PAGE and immunobloting. It was a 30000 single chain fragment (scFv) with antigen-binding specificity. The yield of the scFv was 28.9% of the total bacterial proteins.

Animals↗
PubMed1997

[Analysis of risk factors for infant deaths in urban and rural areas of Shandong Province].

Data of infant deaths in 1984 and 1994 were collected from the disease surveillance system in Shandong Province and 13 risk factors contributing infant death were analyzed with unconditional logistic regression to reveal their change in urban and rural areas in recent ten years. Results showed that risk factors for infant death in urban areas were birth weight, mother's age of childbirth, length of gestation and maternal health status, with comprehensive population attributable risk proportions (PARP) of 92.24 and 93.81 percent in 1984 and 1994, respectively. In addition to the risk factors mentioned above in urban areas, infant death in rural areas was also affected by ways of delivery and unhealthy events during pregnancy, with a comprehensive PARP of 98.07 percent in 1994. Risks for infant death delivered by family midwives reduced greatly in recent ten years, with an odd ratio (OR) of 4.145 in 1984, but it could not be entered a multivariate regression model in 1994.

Cause of Death↗
PubMed1997

Bladder tissue pharmacokinetics and antitumor effect of intravesical 5-fluorouridine.

The present study evaluates whether intravesical 5-fluorouridine (FUR), a potent fluorinated pyrimidine, is effective against bladder cancer. The tissue and plasma pharmacokinetics of i.v. and intravesical FUR were studied in dogs to determine the tissue targeting advantage by the intravesical route. The i.v. study used a bolus FUR dose of 4 mg/kg, which is tolerated in humans. The disposition of FUR was biphasic, with a peak concentration of 8.8 microgram/ml and a clearance of 127 ml/min/kg. 5-Fluorouracil was the major circulating metabolite, reaching a peak concentration of 3.2 microgram/ml. In the intravesical study, FUR (approximately 2 mg/kg in 20 ml of water) was instilled in the dog bladder. At the end of the 2-h treatment, FUR concentration in urine decreased by about 40%, due mainly to dilution by residual and newly produced urine. The concentration at the interface between urothelium and lamina propria was 14 microgram/g, or approximately 2% of the urine concentration, and declined logarithmically to 2 microgram/g in the deep muscles. The concentrations of FUR and 5-fluorouracil in plasma were below the assay detection limit of 20 ng/ml, or > 200-fold lower than the concentration after the i.v. dose (adjusted to the difference in the i.v. and intravesical dose). These data indicate a > 200-fold advantage in the reduction of systemic exposure by the intravesical route. To determine whether the achievable tissue concentrations of FUR produced significant antitumor activity, we studied the effect of FUR against human bladder tumors maintained as 3-dimensional histocultures. The FUR concentrations (IC50s) required to produce 50% inhibition of DNA precursor ([3H]thymidine or bromodeoxyuridine) incorporation in human superficial bladder tumors (i.e., Ta and T1 tumors, n = 4) and muscle-invading tumors (i.e., T3 and T4 tumors, n = 4) were 9 and 22 microgram/ml, respectively. In conclusion, intravesical FUR therapy delivers effective drug concentration to superficial bladder tissues without resulting in appreciable systemic blood concentration. We propose that intravesical FUR represents a potentially effective treatment against superficial bladder cancer.

Administration, Intravesical↗
PubMed1996

Release of the excitotoxic amino acids, glutamate and aspartate, from the isolated ischemic/anoxic rat heart.

An isolated rat Langendorff heart preparation has been developed as a model in which to study the release of glutamate, aspartate and other amino acids during ischemia, anoxia and hypoglycemia. 15 min periods of ischemia resulted in large increases in perfusate levels of glutamate, aspartate, glycine, phosphoethanolamine, serine, alanine, taurine and glutamine. Amino acid levels returned towards pre-ischemic levels in subsequent perfusate collections. Anoxia (15 min duration) increased perfusate levels of most of the measured amino acids, with glutamate and aspartate being particularly affected. In contrast to ischemia, glutamate and aspartate levels declined slowly following reoxygenation. Hypoglycemia (15 min) resulted in small but significantly elevated levels of glutamate and glycine in heart perfusates. As the effects of ischemia or anoxia on glutamate and aspartate release from the heart appear to be comparable to those observed in the brain, it is proposed that the heart preparation may be a suitable model in which to study the ischemia-evoked release of these amino acids in the absence of complications arising from their depolarizing and excitotoxic actions on central neurons.

Animals↗
PubMed1996

Quantitative determination of clonazepam in plasma by gas chromatography-negative ion chemical ionization mass spectrometry.

A gas chromatographic-negative ion chemical ionization mass spectrometric (GC-NCI-MS) method for the quantitative analysis of clonazepam in human plasma is described. Clonazepam (M(r) = 315) was derivatized by N,O-bis-(trimethylsilyl)trifluoroacetamide with 1% trimethylchlorosilane. A pre-conditioning procedure involving injection of a silyl-8 and ethyl acetate extraction solution from plasma reduces the interaction between clonazepam-TMS and the analytical system. The routine limit of quantification was set to be 0.25 ng/ml with an injection volume of 2 microliters and a sample volume of 1 ml. The signal-to-noise ratio was greater than five. The detection limit for clonazepam can reach 0.1 ng/ml. The isotope clonazepam-d5 was used as the internal standard.

Anticonvulsants↗
PubMed1996

Localization of yeast RNA polymerase I core subunits by immunoelectron microscopy.

Immunoelectron microscopy was used to determine the spatial organization of the yeast RNA polymerase I core subunits on a three-dimensional model of the enzyme. Images of antibody-labeled enzymes were compared with the native enzyme to determine the localization of the antibody binding site on the surface of the model. Monoclonal antibodies were used as probes to identify the two largest subunits homologous to the bacterial beta and beta' subunits. The epitopes for the two monoclonal antibodies were mapped using subunit-specific phage display libraries, thus allowing a direct correlation of the structural data with functional information on conserved sequence elements. An epitope close to conserved region C of the beta-like subunit is located at the base of the finger-like domain, whereas a sequence between conserved regions C and D of the beta'-like subunit is located in the apical region of the enzyme. Polyclonal antibodies outlined the alpha-like subunit AC40 and subunit AC19 which were found co-localized also in the apical region of the enzyme. The spatial location of the subunits is correlated with their biological activity and the inhibitory effect of the antibodies.

Amino Acid Sequence↗
PubMed1996

The effect of hypoxia and catecholamines on regional expression of heat-shock protein-72 mRNA in neonatal piglet brain.

The present study has shown that hypoxia leads to expression of heat-shock protein in the brain of newborn piglets and this process is almost completely abolished by depletion of catecholamines prior to the hypoxic episode. The piglets were anesthetized and mechanically ventilated. One hour of hypoxia was generated by decreasing the oxygen fraction in the inspired gas (FiO2) from 22% to 6%-10%. FiO2 was then returned to the control value for a period of 2 h. Following the 2 h of reoxygenation, regional expression of the 72-kDa heat-shock protein (hsp72) mRNA was determined using in situ hybridization and autoradiography. The hypoxic insult (cortical pO2 = 3-10 mmHg) induced expression of hsp72 mRNA in regions of both white and gray matter, with strong expression occurring in the cerebral cortex of individual animals. Depleting the brain of catecholamines prior to hypoxia, by treating the animals with alpha-methyl-p-tyrosine (AMT), resulted in a major change in the hsp72 mRNA expression. In the catecholamine depleted group of animals, the intensity of hsp72 mRNA expression was greatly decreased or almost completely abolished relative to the nondepleted hypoxic group. These results suggest that the catecholamines play a significant role in the expression of the hsp72 gene in response to hypoxic insult in neonatal brain.

Animals↗
PubMed1996

Escherichia coli diacylglycerol kinase is an alpha-helical polytopic membrane protein and can spontaneously insert into preformed lipid vesicles.

Escherichia coli diacylglycerol kinase (DAGK) is a 13.2 kDa enzyme which spans the cytoplasmic membrane three times. Functional DAGK was purified to homogeneity using a polyhistidine tag and Ni(II)-chelate chromatography. Transmission Fourier transform infrared spectroscopy (FT-IR) of DAGK in phosphatidylcholine multilayers led to the conclusion that > or = 90 of DAGK's native 121 residues are alpha-helical, consistent with a model in which DAGK consists of two amphipathic alpha-helices and three transmembrane helices. Polarized attenuated total reflection FT-IR studies of DAGK in oriented multilamellae yielded data consistent with a topological arrangement in which the three transmembrane helices are well-aligned with the bilayer normal while the two amphipathic helices are approximately parallel with the membrane plane. The ability of DAGK to spontaneously insert into preformed lipid vesicles was examined using a novel assay system involving DAGK-catalyzed phosphorylation of a fluorescently tagged diacylglycerol. When micellar DAGK is diluted into L alpha-phase vesicles spontaneous insertion of the enzyme is fairly efficient (ca. 30%). DAGK refolding and insertion from delipidated urea-solubilized DAGK into lipid vesicles is also modestly efficient (3.8 +/- 2.1%) above the gel to liquid crystalline phase transition temperature. The insertion studies indicate that the difference in energy barriers (delta delta G++) between pathways leading to catalytically productive folding and insertion of DAGK relative to unproductive pathways is < 4 kcal/mol. However, additional studies carried out with mutant forms of DAGK indicated that the differences between refolding/insertion pathways for DAGK in vivo and in vitro can be significant.

Amino Acid Sequence↗
PubMed1996

Inhibition of tyrosine phosphorylation attenuates amino acid neurotransmitter release from the ischemic/reperfused rat cerebral cortex.

Protein tyrosine phosphorylation plays an important role in neuronal function. In this study we have examined the effects of inhibition of tyrosine phosphorylation on the extracellular levels of four neurotransmitter amino acids (aspartate, glutamate, gamma-aminobutyric acid (GABA) and glycine) and of the non-transmitter amino acid phosphoethanolamine during cerebral ischemia and reperfusion in a rat four vessel occlusion model. In comparison with the control group, the tyrosine kinase inhibitor genistein significantly depressed ischemia/reperfusion-evoked efflux of these amino acids, with the exception of GABA, into cerebral cortical superfusates. GABA efflux was non-significantly reduced. These results suggest that tyrosine phosphorylation is involved in the ischemia-evoked efflux of amino acids into the extracellular milieu, likely as a consequence of the phosphorylation of microtubule-associated protein kinase (MAP kinase) and downstream activation of PLA2 in the plasma membrane. Amino acid efflux would occur, in part, as a consequence of the ensuing disruption of plasma membrane integrity and leakage of cytoplasmic constituents along their concentration gradients.

Amino Acids↗
PubMed1996

Effect of hypoxia and reoxygenation on regional activity of nitric oxide synthase in brain of newborn piglets.

The activity of nitric oxide synthase (NOS) was measured in homogenates from cortex, striatum, hippocampus, cerebellum, pons, thalamus and midbrain of the brain of newborn piglets and the effects of hypoxia and posthypoxic period on this activity was evaluated. The control activities were 19.7, 31.5, 26.8, 16.7, 33.6, 19.3 and 39.4 pmol/mg protein per min, respectively. A 1 h period of hypoxia (an FiO2 of 7%) resulted in statistically significant decreases in the activity of NOS in every region of the brain except for the cortex, where the activity was not significantly altered compared to control. By 2 h of reoxygenation following such a hypoxic episode, the NOS activities increased to above control levels in all regions of the brain, but this increase was statistically significant compared to control only in thalamus. Since hypoxia induced the greatest decrease in NOS activity in the cerebellum, the kinetic constants of the enzyme were measured in homogenates from this region of brain. The decreased activity following the hypoxic episode was associated with an approximately four-fold increase in the apparent affinity (KM) for arginine with no significant change in the maximal rate of reaction (Vmax). The decrease in NOS activity subsequent to a hypoxic episode may contribute to the disturbances in cellular metabolism in the immature brain induced by episodes of hypoxia-reoxygenation.

Animals↗
PubMed1996

Direct injection isocratic high-performance liquid chromatographic analysis of mitomycin C in plasma.

A direct injection high-performance liquid chromatography method is described for the determination of mitomycin C (MMC) in human plasma. The stationary phase consisted of hydrophilic and hydrophobic functional groups covalently bound to silicone-coated silica beads (CAPCELL PAK MF Ph-1, 150x4.6 mm I.D., 5 microns). A mobile phase using 100% water gave a better separation of MMC from endogenous interferences as compared to a mobile phase with 12.5% acetonitrile and 2.5 mM phosphate buffer (pH 6.9). Using water as the eluent (1 ml/min) and UV detection at 365 nm, MMC was found to elute at 5.0 min with a peak width of 0.3 min, whereas endogenous interferences eluted before 3 min. Total assay time per sample was 6 min. Internal standard was not required because the recovery of MMC was nearly complete, about 90% from 20 to 5000 ng/ml. The standard curve was linear from 20 to 5000 ng/ml in plasma, and the intra-and inter-day variation was between 3 to 6%. The lower detection limit was 5 ng/ml with a 25 microliters sample, which represents a two- to four-fold improvement over the 10 ng/ml detection limit by previous methods using liquid-liquid extraction and comparable sample size. The simplicity of this method, i.e., no sample extraction, no internal standard, 100% aqueous mobile phase, isocratic elution and short analysis time (6 min/sample), makes it suitable for large scale routine sample analysis, whereas its small sample volume requirement and high sensitivity are useful for pharmacokinetic studies in small animals where limited sample is available.

Antibiotics, Antineoplastic↗
PubMed1996

Binding of taxol to plastic and glass containers and protein under in vitro conditions.

This study examined the stability and protein binding of taxol in aqueous solution and tissue culture medium. After storage for 19 h in 1% methanolic aqueous solution, the concentration of taxol declined to about 40% in 1.5 mL glass vials, 67% in 1.5 mL polypropylene tubes, and 55% in 1.5 mL siliconized polypropylene tubes. There was no difference in the decline at two initial concentrations of 0.18 and 1.8 micrograms/mL. The concentration declines were biphasic; the initial phase showed a half-life of 1-2 min and the second phase showed a half-life of 30-350 min. A methanol wash recovered 0%, 16%, and 31% of taxol from glass vials and unsiliconized and siliconized polypropylene tubes, respectively. In culture medium without fetal bovine serum, the concentration of taxol stored in polystyrene tissue culture plates declined to 73% after 24 h, whereas no concentration decline was observed in the presence of 9% fetal bovine serum (FBS). Protein binding of taxol in FBS-containing culture medium was saturable, varying from 79% at 0.5 microgram/mL to 20% at 15 micrograms/mL. These data indicated (a) rapid and nonspecific adsorption of taxol to plastic and glass surfaces, (b) stabilization of taxol in culture medium by FBS, and (c) saturable protein binding of taxol in culture medium.

Animals↗