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Biomedical subjects

G Chen

Publications and source records attributed to G Chen.

At least 937 records · Page 52Linked to original sources
PubMedDate not supplied

Preparation and properties of thermoreversible, phase-separating enzyme-oligo(N-isopropylacrylamide) conjugates.

A thermoreversible N-isopropylacrylamide (NIPAAm) oligomer with an N-hydroxysuccinimide (NHS) ester functional end group has been prepared for coupling to an enzyme, beta-D-glucosidase, to form a thermoreversible, phase-separating polymer-enzyme conjugate. This conjugate can be used for separation, recovery, and recycle of an enzyme simply by applying small temperature changes to the reaction medium. In contrast to the random polymer-enzyme conjugates previously reported by us and others in the literature, in this study the conjugate is formed by a single, end attachment of each oligomer chain to the enzyme. Preliminary studies show that the conjugated enzyme exhibits very high retention of activity, even higher than native enzyme, and shows improved thermal stability compared to native enzyme.

Acrylamides↗
PubMedDate not supplied

Site-specific conjugation of a temperature-sensitive polymer to a genetically-engineered protein.

A genetically-engineered mutant of cytochrome b5, incorporating a unique cysteine residue, was conjugated to maleimide-terminated oligo(N-isopropylacrylamide). The conjugation of the protein by reaction of the cysteine residue, precisely positioned by site-directed mutagenesis techniques, with an activated oligomer containing only one reactive end group in the oligomer chain permits the site-specific and stoichiometric conjugation of the oligomer with the protein. The protein-oligomer conjugate was shown to exhibit lower critical solution temperature (LCST) behavior, similar to the free oligomer. Furthermore, the LCST behavior of the protein-oligomer conjugate is reversible and allows selective precipitation of the conjugate above its LCST.

Acrylic Resins↗
PubMedDate not supplied

Synthesis and purification of thermally sensitive oligomer-enzyme conjugates of poly(N-isopropylacrylamide)-trypsin.

Using chain-transfer polymerization, we have synthesized oligomers of poly(N-isopropylacrylamide) [poly(NIPAAm)] with one carboxyl group at the end of each oligomer chain. The lower critical solution temperature (LCST) of the oligomers is very close to that of homo-poly(NIPAAm) lacking the end carboxyl group. The carboxyl groups were activated in methylene chloride using N,N'-dicyclohexyl-carbodiimide (DCC) and N-hydroxysuccinimide (NHS). A conjugate of trypsin with the preactivated oligomer has been prepared. We studied the effect of oligomer to enzyme (O/E) ratio in the feed on the O/E ratio of the conjugate (the average number of oligomer chains conjugated to one trypsin molecule), assuming that only the primary amino groups of lysine residues and the amino terminal of trypsin would react. The O/E ratio of the conjugate was estimated by determination of the remaining primary amine groups on the trypsin molecule. More than 95% of the conjugate can be recovered by thermally induced precipitation.

Acrylic Resins↗
PubMedDate not supplied

Mitogenic activities of water-soluble and -insoluble insulin conjugates.

Insulin was covalently bound to water-soluble polymers such as poly(oxyethylene) and poly(acrylic acid). The former and the latter product are water-soluble monovalent and multivalent conjugates, respectively. Insulin was also bound to a poly(acrylic acid)-grafted polystyrene film, to form a water-insoluble multivalent conjugate. The matrix polymer was prepared by graft polymerization of acrylic acid initiated by glow-discharged polystyrene film. Insulin coupled with poly(oxyethylene) reduced the mitogenic activity, but the poly(acrylic acid)-insulin conjugate stimulated cell growth more than native insulin. A concentration of immobilized insulin much lower than that of native insulin and the water-soluble insulin conjugates accelerated cell growth. The maximal mitogenic effect of the immobilized insulin was greater than that of native insulin or the water-soluble insulin conjugates. The findings suggest that the mitogenic effect of the water-insoluble, multivalent insulin conjugate lasts longer than that of the water-soluble conjugates, owing to the absence of internalization into the cell.

Acrylic Resins↗
PubMedDate not supplied

Photoimmobilization of sulfated hyaluronic acid for antithrombogenicity.

Anticoagulant polymer sulfated hyaluronic acid was patterned immobilized on a poly(ethylene terephthalate) (PET) film in a specific pattern by photolithography. Hyaluronic acid was sulfated by a sulfur trioxide-pyridine complex. The polymer was coupled with azidoaniline. The derivatized polymer was cast on a PET film from aqueous solution. After drying, the film was photoirradiated in the presence or absence of a photomask. The micropatterning was confirmed by staining with a dye, brilliant green. Since the anticoagulant polymer has negative charges, the cationic dye was adsorbed on the regions where the anticoagulant polymer was immobilized. Platelet adhesion was reduced on the sulfated hyaluronic acid-immobilized areas. The immobilized sulfated hyaluronic acid significantly reduced thrombus formation.

Blood Coagulation↗
PubMedDate not supplied

Micropatterned immobilization of epidermal growth factor to regulate cell function.

Photoreactive epidermal growth factor (EGF) was synthesized by conjugating mouse EGF with photoreactive polyallylamine, which was synthesized by the coupling reaction of polyallylamine with N-[4-(azidobenzoyl)oxy]succinimide. The EGF derivative was pattern-immobilized onto a polystyrene plate by UV irradiation in the presence of a photomask in a prescribed micropattern. The patterned immobilization of EGF on the polystyrene plate was confirmed by immunostaining with anti-EGF antibody. Chinese hamster ovary (CHO) cells overexpressing EGF receptors were cultured on the micropatterned plate. The phosphorylated tyrosine residues of signal proteins, including EGF receptors, were detected only in the cells adhered in the EGF-immobilized area, and cell growth was observed only in the EGF-immobilized area. The cells growing in the EGF-immobilized area were partially stained by anti-phosphotyrosine antibody, when the area of EGF immobilization was smaller than the cell. The partial staining of activated proteins indicates that immobilization of EGF inhibited the free lateral diffusion and internalization of the activated EGF-EGF receptor complex. The enhanced cell growth is due to juxtacrine stimulation realized by immobilized EGF.

Animals↗
PubMedDate not supplied

A quantitative immunoassay utilizing Escherichia coli cells possessing surface-expressed single chain Fv molecules.

A facile, quantitative immunoassay is described that utilizes Escherichia coli (E. coli) bacteria expressing single chain Fv (scFv) antibody fragments attached to the cell surface. A Scatchard analysis demonstrated that the antibodies on the surface of the cells retained full binding activity (Kd = 2.2 x 10(-9) M) and that there are 60,000 scFv molecules per cell. The cells are used as the antibody reagent in the assay, and, following incubation with analyte, simple centrifugation is used to separate the antibody-bound from unbound analyte. The immunoassay is rapid and accurate down to the nanomolar level. In addition, a variety of detection strategies can be used, and the immunoassay is not adversely affected by the presence of animal serum. A key advantage of the new immunoassay is that the antibody reagent can be inexpensively produced in a "ready to use" form by simply growing cultures of the bacteria.

Antigens, Surface↗
PubMedDate not supplied

Photoimmobilization of insulin onto polystyrene dishes for protein-free cell culture.

Photoreactive insulin was synthesized by coupling with azidobenzoic acid. The insulin derivative was immobilized onto the wells of polystyrene culture plates by photoir-radiation. The photoimmobilized insulin enhanced the growth of anchorage-dependent cells such as Chinese hamster ovary CHO-K1 and mouse fibroblast STO cells by more than native or azidophenyl-derivatized insulin. A small amount of photoimmobilized insulin (1-10% of the amount of the native or derivatized insulin) enhanced the growth of CHO-K1 and STO cells. In addition, the maximal mitogenic effect of the immobilized insulin was greater than that of native or derivatized insulin. Photoimmobilization could be a universal means of immobilizing growth factors onto the surface of materials devoid of chemical functional groups scaffolding growth factors and providing a new protein-free cell culture system or tissue engineering materials.

Animals↗
PubMedDate not supplied

(Tetrakis(2-pyridylmethyl)ethylenediamine)iron(II) perchlorate. Study of density functional methods.

On the basis of the data obtained by X-ray diffraction, the properties of two independent crystallographic subsystems in the [Fe(tpen)](ClO4)2.2/3H2O complex are studied in detail with the density functional method B3LYP. The energies of singlet, triplet, and quintet states at different temperatures are obtained, the influences of geometry on energy changes are analyzed, the regularity of the spin-state interconversions is investigated, and the effect of the triplet and action of the anion on spin crossover are discussed. This investigation demonstrates that (1) the energy difference between the high-spin state and singlet state decreases as the Fe-N distance and geometric distortion increase, (2) the spin-equilibrium system is predominantly in low-spin form below room temperature and the proportion of high-spin state rapidly increases above room temperature, (3) one of the two cation sites has a greater presence of the high-spin content, (4) the triplet state may be responsible for the fast rate of spin-state interconversions, and (5) the B3LYP method proves to be very adequate to study the spin-state transition of this complex.

Journal Article↗
PubMedDate not supplied

Influence of substrate structure on the catalytic efficiency of hydroxysteroid sulfotransferase STa in the sulfation of alcohols.

Sulfotransferase a (STa) is an isoform of hydroxysteroid (alcohol) sulfotransferase that catalyzes the formation of sulfuric acid esters from both endogenous and xenobiotic alcohols. Among its various functions in toxicology, STa is the major form of hepatic sulfotransferase in the rat that catalyzes the formation of genotoxic and carcinogenic sulfuric acid esters from hydroxymethyl polycyclic aromatic hydrocarbons. The goal of the present study was to elucidate fundamental quantitative relationships between substrate structure and catalytic activity of STa that would be applicable to these and other xenobiotics. We have modified previous procedures for purification of STa in order to obtain sufficient amounts of homogeneous enzyme for determination of kcat/Km values, a quantitative measure of catalytic efficiency. We determined the catalytic efficiency of STa with benzyl alcohol and eight benzylic alcohols that were substituted with n-alkyl groups (CnH2n + 1, where n = 1-8) in the para position, and the optimum value for kcat/Km in these reactions was obtained with n-pentylbenzyl alcohol. Correlations between logarithms of kcat/Km values and logarithms of partition coefficients revealed that hydrophobicity of the substrate was a major factor contributing to the catalytic efficiency of STa. Primary n-alkanols (CnH(2n+1)OH, where n = 3-16) exhibited an optimum kcat/Km for C9-C11 and a linear decrease in vmax of the reaction for C3-C14; 15- and 16-carbon n-alkanols were not substrates for STa. These results indicated limits to the length of the extended carbon chain in substrates. Such limits may also apply to hydroxysteroids, since cholesterol was inactive as either substrate or inhibitor of STa. Furthermore, the importance of steric effects on the catalytic efficiency of STa was also evident with a series of linear, branched, and cyclic seven-carbon aliphatic alcohols. In conclusion, our results provide fundamental quantitative relationships between substrate structure and catalytic efficiency that yield insight into the specificity of STa for both endogenous and xenobiotic alcohols.

9,10-Dimethyl-1,2-benzanthracene↗
PubMedDate not supplied

A learning deficit related to age and beta-amyloid plaques in a mouse model of Alzheimer's disease.

Mice that overexpress the human mutant amyloid precursor protein (hAPP) show learning deficits, but the apparent lack of a relationship between these deficits and the progressive beta-amyloid plaque formation that the hAPP mice display is puzzling. In the water maze, hAPP mice are impaired before and after amyloid plaque deposition. Here we show, using a new water-maze training protocol, that PDAPP mice also exhibit a separate age-related deficit in learning a series of spatial locations. This impairment correlates with beta-amyloid plaque burden and is shown in both cross-sectional and longitudinal experimental designs. Cued navigation and object-recognition memory are normal. These findings indicate that A beta overexpression and/or A beta plaques are associated with disturbed cognitive function and, importantly, suggest that some but not all forms of learning and memory are suitable behavioural assays of the progressive cognitive deficits associated with Alzheimer's-disease-type pathologies.

Aging↗
PubMedDate not supplied

CT findings after uterine artery embolization.

Asymptomatic uterine leiomyoma can be detected on routine computed tomography (CT) of the pelvis. Leiomyomas have been described as low attenuation masses that can disrupt the smooth contour of a normal uterus. Four women underwent uterine artery embolization for the treatment of uterine leiomyoma. CT findings include initial retention of contrast in fibroids the day of the procedure and central necrosis of the fibroid with subsequent cavitation as early as 1 month postprocedure.

Adult↗
PubMedDate not supplied

Regulation of signal transduction pathways and gene expression by mood stabilizers and antidepressants.

OBJECTIVE: To determine whether the currently available evidence supports the hypothesis that antidepressants and mood stabilizers may bring about some of their long-term therapeutic effects by regulating signal transduction pathways and gene expression in the central nervous system. METHODS: To address this question, we reviewed the evidence showing that chronic administration of antidepressants and mood stabilizers involves alterations in signaling pathways and gene expression in the central nervous system. RESULTS: A large body of data has shown that lithium and valproate exert effects on the protein kinase C signaling pathway and the activator protein 1 family of transcription factors; in contrast, antidepressants affect the cyclic adenosine monophosphate pathway and may bring about their therapeutic effects by modulating cyclic adenosine monophosphate-regulated gene expression in the central nervous system. CONCLUSIONS: Given the key roles of these signaling cascades in the amplification and integration of signals in the central nervous system, the findings have clear implications not only for research into the etiology and pathophysiology of the severe mood disorders but also for the development of novel and innovative treatment strategies.

Antidepressive Agents↗
PubMedDate not supplied

Evans syndrome: results of a national survey.

PURPOSE: Our goal was to improve the management of Evans Syndrome, an uncommon and frequently refractory condition. We conducted a retrospective survey to assess the demography, presentation, clinical course, and treatment response of affected children. PATIENTS AND METHODS: Information was analyzed from a detailed questionnaire completed by pediatric hematologists mainly in the U.S. and Canada. We sought information regarding demographics, findings at presentation, approach to diagnosis, treatments used (with specific reference to splenectomy, corticosteroids, and intravenous immunoglobulin (IVIG)), course of the disease with emphasis on recurrences, and status at last follow-up. RESULTS: Forty-two patients (22 male, 20 female) were included in the study. The median age was 7.7 years (range 0.2-26.6 years). At presentation, thrombocytopenia (32 patients) and anemia (28) were common; neutropenia occurred in 10 and pancytopenia in 6. Patients received a median of 5 (range 0-12) modalities of treatment. Courses of IVIG and corticosteroids were given to almost all patients; responses were varied but the effects lasted as long as 2 years. Splenectomy was performed for 15 patients but the median duration of response was only 1 month. Other treatments included cyclosporine, vincristine, danazol, azathioprine, cyclophosphamide, and plasmapheresis. The course of the disease was characterized by recurrent thrombocytopenia, hemolytic anemia, and neutropenia. After a median follow-up of 3 years, 3 patients had died, 20 had active disease on treatment, 5 had persistent disease (not on treatment), and 14 had no evidence of disease. CONCLUSION: Evans syndrome is a chronic and recurrent condition which is often refractory to IVIG, corticosteroids, and splenectomy. Responses to other agents have been anecdotal and inconclusive. A prospective study involving these agents is needed to determine optimal therapeutic combinations.

Adolescent↗
PubMedDate not supplied

Noninvasive diagnostic testing of coronary artery disease in women.

Noninvasive diagnostic testing of coronary artery disease (CAD) is widely recognized as an area that is less studied and less accurate with regard to women than to men. Accurate and safe diagnostic testing constitutes the crucial link between early detection and optimal management of CAD. Many noninvasive diagnostic modalities are available to the clinician, including traditional electrocardiography, the relatively novel imaging of echocardiography, the emerging nuclear perfusion technology of electron beam computed tomography, exercise testing, and pharmacologic testing. The most accurate and cost-effective diagnostic method for patients depends on the patients' pretest likelihood of the disease as determined by factors such as sex, age, and cardiovascular risk factors. Noninvasive tests are most useful in the diagnosis of CAD in patients with intermediate pretest likelihood of CAD. Patients with low pretest likelihood of CAD with normal electrocardiograms may benefit from noninvasive tests or a watchful waiting strategy. Patients with a high pretest likelihood of CAD may benefit greatly from direct referral to coronary angiography. Among the noninvasive diagnostic methods, exercise electrocardiography is the most studied and least accurate with regard to women patients. Electrocardiography improves in accuracy when combined with imaging techniques such as echocardiography or nuclear single photon emission computed tomography. Combining data from all studies has shown that exercise echocardiography yields the highest diagnostic accuracy in women among all of the exercise stress tests. Patients who are unable to achieve maximal exercise capacity may undergo pharmacologic testing using dipyridamole or adenosine radionuclide perfusion or dobutamine echocardiography. Recent development of electron beam computed tomography accurately detects coronary artery calcium but has not been validated yet as a standard diagnostic test for CAD.

Coronary Angiography↗