Biomedical subjects

C H Lin

Publications and source records attributed to C H Lin.

At least 37 records · Page 2Linked to original sources

PubMed1997

Making chemistry selectable by linking it to infectivity.

The link between recognition and replication is fundamental to the operation of the immune system. In recent years, modeling this process in a format of phage-display combinatorial libraries has afforded a powerful tool for obtaining valuable antibodies. However, the ability to readily select and isolate rare catalysts would expand the scope of library technology. A technique in which phage infection controlled the link between recognition and replication was applied to show that chemistry is a selectable process. An antibody that operated by covalent catalysis to form an acyl intermediate restored phage infectivity and allowed selection from a library in which the catalyst constituted 1 in 10(5) members. Three different selection approaches were examined for their convenience and generality. Incorporating these protocols together with well known affinity labels and mechanism-based inactivators should allow the procurement of a wide range of novel catalytic antibodies.

Antibodies, Catalytic↗

PubMed1997

Application of capillary electrophoresis-fluorescence line-narrowing spectroscopy for on-line spectral characterization of closely related analytes.

Capillary electrophoresis (CE) interfaced with low-temperature (4.2 K) fluorescence line-narrowing spectroscopy (FLNS) is used for the separation and spectral characterization of closely related analytes. In this paper, the CE-FLNS system is applied to the analysis of a mixture of deuterated and protonated benzo[a]pyrene, a mixture of structurally similar benzo[a]pyrene and benzo[e]pyrene, and mixtures of dibenzo[a,l]pyrene-derived adenine DNA adducts. The CE-FLNS system provides on-line separation and high-resolution spectroscopic identification of CE-separated analytes, via fingerprint structure of vibrationally resolved FLN spectra at 4.2 K. The combination of the separation power of CE and the spectral selectivity of FLNS provide a methodology that has potential to become a powerful tool for molecular analyte characterization. The main applications of the CE-FLNS system, due to its selectivity, should be in the chemical analysis of structurally similar analytes and applications where analyte purity and detailed structural characterization are required.

Benzopyrenes↗

PubMed1997

Bacterial resistance to vancomycin: overproduction, purification, and characterization of VanC2 from Enterococcus casseliflavus as a D-Ala-D-Ser ligase.

The VanC phenotype for clinical resistance of enterococci to vancomycin is exhibited by Enterococcus gallinarum and Enterococcus casseliflavus. Based on the detection of the cell precursor UDP-N-acetylmuramic acid pentapeptide intermediate terminating in D-Ala-D-Ser instead of D-Ala-D-Ala, it has been predicted that the VanC ligase would be a D-Ala-D-Ser rather than a D-Ala-D-Ala ligase. Overproduction of the E. casseliflavus ATCC 25788 vanC2 gene in Escherichia coli and its purification to homogeneity allowed demonstration of ATP-dependent D-Ala-D-Ser ligase activity. The kcat/Km2 (Km2 = Km for D-Ser or C-terminal D-Ala) ratio for D-Ala-D-Ser/D-Ala-D-Ala dipeptide formation is 270/0.69 for a 400-fold selection against D-Ala in the C-terminal position. VanC2 also has substantial D-Ala-D-Asn ligase activity (kcat/Km2 = 74 mM-1min-1).

Bacterial Proteins↗

PubMed1997

Role of modified glutamic acid in the helical structure of conantokin-T.

Circular dichroism (CD) and 2-dimensional NMR were used to study the solution conformation of conantokin-T (Con-T), a small peptide toxin found in the venom of fish-hunting cone snails, and its Glu-substituted analog. Con-T lacks disulfide bonds but contains many gamma-carboxyglutamic acids (Gla), a post-translationally modified residue. Our results show that Con-T adopts an alpha-helical conformation in aqueous solution even in the absence of calcium. Glu replacements diminish both helicity and function of Con-T. The helical content of Con-T is higher than most natural helical peptides of this length in aqueous solution. The sequence of this small toxin incorporates several known elements that stabilize alpha-helical structure in peptides. Gla residues form several salt bridges that stabilize helical conformation of Con-T.

1-Carboxyglutamic Acid↗

PubMed1997

Transcription and processing of the gene for spinach chloroplast threonine tRNA in a homologous in vitro system.

An in vitro system was established to study the transcription and processing of threonine tRNA using spinach chloroplast enzyme extract. Experiments using a series of 5' deletion mutants demonstrated that the transcription of trnT gene required no 5' upstream promoter elements. Four plasmid DNA templates containing trnT were constructed for tRNA processing assay. The processing reaction was carried out either with exogenously added precursor-tRNAs made by T7 RNA polymerase or with RNAs synthesized by the transcription activity in the same processing enzyme extract. Both assays demonstrated that the 5' and 3' ends of mature tRNA were processed endonucleolytically and the processing of the 5' end preceded the maturation of the 3' end. The activity of nucleotidyl transferase that adds CCA nucleotides to the 3' end of tRNA was also observed. The use of a coupled transcription and processing system provides us with a better insight to the tRNA processing mechanism of the chloroplast.

Base Sequence↗

PubMed1997

Protein-tyrosine phosphatases PTP1B and syp are modulators of insulin-stimulated translocation of GLUT4 in transfected rat adipose cells.

The protein-tyrosine phosphatases PTP1B and Syp have both been implicated as modulators of the mitogenic actions of insulin. However, the roles of these protein-tyrosine phosphatases in the metabolic actions of insulin are not well characterized. In this study, we directly assessed the ability of PTP1B and Syp to modulate insulin-stimulated translocation of the insulin-responsive glucose transporter GLUT4 in a physiologically relevant insulin target cell. Primary cultures of rat adipose cells were transiently transfected with either wild-type PTP1B (PTP1B-WT), wild-type Syp (Syp-WT), or the catalytically inactive mutants PTP1B-C/S or Syp-C/S. The effects of overexpression of these constructs on insulin-stimulated translocation of a co-transfected epitope-tagged GLUT4 were studied. Cells overexpressing either PTP1B-C/S or Syp-WT had insulin dose-response curves similar to those obtained with control cells expressing only epitope-tagged GLUT4. In contrast, for cells overexpressing PTP1B-WT the level of GLUT4 on the cell surface at each insulin dose (ranging from 0 to 60 nM) was significantly lower than that observed in the control cells. Interestingly, cells overexpressing the dominant inhibitory mutant Syp-C/S also had a small but statistically significant impairment in insulin responsiveness. At a maximally stimulating concentration of insulin (60 nM), cell surface epitope-tagged GLUT4 was approximately 20% less than that of the control cells. It is possible that effects from high level overexpression of Syp and PTP1B constructs may not reflect what occurs under physiological conditions. Nevertheless, our data raise the possibility that PTP1B may be a negative regulator of insulin-stimulated glucose transport, while Syp may have a small role as a positive mediator of the metabolic actions of insulin.

Adipocytes↗

PubMed1997

Suppression of myelopoiesis and myeloid leukemia cell line proliferation by a novel bone marrow-derived factor, reptimed.

Bone marrow is the major site of hematopoiesis in the adult mammal. Bone marrow contains a highly organized microenvironment for the support of hematopoietic stem and progenitor cells, including the production of growth factors. Bone marrow cells also produce negative regulatory factors which may regulate hematopoiesis and inflammatory responses. In this paper we describe Reptimed, a unique bone marrow-derived factor with inhibitory activity for myelopoiesis and in vitro growth of myeloid cell lines. Reptimed was partially purified from bone marrow supernatants using a combination of solid-phase extraction and size exclusion chromatography. Reptimed is < 1000 Da MW and is water soluble. Reptimed inhibited growth of granulocyte-macrophage and macrophage colonies as well as proliferation of several myeloid leukemia cell lines. Reptimed may be part of a hemoregulatory circuit.

Animals↗

PubMed1997

Why is CMP-ketodeoxyoctonate highly unstable?

CMP-ketodeoxyoctonate (CMP-KDO) and analogs, including CMP-5-deoxy-5-fluoro-KDO, CMP-5-deoxy-KDO, and CMP-5-epi-KDO, were prepared from CTP and the corresponding KDO sugars catalyzed by CMP-KDO synthetase. These analogs were found to be much more stable than CMP-KDO (t1/2 = 0.57 h) yet less stable than CMP-sialic acid (t1/2 = 151 h). Fluorination at the 5-position of CMP-KDO has a 200-fold enhanced stability compared to the 156-fold enhancement for the 3R-fluoro analog, probably due to the loss of H-bonding interactions (for the 5-F derivative) and the cause of remote inductive effect (for the 3- and the 5-F analogs) on the glycosidic cleavage. Hydrolysis of CMP-KDO is perhaps facilitated by an intramolecular hydrogen bond from the 5-OH group with the phosphate oxygen as demonstrated by the 3-5-fold enhanced stability of CMP-5-epi-KDO and CMP-5-deoxy-KDO compared to CMP-KDO and by molecular modeling studies of water-solvated CMP-KDO. Hydrolysis of CMP-KDO also was found to be subject to a substantial solvent isotope effect (kH/kD = 2.7), which is significantly different from the reported solvent isotope effect for the hydrolysis of sialyglycosides (kH/kD = 0.86) and dependent on both buffer and magnesium ion concentrations. Considering these results and molecular modeling studies, it is proposed that the hydrolysis of CMP-KDO under neutral conditions proceeds through a glycosidic cleavage which occurs at the electronically favorable twist-boat conformation, facilitated by intramolecular H-bonding interaction of the 4-, 5- and 7- (or 8-) OH groups and the phosphate oxygen and by the leaving group magnesium ion complexation.

Buffers↗

PubMed1997

Dissection of glutathionylspermidine synthetase/amidase from Escherichia coli into autonomously folding and functional synthetase and amidase domains.

The bifunctional glutathionylspermidine synthetase/amidase from Escherichia coli catalyzes both the ATP-dependent formation of an amide bond between N1 of spermidine (N-(3-amino)propyl-1, 4-diaminobutane) and the glycine carboxylate of glutathione (gamma-Glu-Cys-Gly) and the opposing hydrolysis of this amide bond (Bollinger, J. M., Jr., Kwon, D. S., Huisman, G. W., Kolter, R., and Walsh, C. T. (1995) J. Biol. Chem. 270, 14031-14041). In our previous work describing its initial characterization, we proposed that the 619-amino acid (70 kDa) protein might possess separate amidase (N-terminal) and synthetase (C-terminal) domains. In the present study, we have confirmed this hypothesis by expression of independently folding and functional amidase and synthetase modules. A fragment containing the C-terminal 431 amino acids (50 kDa) has synthetase activity only, with steady-state kinetic parameters similar to the full-length protein. A fragment containing the N-terminal 225 amino acids (25 kDa) has amidase activity only and is significantly activated relative to the full-length protein for hydrolysis of glutathionylspermidine analogs. This observation suggests that the amidase activity in the full-length protein is negatively autoregulated. The amidase active site catalyzes hydrolysis of amide and ester derivatives of glutathione (e.g. glutathione ethyl ester and glutathione amide) but lacks activity toward acetylspermidine (N1 and N8) and acetylspermine (N1), indicating that glutathione provides the primary recognition determinants for glutathionylspermidine amide bond cleavage. No metal ion is required for the amidase activity. A tetrahedral phosphonate analogue of glutathionylspermidine, designed as a mimic of the proposed tetrahedral intermediate for either reaction, inhibits the synthetase activity (Ki approximately 10 microM) but does not inhibit the amidase activity.

Amide Synthases↗

PubMed1997

Early postnatal dexamethasone therapy may lessen lung inflammation in premature infants with respiratory distress syndrome on mechanical ventilation.

Early postnatal use of dexamethasone has recently been shown to be effective in improving the pulmonary status in premature infants with respiratory distress syndrome (RDS). To study the effect of dexamethasone on pulmonary inflammatory responses, we studied ten infants treated with dexamethasone and ten infants without this treatment. Serial tracheal aspirates were obtained for cell counts, neutrophil counts, total protein concentrations, and leukotriene B4 (LTB4) and 6-keto prostaglandin (PG)F(1 alpha) levels before and after starting the study. Infants in the dexamethasone-treated group required significantly lower mean airway pressures for ventilation and had lower PaCO2 values from day 3 to day 14 than infants in the control group, suggesting better pulmonary function. For infants in the dexamethasone group, the tracheal aspirates showed significantly lower cell and neutrophil counts, protein concentrations, and 6-keto-PGF(1 alpha) and LTB4 levels than in the control group. We conclude that early postnatal dexamethasone therapy may lessen lung inflammation and improve pulmonary function in infants with RDS.

6-Ketoprostaglandin F1 alpha↗

PubMed1997

The effect of the selective PAF antagonist CIS-19 on PAF- and antigen-induced bronchoconstriction, microvascular leakage and bronchial hyperreactivity in guinea-pigs.

We investigated the effects of a novel platelet-activating factor (PAF) receptor antagonist, CIS-19 [cis-2-(3, 4-dimethoxyphenyl)-6-isopropoxy-7-methoxy-1-(N-methyl-formamido)-1, 2, 3, 4-tetrahydronaphthalene], on PAF-, histamine-, substance P- and antigen-induced bronchoconstriction and microvascular leakage, as well as PAF- and antigen-induced bronchial hyperreactivity to methacholine in urethane-anesthetized guinea-pigs. Administration of CIS-19 (0.5-5 mg/kg, i.v.) inhibited the increase in lung resistance induced by PAF (30 ng/kg, i.v.) in a dose-dependent manner, but failed to inhibit the increase induced by histamine (30 micrograms/kg, i.v.) or substance P (6.5 micrograms/kg, i.v.). CIS-19 (5 mg/kg, i.v.) did not inhibit the increase in lung resistance induced by ovalbumin (2 mg/kg, i.v.) in actively sensitized guinea-pigs. PAF (30 ng/kg, i.v.)-induced microvascular leakage, measured by the extravasation of Evans blue dye, was dose-dependently inhibited by CIS-19 (0.5-5 mg/kg, i.v.) in the trachea, main bronchi and intrapulmonary airways, but it did not affect histamine (30 micrograms/kg, i.v.)- or substance P (6.5 micrograms/kg, i.v.)-induced microvascular leakage at all airway levels. CIS-19 (2.5 and 5 mg/kg) did not affect ovalbumin (2 mg/kg, i.v.)-induced microvascular leakage in all airway levels in actively sensitized guinea-pigs, CIS-19 (2.5 and 5 mg/kg, i.v.) significantly inhibited PAF-induced enhancement of the bronchial response to methacholine, but had no effect on ovalbumin (0.05 mg/kg, i.v.)-induced bronchial hyperreactivity in actively sensitized guinea-pigs. It is concluded that CIS-19 is a potent PAF receptor antagonist which inhibits PAF- but not antigen-induced bronchoconstriction, microvascular leakage and bronchial hyperreactivity. These results suggest that PAF plays little or no role in early airway responses following antigen challenge.

Airway Resistance↗

PubMed1997

The acetylcholine releaser linopirdine increases parietal regional cerebral blood flow in Alzheimer's disease.

Centrally acting cholinergic drugs have been reported to increase regional cerebral blood flow (rCBF) as measured by single photon emission computed tomography (SPECT) in brain regions affected by Alzheimer's disease (AD). We studied the effects of the acetylcholine releaser linopirdine (LPD) on SPECT rCBF in patients with probable AD. Twenty-four AD patients (12 M, 12 F; mean age +/- SD = 68.9 +/- 8.2 years) and 13 healthy controls (8 M, 5 F; 68.4 +/- 8.0 years) participated. AD patients were scanned with 20 mCi of Tc99m-ECD at baseline and following 4 weeks of treatment with LPD 40 mg TID (n = 15) or placebo TID (n = 9) in a double-blind trial. Healthy subjects were scanned for comparison with baseline AD scans. Cortical/cerebellar rCBF ratios were derived for nine cortical structures. The combined parietal association cortex showed a 20.6% reduction in patients relative to controls. Patients treated with LPD showed an increase in parietal rCBF of 4.1 +/- 5.8%; whereas those treated with placebo showed a decrease of -2.0 +/- 7.4% (F = 5.13; df = 1, 22; P = 0.03). These data support the conclusion that rCBF abnormalities in AD are, in part, truly "functional" and can be selectively altered with pharmacological interventions. The parietal activation seen with LPD and other cholinergic AD drug therapies suggests the importance of measuring parietal lobe neuropsychological function in the course of evaluating these drugs.

Acetylcholine↗

PubMed1997

The correlation of depression with functional activity in Parkinson's disease.

We evaluated 109 Chinese patients with Parkinson's disease (PD) in three ways: using a standardized psychiatric interview for depression and anxiety, using standardized neurological evaluation for motor disability, and using cognitive assessment for cognitive impairment. Six of the 109 patients who had dementia and another two afflicted with organic delusional disorder were excluded from further analysis. The remaining 101 PD patients were divided into the following three groups according to the DSM-III-R criteria: major depressive disorder (n = 18), other depressive disorders (n = 25) including dysthymic disorder and depressive disorder not otherwise specified, and no depression (n = 58). The frequency of major depressive disorder of the 109 PD patients was 16.5%, and the frequency of major and other depressive disorders, taken together, was 42.2%. Using the percentage points measured on the Schwab & England Activities of Daily Living Scale as the dependent variable to fit a multivariate regression model, we found the lower score significantly correlated with the diagnosis of depressive disorder and higher score of the Hamilton Depression Rating Scale, in addition to motor disability and disease severity of PD. Given the high frequency of depression and the significant correlation between depression and performance in daily functional activites, we believe that an evaluation of PD patients for coexisting depression is necessary for a better therapeutic outcome.

Aged↗

PubMed1997

Congenital bilateral agenesis of diaphragm: report of a case.

UNLABELLED: Bilateral agenesis of the diaphragm is a rare, life-threatening malformation. Infants with this defect rarely survive to have surgical intervention. We report a 32-week premature female infant who was born to a 36-year-old mother via vaginal delivery. The pregnancy course was complicated by hypertension and polyhydramnios. Cytogenetic study showed a normal 46 XX female karyotype. She had cyanosis, respiratory distress and scaphoid abdomen at birth. A roentgenograph confirmed the diagnosis of diaphragmatic hernia. Surgery was performed at 21 h of age. Bilateral agenesis of diaphragm, herniation of abdominal organs and oesophagus and pulmonary hypoplasia were noted. Furthermore, stomach and spleen were adherent to the mediastinum and vertebrae. The patient developed hypotension and persistent hypoxaemia and expired at age of 26 h. Autopsy revealed bilateral agenesis of diaphragm, hypoplasia of lungs, and pancreas fibrosis with mild hypoplasia of islets of Langerhans. CONCLUSION: Bilateral agenesis of diaphragm associated with pancreas fibrosis is a rare entity, and its clinical significance needs further investigation.

Abnormalities, Multiple↗

PubMed1997

A study of the referral patterns of obstetric clinics and the performance of receiving neonatal intensive care units in Taiwan.

To study the referral patterns of obstetric clinics, and the performance of receiving intensive care units measured by the survival of transported neonates, transport records were collected prospectively between July, 1991 and June, 1992. Two hundred and fifty-four transported neonates born in 51 obstetric clinics (level I units) in Tainan City and County, in southern Taiwan, were enrolled in this study. Nineteen percent of the transported neonates were very low birthweight infants (< 1500 g). Nearly equal numbers of them were transported to eight district hospitals (level II units) and to a tertiary center (level III unit), but these infants were 1.5 times more likely to die in a level II unit than a level III unit. In addition, equal numbers of infants assisted by mechanical ventilators were transported to level II and III units, but these infants were three times more likely to die in a level II unit than a level III unit (P = 0.006). Seventy-seven percent of the normal birthweight infants (> or = 2500 g) were transported to level II units, and the mortality in this group was 12.3% compared with 0% in those transported to the level III unit. Approximately 56% of these normal birthweight infants in level II units died of severe birth asphyxia. The referral patterns of level 1 units had an unfavorable effect on the survival of neonates requiring mechanical ventilation. Enhancing the skills of the staff in level I units to recognize and stabilize such infants, elevating the capability of level II units in treating some of these cases, and increasing the hospital beds for level III care are necessary to increase their chance of survival.

Health Services Research↗

PubMed1997

Conantokin-T selectively antagonizes N-methyl-D-aspartate-evoked responses in rat hippocampal slice.

This study investigated the mode of action of conantokin-T, a 21 amino acid peptide toxin isolated from the venom of the fish-hunting cone snail Conus tulipa, on excitatory synaptic transmission in rat hippocampal slices using intracellular recording techniques. Superfusion of conantokin-T (1-500 nM) specifically and irreversibly decreased the pharmacologically isolated N-methyl-D-aspartate receptor (NMDA)-mediated excitatory postsynaptic potential (EPSPNMDA) in a concentration-dependent manner but had no effect on normal excitatory synaptic transmission (EPSP). The sensitivity of postsynaptic neurons to NMDA but not to alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid was also antagonized by conantokin-T pretreatment. In addition, the conantokin-T-induced depression of EPSPNMDA could be antagonized by prior treatment of hippocampal slices with either DL-2-amino-5-phosphonovaleate (10 microM) or ifenprodil (20 microM). However, 7-chlorokynurenic acid (1 microM) had no effect on the action of conantokin-T. These findings indicated that conantokin-T modulates the NMDA receptor by an interaction with its glutamate binding site and polyamine recognition site.

Animals↗

PubMed1997

Swallowing difficulty in Parkinson's disease.

Dysphagia is a frequent and potentially serious complication of Parkinson's disease (PD). We examined the oropharyngeal swallowing ability in 19 PD patients (15 men and 4 women, mean age 68.42 years, mean Hoehn and Yahr stage 1.8) using modified barium swallow before and after administering oral levodopa (in combination with benserazide). Twelve (63.2%) patients demonstrated objective evidence of swallowing abnormalities; although only six patients (31.6%) had subjective complaints. Vallecula sinus and pyriform sinus residues were the most frequent abnormalities (47.4% and 42.1%); followed by delayed swallowing reflex (26.3%). Three patients demonstrated silent aspiration. In the 12 patients with abnormal swallowing, six (50%) showed objective improvement after levodopa treatment, while the remaining six showed no change. Of the former group of six, one patient showed improvement in the oral phase, but deterioration in the pharyngeal phase. We concluded that PD patients had a high percentage of objective swallowing abnormalities which could be reduced in half of the patients through the administration of levodopa treatment.

Adult↗

PubMed1997

Structural basis of DNA folding and recognition in an AMP-DNA aptamer complex: distinct architectures but common recognition motifs for DNA and RNA aptamers complexed to AMP.

BACKGROUND: Structural studies by nuclear magnetic resonance (NMR) of RNA and DNA aptamer complexes identified through in vitro selection and amplification have provided a wealth of information on RNA and DNA tertiary structure and molecular recognition in solution. The RNA and DNA aptamers that target ATP (and AMP) with micromolar affinity exhibit distinct binding site sequences and secondary structures. We report below on the tertiary structure of the AMP-DNA aptamer complex in solution and compare it with the previously reported tertiary structure of the AMP-RNA aptamer complex in solution. RESULTS: The solution structure of the AMP-DNA aptamer complex shows, surprisingly, that two AMP molecules are intercalated at adjacent sites within a rectangular widened minor groove. Complex formation involves adaptive binding where the asymmetric internal bubble of the free DNA aptamer zippers up through formation of a continuous six-base mismatch segment which includes a pair of adjacent three-base platforms. The AMP molecules pair through their Watson-Crick edges with the minor groove edges of guanine residues. These recognition G.A mismatches are flanked by sheared G.A and reversed Hoogsteen G.G mismatch pairs. CONCLUSIONS: The AMP-DNA aptamer and AMP-RNA aptamer complexes have distinct tertiary structures and binding stoichiometries. Nevertheless, both complexes have similar structural features and recognition alignments in their binding pockets. Specifically, AMP targets both DNA and RNA aptamers by intercalating between purine bases and through identical G.A mismatch formation. The recognition G.A mismatch stacks with a reversed Hoogsteen G.G mismatch in one direction and with an adenine base in the other direction in both complexes. It is striking that DNA and RNA aptamers selected independently from libraries of 10(14) molecules in each case utilize identical mismatch alignments for molecular recognition with micromolar affinity within binding-site pockets containing common structural elements.

Adenosine Monophosphate↗