Article

Leptin is key to peroxynitrite-mediated oxidative stress and Kupffer cell activation in experimental nonalcoholic steatohepatitis.

Environmental Health and Disease Laboratory, Department of Environmental Health Sciences, University of South Carolina, Columbia 29208 USA
Journal of Hepatology (Impact Factor: 10.4). 11/2012; DOI: 10.1016/j.jhep.2012.11.035
Source: PubMed

ABSTRACT BACKGROUND AND AIMS: Progression from steatosis to steatohepatitic lesions is hypothesized to require a second hit. These lesions have been associated with increased oxidative stress, often ascribed to high levels of leptin and other proinflammatory mediators. Here we have examined the role of leptin in inducing oxidative stress and Kupffer cell activation in CCl(4)-mediated steatohepatitic lesions of obese mice. METHODS: Male C57BL/6 mice fed with a high fat diet (60%kcal) at 16 weeks were administered CCl(4) to induce steatohepatitic lesions. Approaches included use of immuno-spin trapping for measuring free radical stress, gene-deficient mice for leptin, p47 phox, iNOS and adoptive transfer of leptin primed macrophages in vivo. RESULTS: Diet-induced obese (DIO) mice, treated with CCl(4) increased serum leptin levels. Oxidative stress was significantly elevated in DIO mice liver but not in OB/OB mice, or in DIO mice treated with leptin antibody. In OB/OB mice, leptin supplementation restored markers of free radical generation. Markers of free radical formation were significantly decreased by the peroxynitrite decomposition catalyst FeTPPS, the iNOS inhibitor 1400W, the NADPH oxidase inhibitor apocynin, or in iNOS or p47 phox-deficient mice. These results correlated with the decreased expression of TNF-alpha and MCP-1. Kupffer cell depletion eliminated oxidative stress and inflammation, whereas in macrophage-depleted mice, the adoptive transfer of leptin-primed macrophages significantly restored inflammation. CONCLUSIONS: These results, for the first time, suggest that leptin action in macrophages of steatotic liver through induction of iNOS and NADPH oxidase caused peroxynitrite-mediated oxidative stress thus activating Kupffer cells.

2 Followers
 · 
120 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: Altered epididymal sperm count and morphology following leptin treatment has been reported recently. This study examined the effects of 42 days of leptin treatment on sperm count and morphology and their reversibility during a subsequent 56-day recovery period. Twelve-week-old male Sprague-Dawley rats were randomised into four leptin and four saline-treated control groups (n = 6). Intraperitoneal injections of leptin were given daily (60 μg Kg−1 body weight) for 42 days. Controls received 0.1 ml of 0.9% saline. Leptin-treated animals and their respective age-matched controls were euthanised on either day 1, 21, 42 or 56 of recovery for collection of epididymal spermatozoa. Sperm concentration was determined using a Makler counting chamber. Spermatozoa were analysed for 8-hydroxy-2-deoxyguanosine and DNA fragmentation (Comet assay). Data were analysed using anova. Sperm concentration was significantly lower but fraction of abnormal spermatozoa, and levels of 8-hydroxy-2-deoxyguanosine were significantly higher in leptin-treated rats on day 1 of recovery. Comet assays revealed significant DNA fragmentation in leptin-treated rats. These differences were reduced by day 56 of recovery. It appears that 42 days of leptin treatment to Sprague-Dawley rats has significant adverse effects on sperm count and morphology that reverse following discontinuation of leptin treatment.
    Andrologia 09/2014; DOI:10.1111/and.12325 · 1.17 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatic fibrosis in NASH is the common pathophysiologic process resulting from chronic liver inflammation and oxidative stress. Though significant research has been carried out on the role of leptin induced NADPH oxidase in fibrogenesis, the molecular mechanisms that connect leptin-NADPH oxidase axis in upregulation of TGF-β signaling has been unclear. We aimed to investigate the role of leptin mediated upregulation of NADPH oxidase and its subsequent induction of miR21 in fibrogenesis. Human NASH livers and a high fat (60% kCal) diet fed chronic mouse model where hepatotoxin bromodichloromethane was used to induce NASH, were used for this study. To prove the role of Leptin-NADPH oxidase-miR21 axis, mouse deficient in genes for leptin, p47phox and miR21 were used. Results showed that wild type mice and human livers with NASH had increased oxidative stress, increased p47phox expression, augmented NF-kB activation and increased miR21 levels. These mice and human livers showed increased TGF-β, SMAD2/3-SMAD4 colocalizations in the nucleus, increased Col1α and α-SMA expression with a concomitant decrease in protein levels of SMAD7. Mice that were deficient in leptin or p47phox had decreased activated NF-kB and miR21 levels suggesting the role of leptin and NADPH oxidase in inducing NF-kB mediated miR21 expression. Further miR21 KO mice had decreased co-localization events of SMAD2/3-SMAD4 in the nucleus, increased SMAD7 levels and decreased fibrogenesis. Taken together the studies show the novel role of leptin-NADPH oxidase induction of miR21 as a key regulator of TGF-β signaling and fibrogenesis in experimental and human NASH. Copyright © 2014, American Journal of Physiology- Gastrointestinal and Liver Physiology.
    AJP Gastrointestinal and Liver Physiology 12/2014; 308(4):ajpgi.00346.2014. DOI:10.1152/ajpgi.00346.2014 · 3.74 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Activation of M1 macrophages in the nonalcoholic steatohepatitis (NASH) following several external or endogenous factors viz inflammatory stimuli, oxidative stress and cytokines are known. However, any direct role of oxidative stress in causing M1 polarization in NASH has been unclear. We hypothesized that CYP2E1-mediated oxidative stress causes M1 polarization in experimental NASH and NO donor administration inhibits CYP2E1 mediated inflammation with concomitant attenuation of M1 polarization. Since CYP2E1 takes center stage in these studies we use a toxin model of NASH which uses a ligand and a substrate of CYP2E1 for inducing NASH. Subsequently we use a methionine and choline deficient diet induced rodent NASH model where CYP2E1 role in disease progression has been shown. Results show that CYP2E1 causes M1 polarization bias that includes a significant increase in IL-1β and IL-12 in both models of NASH while CYP2E1 null mice or diallyl sulfide administration prevented it. Administration of GdCl3, a macrophage toxin attenuated both the initial M1 response and subsequent M2 response showing the observed increase in cytokine levels is primarily from macrophages. Based on the evidence of an adaptive NO increase, NO donor administration in vivo, that mechanistically inhibited CYP2E1 catalyzed oxidative stress during the entire study in NASH abrogated M1 polarization and NASH progression. The results obtained show the association of CYP2E1 in M1 polarization and that inhibition of CYP2E1 catalyzed oxidative stress by NO donor (DETA NONOate) can be a promising therapeutic strategy in NASH.
    Journal of Pharmacology and Experimental Therapeutics 10/2014; DOI:10.1124/jpet.114.218131 · 3.86 Impact Factor