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Y C Chai

Publications and source records attributed to Y C Chai.

10 recordsLinked to original sources
PubMed2000

Relationship of molecular structure to the mechanism of lysophospholipid-induced smooth muscle cell proliferation.

We previously reported that oxidized low-density lipoprotein and one of its constituents, lysophosphatidylcholine (lysoPC), caused smooth muscle cell proliferation that was inhibitable by vitamin E and by a neutralizing antibody against basic fibroblast growth factor-2 (FGF-2). We now show that the mitogenic activity of lysolipids is highly dependent on structure. Phospholipids with palmitoyl fatty acid and phosphocholine induced DNA synthesis optimally. Shorter and longer fatty acids were significantly less potent, as were phosphoserine and phosphoethanolamine head groups. Structurally related phospholipids [platelet-activating factor (PAF) and lysoPAF] were also mitogens and acted via an analogous FGF-2-dependent, vitamin E-inhibitable mechanism. The mechanism of lysoPC stimulation was distinct from that of another phospholipid mitogen, lysophosphatidic acid (lysoPA), in that lysoPC stimulation was not pertussis toxin inhibitable. Furthermore, lysoPA stimulation was not inhibitable by vitamin E. Despite its distinct cellular pathway for stimulation, lysoPA also ultimately led to FGF-2 release. Our data show that specific structural attributes of lysoPC, PAF, and lysoPAF enable these agents to mediate smooth muscle cell release of FGF-2, which in turn stimulates proliferation.

Animals↗
PubMed1998

iNOS expression in human intestinal microvascular endothelial cells inhibits leukocyte adhesion.

Increased nitric oxide (NO) production by inducible nitric oxide synthase (iNOS) has been associated with intestinal inflammation, including human inflammatory bowel disease. However, NO can downregulate endothelial activation and leukocyte adhesion, critical steps in the inflammatory response. Using primary cultures of human intestinal microvascular endothelial cells (HIMEC), we determined the role of NO in the regulation of HIMEC activation and interaction with leukocytes. Both nonselective (NG-monomethyl-L-arginine) and specific (N-iminoethyl-L-lysine) competitive inhibitors of iNOS significantly increased binding of leukocytes by HIMEC activated with cytokines and lipopolysaccharide. Increased adhesion was reversible with the NOS substrate L-arginine and was not observed in human umbilical vein endothelial cells (HUVEC). Activation of HIMEC significantly upregulated HIMEC iNOS expression and NO production. NOS inhibitors did not augment cell adhesion molecule levels in activated HIMEC but did result in sustained increases in intracellular reactive oxygen species. In addition, antioxidant compounds reversed the effect of NOS inhibitors on HIMEC-leukocyte interaction. Taken together, these data suggest that after HIMEC activation, iNOS-derived NO is an endogenous antioxidant, downregulating leukocyte binding and potentially downregulating intestinal inflammation.

Arginine↗
PubMed1996

Oxidized low density lipoprotein and lysophosphatidylcholine stimulate cell cycle entry in vascular smooth muscle cells. Evidence for release of fibroblast growth factor-2.

We have previously shown that oxidized low density lipoprotein (LDL) but not native LDL stimulated DNA synthesis in cultured smooth muscle cells (SMC) and that alpha-tocopherol (vitamin E) inhibited this proliferative response (Lafont, A., Chai, Y. C., Cornhill, J. F. , Whitlow, P. L., Howe, P. H., and Chisolm, G. M.(1995) J. Clin. Invest. 95, 1018-1025). The moiety of oxidized LDL that stimulates DNA synthesis and the cellular mechanism for this potentially mitogenic effect are not known. We now report that lipid fractions containing lysophospholipids from oxidized LDL or phospholipase A2-treated native LDL stimulated SMC DNA synthesis as did palmitoyl lysophosphatidylcholine (lysoPC). Protein kinase C inhibitors and down-regulation of protein kinase C activity by phorbol ester inhibited oxidized LDL- and lysoPC-induced DNA synthesis. A neutralizing monoclonal antibody against fibroblast growth factor-2 significantly inhibited oxidized LDL and lysoPC-induced DNA synthesis in SMC; irrelevant antibodies were ineffective. Vitamin E inhibited the DNA synthesis stimulated by lysoPC, an observation that distinguished this effect from DNA synthesis induced by another detergent, digitonin. These results suggest that oxidized LDL and its lysoPC moiety stimulate SMC to enter the cell cycle via an oxidative mechanism that causes the release of fibroblast growth factor-2 and a subsequent autocrine or paracrine response.

Animals↗
PubMed1995

Effect of alpha-tocopherol on restenosis after angioplasty in a model of experimental atherosclerosis.

The ability of alpha-tocopherol to reduce restenosis after angioplasty was tested in a rabbit model in which angioplasty was performed on established atherosclerotic lesions. Lesions induced by 4 wk of cholesterol feeding after focal desiccation of femoral arteries were balloon dilated. 3 wk after angioplasty, angiographically determined minimum luminal diameters were less in the untreated group (0.80 +/- 0.51 mm) than in the group treated with oral alpha-tocopherol beginning 19 d before angioplasty (1.38 +/- 0.29 mm; P < 0.01). The cross-sectional area of the intima-media was greater in the untreated group (1.18 +/- 0.48 mm2) than in the alpha-tocopherol group (0.62 +/- 0.25 mm2, P < 0.0001). These differences were not due to vasoconstriction or altered plasma cholesterol. Alpha-tocopherol thus reduced restenosis after angioplasty in this model. In rabbit vascular smooth muscle cells, oxidized low density lipoprotein stimulated DNA synthesis. Alpha-tocopherol treatment inhibited DNA synthesis stimulated by oxidized low density lipoprotein, but not by serum. The findings are consistent with the hypothesis that oxidized lipids can stimulate hyperplasia and that antioxidants may limit hyperplasia by inhibiting either the oxidation or the proliferative effects of oxidants on cells.

Administration, Oral↗
PubMed1994

S-thiolation and irreversible oxidation of sulfhydryls on carbonic anhydrase III during oxidative stress: a method for studying protein modification in intact cells and tissues.

S-thiolation of carbonic anhydrase III (CA III) in cultured rat hepatocytes under oxidative stress was studied by immunodetection on nitrocellulose blots of isoelectrofocusing gels. In cells treated with menadione, three S-thiolated forms of CA III were detected, whereas only two forms were observed in hepatocytes treated with t-butyl hydroperoxide. Two "nonreducible" oxidized forms of CA III were also detected on nitrocellulose blots. These forms increased with the amount of stress and were the only modified forms of CA III in buthionine sulfoxide-treated hepatocytes containing 10-fold less glutathione than control hepatocytes. These experiments support the concept that S-thiolation protects CA III from irreversible oxidation during oxidative stress. Partly and fully S-thiolated forms of CA III were easily detected in both male and female hepatocytes by the immunoblotting method, although female cells contained 15-fold less CA III than did male liver. S-thiolated forms of CA III were also detected in rat skeletal muscle and heart showing the utility of this method for determining the effect of oxidative stress on specific S-thiolatable protein in several tissues in vivo.

Animals↗
PubMed1994

Protein S-thiolation in hepatocytes stimulated by t-butyl hydroperoxide, menadione, and neutrophils.

In order to examine potentially important S-thiolated proteins, 35S-labeled hepatocytes were exposed to oxidative stress. A similar group of S-thiolated proteins including carbonic anhydrase III was observed in cells treated with t-butyl hydroperoxide, menadione, or stimulated neutrophils. The radioactive thiols bound to hepatocyte proteins were identified by HPLC and more than 85% was glutathione. In menadione-treated hepatocytes, proteins were gradually S-thiolated over 30 min and 25% of the cellular glutathione pool became protein-bound. In t-butyl hydroperoxide-treated cells, S-thiolation was more transient and 11% of the glutathione was protein-bound. Neutrophil-treated hepatocytes had nearly the same amount of protein S-thiolation (8% after 25 min). Two major proteins that were S-thiolated in untreated hepatocytes did not increase during any form of oxidative stress. In neutrophil-treated hepatocytes protein S-thiolation was not accompanied by either formation of glutathione disulfide or a measurable change in the total amount of glutathione. In both t-butyl hydroperoxide-and menadione-treated cells there was extensive formation of glutathione disulfide and in menadione-treated cells a significant increase in the total hepatocyte glutathione pool was observed. This result suggests that protein S-thiolation may occur by mechanisms that do not result from thiol/disulfide exchange between glutathione disulfide and protein sulfhydryls. It is suggested that a thiyl radical intermediate is important in neutrophil-mediated protein S-thiolation.

Animals↗
PubMed1994

S-thiolation of individual human neutrophil proteins including actin by stimulation of the respiratory burst: evidence against a role for glutathione disulfide.

Protein S-thiolation, a reversible modification of protein sulfhydryls resulting in formation of mixed-disulfides, was studied in human neutrophils stimulated with phorbol diester to produce superoxide anion. Rapid S-thiolation of several proteins was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Glutathione was identified as the primary protein-bound thio by HPLC chromatography, contributing considerably more than 85% of the total. Minor amounts of homocysteine and/or cysteine were also detected as protein-bound thiols. During the first 30 min after stimulation, 10% of the cellular glutathione became protein bound (2 nmol/mg of protein). There was no increase in glutathione disulfide suggesting that S-thiolation of the proteins did not occur by thiol/disulfide exchange. Approximately 10 mol% of one heavily modified band (29 kDa) was S-thiolated after 30 min. A second major band of 42 kDa was identified as actin. It contained 1/10th of the total protein-bound glutathione and approximately 5 mol% was S-thiolated after 30 min. These experiments identify a subset of S-thiolated neutrophil proteins, including actin, whose modification is related to the phorbol diester stimulation of superoxide anion production in human neutrophils. Ten percent of the total glutathione pool became protein-bound without an appreciable change in non-bound concentration of glutathione or glutathione disulfide. These results suggest that glutathione was synthesized during initial phases of the respiratory burst, compensating for the amount of glutathione that became protein-bound. Since there was no significant increase in glutathione disulfide, it was probably not important in the observed protein S-thiolation.

Actins↗
PubMed1991

Identification of an abundant S-thiolated rat liver protein as carbonic anhydrase III; characterization of S-thiolation and dethiolation reactions.

An S-thiolated 30-kDa protein has been purified from rat liver by two steps of ion-exchange chromatography. This monomeric protein has two "reactive" sulfhydryls that can be S-thiolated by glutathione (form a mixed disulfide with glutathione) in intact liver. The protein has been identified as carbonic anhydrase III by sequence analysis of tryptic peptides from the pure protein. The two "reactive" sulfhydryls on this protein can produce three different S-thiolated forms of the protein that can be separated by isoelectric focusing. Using this technique it was possible to study the S-thiolation and dethiolation reactions of the pure protein. The reduced form of this protein was S-thiolated both by thiol-disulfide exchange with glutathione disulfide and by oxyradical-initiated S-thiolation with reduced glutathione. The S-thiolation rate of this 30-kDa protein was somewhat slower than that of glycogen phosphorylase b by both S-thiolation mechanisms. The S-thiolated form of this protein was poorly dethiolated (i.e., reduced) by glutathione, cysteine, cysteamine, or coenzyme A alone. Enzymatic catalysis by two different enzymes (glutaredoxin and thioredoxin-like) greatly enhanced the dethiolation rate. These experiments suggest that carbonic anhydrase III is a major participant in the liver response to oxidative stress, and that the protein may be S-thiolated by two different non-enzymatic mechanisms and dethiolated by enzymatic reactions in intact cells. Thus, the S-thiolation/dethiolation of carbonic anhydrase III resembles glycogen phosphorylase and not creatine kinase.

Amino Acid Sequence↗