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Biomedical subjects

S Wei

Publications and source records attributed to S Wei.

At least 145 records · Page 8Linked to original sources
PubMed1996

The highly stereoselective oxidation of polyunsaturated fatty acids by cytochrome P450BM-3.

Cytochrome P450BM-3 catalyzes NADPH-dependent metabolism of arachidonic acid to nearly enantiomerically pure 18(R)-hydroxyeicosatetraenoic acid and 14(S), 15(R)-epoxyeicosatrienoic acid (80 and 20% of total products, respectively). P450BM-3 oxidizes arachidonic acid with a rate of 3.2 +/- 0.4 micromol/min/nmol at 30 degrees C, the fastest ever reported for an NADPH-dependent, P450-catalyzed reaction. Fatty acid, oxygen, and NADPH are utilized in an approximately 1:1:1 molar ratio, demonstrating efficient coupling of electron transport to monooxygenation. Eicosapentaenoic and eicosatrienoic acids, two arachidonic acid analogs that differ in the properties of the C-15-C-18 carbons, are also actively metabolized by P450BM-3 (1.4 +/- 0.2 and 2.9 +/- 0.1 micromol/min/nmol at 30 degrees C, respectively). While the 17,18-olefinic bond of eicosapentaenoic acid is epoxidized with nearly absolute regio- and stereochemical selectivity to 17(S),18(R)-epoxyeicosatetraenoic acid (>/=99% of total products, 97% optical purity), P450BM-3 is only moderately regioselective during hydroxylation of the eicosatrienoic acid omega-1, omega-2, and omega-3 sp3 carbons, with 17-, 18-, and 19-hydroxyeicosatrienoic acid formed in a ratio of 2.4:2.2:1, respectively. Based on the above and on a model of arachidonic acid-bound P450BM-3, we propose: 1) the formation by P450BM-3 of a single oxidant species capable of olefinic bond epoxidation and sp3 carbon hydroxylation and 2) that product chemistry and, thus, catalytic outcome are critically dependent on active site spatial coordinates responsible for substrate binding and productive orientation between heme-bound active oxygen and acceptor carbon bond(s).

8,11,14-Eicosatrienoic Acid↗
PubMed1996

STAT3 activation is a critical step in gp130-mediated terminal differentiation and growth arrest of a myeloid cell line.

Myeloid leukemia M1 cells can be induced for growth arrest and terminal differentiation into macrophages in response to interleukin 6 (IL-6) or leukemia inhibitory factor (LIF). Recently, a large number of cytokines and growth factors have been shown to activate the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) signaling pathway. In the case of IL-6 and LIF, which share a signal transducing receptor gp130, STAT3 is specifically tyrosine-phosphorylated and activated by stimulation with each cytokine in various cell types. To know the role of JAK-STAT pathway in M1 differentiation, we have constructed dominant negative forms of STAT3 and established M1 cell lines that constitutively express them. These M1 cells that overexpressed dominant negative forms showed no induction of differentiation-associated markers including Fc gamma receptors, ferritin light chain, and lysozyme after treatment with IL-6. Expression of either c-myb or c-myc was not downregulated. Furthermore, IL-6- and LIF-mediated growth arrest and apoptosis were completely blocked. Thus these findings demonstrate that STAT3 activation is the critical step in a cascade of events that leads to terminal differentiation of M1 cells.

Acute-Phase Proteins↗
PubMed1996

Identification of an amphotericin B resistant strain of Candida albicans using a rapid 3H-glucose incorporation microassay.

Using a 3H-glucose incorporation assay, antifungal sensitivity testing undertaken on an isolate of Candida albicans cultured from the blood of a bone marrow transplant patient documented resistance to amphotericin B but sensitivity to fluconazole and itraconazole. Information obtained from in vitro antifungal sensitivity testing can be used to direct in vivo antifungal therapy. Widespread application of standardized in vitro antifungal sensitivity testing is needed.

Adult↗
PubMed1996

[The effects of intensive DMPA regimen in the treatment of Graves' ophthalmopathy].

35 patients with Graves' ophthalmopathy were treated with DMPA regimen which included dexamethasone (D) 20-30 mg and Methotrexate (M) 15-20 mg i.v. once a week and prednisone (P) 20 mg/d p. o. 4 times a week for 4 weeks followed by addition of azathioprine (A) or 6-MP 75-100 mg/d and the increment of prednisone (20 mg/d) to 5 times a week in the fifth and sixth weeks. The full intensive course of therapy lasted for 6 weeks. If necessary, the similar course of therapy might be repeated. Later on, each patient was mintainted on a therapy with P and A or P and 6-MP for another 1-2 months then the P and A or P and 6-MP for another 1-2 months, and then the P and A or 6-MP were tapered and finally discontinued at the end of sixth month. The symptoms and signs of the opthalmopathy were evaluated blindedly at the beginning and the end of the first and second courses of the therapy by two ophthalmologists. The results showed that the symptoms and signs were significantly improved in 33 (94.3%) cases. A decrease of eyeball protrusion of 1.22 +/- 1.15 and 1.28 +/- 1.15 mm in the left and right eyes respectively was shown by eophthalmometry reading (both P < 0.0005, compared with before and after the therapy). The decrease of exophthalmos was more remarkable in the cases with ophthalmopathy for < or = 1 year than that > 1 year. 11/22 patients recovered their ability to close eyes adequately. The diplopia was corrected in 8/15 cases. The mobility of eye ball returned to normal in 5/15 cases. Exposed lesion and/or ulceration of the cornea found in 7 patients were cured. Serious side and/or toxic effects of the drugs were not present during the course of therapy. It is suggested that DMPA regimen is an effective, safe and cheap approach to the treatment of Graves' ophthalmopathy.

Adult↗
PubMed1996

[A direct transnasal transsphenoidal approach to pituitary tumors].

38 cases of pituitary adenomas were resected via direct transnasal trans-sphenoidal approach. Complete or subtotal tumor excision was achieved in 36 patients, and partial excision in 2. The operative mortality was 0. The operative techniques were, in patients with small nostrile, a 5-6 mm small incision was made in the base of lateral alar, but the speculum straightly upon to the anterior wall of the sphenoidal sinus. As expanding the speculum, the middle septum was pressed to the opposite side. Under the operating microscope, the anterior wall mucose was dissected away, the bony wall drill off and then the sphenoidal sinus approached. Other steps were as usual. All cases achieved good result. The advantages of this approach were less destructive, less time-concuming, less complications and easy to perform.

Adenoma↗
PubMed1995

Role of extracellular adenosine triphosphate in the cytotoxic T-lymphocyte-mediated lysis of antigen presenting cells.

The lysis of antigen presenting cells (APCs) by cytotoxic T lymphocytes (CTLs) may be one mechanism whereby an immune response is downregulated by Staphylococcus superantigens. Disappearance of monocytes/macrophages from staphylococcal enterotoxin A (SEA)-activated peripheral blood mononuclear cell (PBMC) cultures, but not from control PBMC cultures was seen by flow cytometry. Recently, adenosine triphosphate (ATP) has been described as an effector molecule in CTL-mediated lysis of some murine tumor target cells. We have also shown that ATP caused the lysis of human macrophages, and that treatment of cells with interferon gamma (IFN gamma) rendered macrophages significantly more sensitive to ATP than untreated cells. To show that this purine nucleotide may play a role in modulating the immune system, we generated human CTLs that were stimulated with SEA, and used them as effector cells against SEA-pulsed autologous macrophages. CTLs were found to specifically lyse SEA-pulsed macrophages, while control, unpulsed, macrophages were unaffected. The addition of hexokinase, an enzyme that hydrolyzes ATP, significantly abrogated the killing of SEA-pulsed cells during the assay. In examining the mechanism of cytotoxicity, electron microscopy showed that macrophages incubated with both ATP and CTLs underwent necrosis, rather than apoptosis. From these results, it is suggested that ATP is released from CTLs during antigen presentation, and that IFN gamma-activated macrophages, which are inherently more sensitive to this mediator, are readily lysed and therefore removed from circulation, thus downregulating an immune response.

Adenosine Triphosphate↗
PubMed1995

Studies on the metabolism of retinol and retinol-binding protein in transthyretin-deficient mice produced by homologous recombination.

Tissue needs for retinoids are believed to be satisfied through the delivery in the circulation of retinol by its specific plasma transport protein, retinol-binding protein (RBP), which circulates as a 1-to-1 protein complex with transthyretin (TTR). The binding of RBP to TTR is thought to prevent filtration of retinol-RBP in the kidney and to play a role in secretion of RBP from hepatocytes. Recently a strain of mice (TTR-) that totally lacks immunoreactive TTR was produced by targeted mutagenesis. We have explored the effects of TTR deficiency on retinol and RBP metabolism in this mutant strain. In pooled plasma from the TTR- mice retinol levels averaged 6% of those of wild type animals. Similarly, plasma RBP in the TTR- mice was found to be 5% of wild type levels. Hepatic retinol and retinyl ester levels were similar for mutant and wild type mice, suggesting that the mutation affects neither the uptake nor storage of dietary retinol. Levels of retinol and retinyl esters in testis, kidney, spleen, and eye cups from TTR- mice were normal. Plasma all-trans-retinoic acid levels for the TTR- mice were 2.3-fold higher than those of wild type (425 versus 190 ng/dl). Kidney RBP levels were similar for the mutant and wild type mice and we were unable to detect intact RBP in urine from TTR- mice. Hepatic RBP levels in the TTR- mice were 60% higher than those of wild type mice (39.8 versus 25.0 micrograms of RBP/g of tissue). These data may suggest that there is a partial blockage in RBP secretion from TTR- hepatocytes that leads to lessened plasma levels of retinol-RBP.

Animals↗
PubMed1995

Metabolism of epoxyeicosatrienoic acids by cytosolic epoxide hydrolase: substrate structural determinants of asymmetric catalysis.

The metabolism of cis-epoxyeicosatrienoic acids (EETs), methyl cis-epoxyeicosatrienoates, and cis-epoxyeicosanoic acids by cytosolic epoxide hydrolase was studied to identify substrate structural features important for stereoselective metabolism and chiral diol formation. 14(R), 15(S)-, 11(S),12(R)-, and 8(S),9(R)-EET, the predominant enantiomers present endogenously in rat organs, were metabolized at substantially higher rates than their antipodes. With the exception of 8(R),9(S)-EET (Km = 41 microM), differences in enantiomer hydration rates appear to be caused by Km-independent factors since the apparent Km values for the enantiomers of 14,15-, 11,12-, and 8(S),9(R)-EET were similar (between 3 and 5 microM). Chiral analysis of the diols resulting from enzymatic hydration of homochiral EETs showed that the regio and/or stereochemistry of water addition was EET regioisomer dependent. For the 11,12-EET enantiomers, water addition was nonregioselective; whereas, with both 8,9-EET antipodes water addition occurred predominantly at C9. Importantly, for 14,15-EET the regiochemistry of water addition was enantiomer-dependent. Only with 14(R),15(S)-EET did enzymatic hydration result in regiospecific addition at C15. Hence, enantioselective EET hydration is determined, principally, by enantiomer specific differences in rates of catalytic turnover and/or substrate binding parameters. On the other hand, the chirality of the diol products is determined by EET enantiomer-dependent differences in the regiochemistry of enzymatic oxirane cleavage and water addition. Esterification resulted in an overall reduction in the rates of epoxide hydration for all three EET-methyl esters (59, 89, and 68% of the EET rate for 8,9-, 11,12-, and 14,15-EET-methyl ester, respectively) and in the loss of regioselectivity during methyl 8(S),9(R)-EET oxirane cleavage. Catalytic EET hydrogenation reduced the rates of EET hydration (56, 45, and 23% of the EET rates for 8,9-, 11,12-, and 14,15-epoxyeicosanoic acids, respectively). Compared to 14,15-EET, enzyme catalyzed hydration of 14,15-epoxyeicosanoic acid was less regioselective and yielded products with a substantially lower chiral purity. Based on these data, as well as on the documentation of 14(R),15(R)-dihydroxyeicosatrienoic acid as an endogenous constituent of rat urine we concluded that: (1) cytosolic epoxide hydrolase plays a significant role in the regio- and stereoselective metabolism of endogenous EETs; (2) differences in the affinities and/or turnover rates of the enzyme for the individual EET antipodes may be responsible for enantioselective EET metabolism; and (3) for 14,15- and 8,9-EET, regioselective and/or enantioselective oxirane water addition is responsible for asymmetric diol formation.(ABSTRACT TRUNCATED AT 400 WORDS)

8,11,14-Eicosatrienoic Acid↗
PubMed1995

Expression of interleukin-2 receptor gamma on human monocytes: characterization of lineage specific post-translational modifications.

Functional interleukin-2 receptors (IL-2R) on lymphocytes contain both IL-2R beta and gamma chains. Whereas constitutive expression of IL-2R beta has been found on monocytes, the expression of IL-2R gamma on these phagocytes has not been examined. We performed reverse-transcription-polymerase chain reaction with Southern blot analysis on RNA derived from purified human monocytes and discovered that they constitutively produce IL-2R gamma mRNA. Western immunoblotting revealed 58- and 64-kDa forms of IL-2R gamma on YT-1 and human monocytes, whereas 58-, 64-, and 69-kDa bands were detected using peripheral blood mononuclear cells and non-adherent lymphocytes. These different forms resulted from variable N-linked glycosylation since culture of the cells in tunicamycin resulted in detection of a single 39-kDa band which corresponds to the molecular weight predicted from the deduced amino acid sequence. By co-immunoprecipitation, the IL-2R beta subunit associates with only the 64-kDa IL-2R gamma protein band in monocytes.

Blotting, Southern↗
PubMed1995

Visualization of electronic properties of molecules in chemical reactions.

Modern computational methods allow for the tracking of entire chemical reactions, ranging from initial reactants, through transition states, and to the final products. They also permit the computation of a variety of properties that can change as the reaction proceeds from start to finish. Visualization of these reactions is often difficult and usually limited to static displays of specific steps along the reaction paths. This article describes a program, Reaction Viewer, that we have developed to visualize a chemical reaction dynamically. The article also describes the use of this program to see the movement of electrons and other electronic effects, as well as steric ramifications during the reaction.

Computer Graphics↗
PubMed1995

Role of cytochrome P-450 epoxygenase metabolites in EGF signaling in renal proximal tubule.

Epidermal growth factor (EGF) is a potent epithelial cell mitogen and induces eicosanoid production in many cell types. The present study examined signaling mechanisms for EGF in the renal proximal tubule, where high concentrations of cytochrome P-450 epoxygenase have been reported. In primary cultures of rabbit proximal tubule cells, EGF (30 nM) increased endogenous epoxyeicosatrienoic acid (EET) levels 5.3 +/- 1.4-fold within 10 min (n = 6). In these cells EGF-stimulated [3H]thymidine incorporation was significantly inhibited by the cytochrome P-450 inhibitors ketoconazole or clotrimazole but not by the cyclooxygenase inhibitor indomethacin. In fura 2-loaded proximal tubule cells, EGF caused a concentration-dependent increase in cytosolic Ca2+ concentration ([Ca2+]i), due to Ca2+ influx, which was inhibited by either ketoconazole or SKF-525A but not by indomethacin. Addition of 5,6-EET (0.5 microM) also induced Ca2+ influx in proximal tubule cells, whereas 8,9-11,12-, or 14,15-EET did not. In cells treated with bis(2-amino-5-methylphenoxy)ethane N,N,N',N'-tetraacetic acid tetraacetoxy-methyl ester to chelate [Ca2+]i, EGF-stimulated [3H]thymidine incorporation. These results indicate that EGF increases EET levels in proximal tubule and suggest that 5,6-EET or its metabolites may be a modulator of EGF-induced [Ca2+]i increases and involved in mitogenesis.

8,11,14-Eicosatrienoic Acid↗