This study investigates the effects and possible mechanism of an agonist of PPARα, Wy14643, on primary hepatocytes subjected to H/R injury in rats. H/R induced a significant increase ALT, AST, MDA in the culture medium and ROS in the hepatocytes. These effects were reversed by pretreatment with Wy14643 in the dose-dependent manner. The activity of SOD and the level of GSH in the hepatocytes were decreased after H/R, which were increased by Wy14643 pretreatment. Moreover, the mRNA expressions of PPARα significantly increased in H/R+Wy14643 groups when compared with that in H/R group. A PPARα agonist, Wy14643, exerts significant protective effect against H/R injury in primary hepatocytes via PPARα activation and attenuating oxidative stress. 1. Introduction Ischemia/reperfusion (I/R) injury is a serious complication precipitated during short-term expansion of the invading parasitic pathogens, such as by Entamoeba histolytica as the infections lead to local tissue damage and hypoxia [1]. The condition has also been reported to occur in certain intracellular bacterial infections, for example, Chlamydia species. Mechanisms of I/R injury involve complex and multiple pathways, including the direct ischemic cellular damage as well as the cell injury due to the activation of inflammatory response after reperfusion [2]. A hepatic, in vitro hypoxia/reoxygenation (H/R) model can be use to study the pathophysiology of this injury [3]. Previous study showed that the generation of reactive oxygen species (ROS) is likely to be an important factor in H/R-induced cell damage. Thus, ROS are generated immediately after H/R and activate proapoptotic/inflammatory signalling in the cell or directly damage cell organelles. These include direct oxidation of cellular components and lipids (lipid peroxidation), activation of inflammatory gene transcription, and possible activation of the innate immune response [2, 3]. Administration of antioxidants such as glutathione could afford protection against I/R injury [4, 5]. Oxygen deprivation (hypoxia) during ischemia and subsequent reoxygenation upon reperfusion are thought to be the major factors contributing to ROS production and the subsequent cellular damage [6]. Hypoxia increases mitochondrial reactive oxygen species (ROS) generation at Complex III, and the predominant source of ROS by oxygen limitation originates from mitochondria [7]. Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear receptor related to retinoid, steroid, and thyroid hormone receptors. Peroxisome proliferator-activated receptor-α
References
[1]
A. Olivos-García, M. Nequiz-Avenda？o, E. Tello et al., “Inflammation, complement, ischemia and amoebic survival in acute experimental amoebic liver abscesses in hamsters,” Experimental and Molecular Pathology, vol. 77, no. 1, pp. 66–71, 2004.
[2]
J. A. Wang, T. L. Chen, J. Jiang et al., “Hypoxic preconditioning attenuates hypoxia/reoxygenation-induced apoptosis in mesenchymal stem cells,” Acta Pharmacologica Sinica, vol. 29, no. 1, pp. 74–82, 2008.
[3]
R. Monno, L. Fumarola, P. Trerotoli et al., “Seroprevalence of Chlamydophila pneumoniae in ischaemic heart disease,” New Microbiologica, vol. 33, no. 4, pp. 381–385, 2010.
[4]
R. J. Schauer, S. Kalmuk, A. L. Gerbes et al., “Intravenous administration of glutathione protects parechymal and non-paranchymal liver cells against reperfusion injury following rat liver transplantation,” World Journal of Gastroenterology, vol. 10, no. 6, pp. 864–870, 2004.
[5]
J. F. Su, C. J. Guo, J. Y. Wei, J. J. Yang, Y. G. Jiang, and Y. F. Li, “Protection against hepatic ischemia-reperfusion injury in rats by oral pretreatment with quercetin,” Biomedical and Environmental Sciences, vol. 16, no. 1, pp. 1–8, 2003.
[6]
J. M. Lluis, A. Morales, C. Blasco et al., “Critical role of mitochondrial glutathione in the survival of hepatocytes during hypoxia,” Journal of Biological Chemistry, vol. 280, no. 5, pp. 3224–3232, 2005.
[7]
N. S. Chandel, D. S. McClintock, C. E. Feliciano et al., “Reactive oxygen species generated at mitochondrial complex III stabilize hypoxia-inducible factor-1α during hypoxia: a mechanism of O2 sensing,” Journal of Biological Chemistry, vol. 275, no. 33, pp. 25130–25138, 2000.
[8]
J. C. Fruchart, P. Duriez, and B. Staels, “Peroxisome proliferator-activated receptor-alpha activators regulate genes governing lipoprotein metabolism, vascular inflammation and atherosclerosis,” Current Opinion in Lipidology, vol. 10, no. 3, pp. 245–257, 1999.
[9]
L. A. Moraes, L. Piqueras, and D. Bishop-Bailey, “Peroxisome proliferator-activated receptors and inflammation,” Pharmacology and Therapeutics, vol. 110, no. 3, pp. 371–385, 2006.
[10]
S. Q. Xu, Y. H. Li, S. H. Hu, K. Chen, and L. Y. Dong, “Effects of Wy14643 on hepatic ischemia reperfusion injury in rats,” World Journal of Gastroenterology, vol. 14, no. 45, pp. 6936–6942, 2008.
[11]
T. L. Yue, W. Bao, B. M. Jucker et al., “Activation of peroxisome proliferator-activated receptor-α protects the heart from ischemia/reperfusion injury,” Circulation, vol. 108, no. 19, pp. 2393–2399, 2003.
[12]
A. Sivarajah, P. K. Chatterjee, Y. Hattori et al., “Agonists of peroxisome-proliferator activated receptor-α (clofibrate and WY14643) reduce renal ischemia/reperfusion injury in the rat,” Medical Science Monitor, vol. 8, no. 12, pp. BR532–BR539, 2002.
[13]
M. Collino, M. Aragno, R. Mastrocola et al., “Oxidative stress and inflammatory response evoked by transient cerebral ischemia/reperfusion: effects of the PPAR-α agonist WY14643,” Free Radical Biology and Medicine, vol. 41, no. 4, pp. 579–589, 2006.
[14]
P. Delerive, K. De Bosscher, S. Besnard et al., “Peroxisome proliferator-activated receptor α negatively regulates the vascular inflammatory gene response by negative cross-talk with transcription factors NF-κB and AP-1,” Journal of Biological Chemistry, vol. 274, no. 45, pp. 32048–32054, 1999.
[15]
T. Okaya and A. B. Lentsch, “Peroxisome proliferator-activated receptor-α regulates postischemic liver injury,” American Journal of Physiology, vol. 286, no. 4, pp. G606–G612, 2004.
[16]
P. Razeghi, M. E. Young, S. Abbasi, and H. Taegtmeyer, “Hypoxia in vivo decreases peroxisome proliferator-activated receptor α-regulated gene expression in rat heart,” Biochemical and Biophysical Research Communications, vol. 287, no. 1, pp. 5–10, 2001.
[17]
P. O. Seglen, “Preparation of isolated rat liver cells,” Methods in Cell Biology, vol. 13, pp. 29–83, 1976.
[18]
E. Tüzüner, L. Liu, M. Shimada et al., “Heme oxygenase-1 protects human hepatocytes in vitro against warm and cold hypoxia,” Journal of Hepatology, vol. 41, no. 5, pp. 764–772, 2004.
[19]
P. S. Smith, W. Zhao, D. R. Spitz, and M. E. Robbins, “Inhibiting catalase activity sensitizes 36B10 rat glioma cells to oxidative stress,” Free Radical Biology and Medicine, vol. 42, no. 6, pp. 787–797, 2007.
[20]
J. A. Buege and S. D. Aust, “Microsomal lipid peroxidation,” Methods in Enzymology, vol. 52, pp. 302–310, 1978.
[21]
F. Tietze, “Enzymic method for quantitative determination of nanogram amounts of total and oxidized glutathione: applications to mammalian blood and other tissues,” Analytical Biochemistry, vol. 27, no. 3, pp. 502–522, 1969.
[22]
H. Jaeschke, “Mechanisms of reperfusion injury after warm ischemia of the liver,” Journal of Hepato-Biliary-Pancreatic Surgery, vol. 5, no. 4, pp. 402–408, 1998.
[23]
K. D. Mansfield, M. C. Simon, and B. Keith, “Hypoxic reduction in cellular glutathione levels requires mitochondrial reactive oxygen species,” Journal of Applied Physiology, vol. 97, no. 4, pp. 1358–1366, 2004.
[24]
L. A. Dada, N. S. Chandel, K. M. Ridge, C. Pedemonte, A. M. Bertorello, and J. I. Sznajder, “Hypoxia-induced endocytosis, of Na,K-ATPase in alveolar epithelial cells is mediated by mitochondrial reactive oxygen species and PKC-zeta,” Journal of Clinical Investigation, vol. 111, no. 7, pp. 1057–1064, 2003.
[25]
B. J. Keller, B. U. Bradford, D. S. Marsman et al., “The nongenotoxic hepatocarcinogen wy-14,643 is an uncoupler of oxidative phosphorylation,” Toxicology and Applied Pharmacology, vol. 119, no. 1, pp. 52–58, 1993.
[26]
H. Trimarchi, M. R. Mongitore, P. Baglioni et al., “N-acetylcysteine reduces malondialdehyde levels in chronic hemodialysis patients—A pilot study,” Clinical Nephrology, vol. 59, no. 6, pp. 441–446, 2003.
[27]
S. M. Freudenthaler, K. H. Schreeb, A. Wiese, J. Pilz, and C. H. Gleiter, “Influence of controlled hypoxia and radical scavenging agents on erythropoietin and malondialdehyde concentrations in humans,” Acta Physiologica Scandinavica, vol. 174, no. 3, pp. 231–235, 2002.
[28]
T. Toyama, H. Nakamura, Y. Harano et al., “PPARα ligands activate antioxidant enzymes and suppress hepatic fibrosis in rats,” Biochemical and Biophysical Research Communications, vol. 324, no. 2, pp. 697–704, 2004.
[29]
J. C. Fernández-Checa, “Redox regulation and signaling lipids in mitochondrial apoptosis,” Biochemical and Biophysical Research Communications, vol. 304, no. 3, pp. 471–479, 2003.
[30]
M. A. Avila, M. V. Carretero, E. N. Rodriguez, and J. M. Mato, “Regulation by hypoxia of methionine adenosyltransferase activity and gene expression in rat hepatocytes,” Gastroenterology, vol. 114, no. 2, pp. 364–371, 1998.
[31]
S. Khan and P. J. O'Brien, “Rapid and specific efflux of glutathione before hepatocyte injury induced by hypoxia,” Biochemical and Biophysical Research Communications, vol. 238, no. 2, pp. 320–322, 1997.
[32]
P. M. Barger, J. M. Brandt, T. C. Leone, C. J. Weinheimer, and D. P. Kelly, “Deactivation of peroxisome proliferator-activated receptor-α during cardiac hypertrophic growth,” Journal of Clinical Investigation, vol. 105, no. 12, pp. 1723–1730, 2000.
[33]
J. M. Huss, F. H. Levy, and D. P. Kelly, “Hypoxia inhibits the peroxisome proliferator-activated receptor α/retinoid X receptor gene regulatory pathway in cardiac myocytes: a mechanism for O2-dependent modulation of mitochondrial fatty acid oxidation,” Journal of Biological Chemistry, vol. 276, no. 29, pp. 27605–27612, 2001.
[34]
P. Razeghi, M. E. Young, S. Abbasi, and H. Taegtmeyer, “Hypoxia in vivo decreases peroxisome proliferator-activated receptor α-regulated gene expression in rat heart,” Biochemical and Biophysical Research Communications, vol. 287, no. 1, pp. 5–10, 2001.
