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Fructose Mediated Non-Alcoholic Fatty Liver Is Attenuated by HO-1-SIRT1 Module in Murine Hepatocytes and Mice Fed a High Fructose Diet.

Sodhi K, Puri N, Favero G, Stevens S, Meadows C, Abraham NG, Rezzani R, Ansinelli H, Lebovics E, Shapiro JI - PLoS ONE (2015)

Bottom Line: Fructose increased oxidative stress markers and decreased HO-1 and SIRT1 levels in hepatocytes (p<0.05).Increased levels of HO-1 increased SIRT1 levels and ameliorated fructose-mediated lipid accumulation and fibrosis in liver along with decreasing vascular dysfunction (p<0.05 vs. fructose).Taken together, our study demonstrates, for the first time, that HO-1 induction attenuates fructose-induced hepatic lipid deposition, prevents the development of hepatic fibrosis and abates NAFLD-associated vascular dysfunction; effects that are mediated by activation of SIRT1 gene expression.

View Article: PubMed Central - PubMed

Affiliation: Departments of Medicine and Surgery, Joan C. Edwards School of Medicine, Marshall University, Huntington, West Virginia, United States of America.

ABSTRACT

Background: Oxidative stress underlies the etiopathogenesis of nonalcoholic fatty liver disease (NAFLD), obesity and cardiovascular disease (CVD). Heme Oxygenase-1 (HO-1) is a potent endogenous antioxidant gene that plays a key role in decreasing oxidative stress. Sirtuin1 (SIRT1) belongs to the family of NAD-dependent de-acyetylases and is modulated by cellular redox.

Hypothesis: We hypothesize that fructose-induced obesity creates an inflammatory and oxidative environment conducive to the development of NAFLD and metabolic syndrome. The aim of this study is to determine whether HO-1 acts through SIRT1 to form a functional module within hepatocytes to attenuate steatohepatitis, hepatic fibrosis and cardiovascular dysfunction.

Methods and results: We examined the effect of fructose, on hepatocyte lipid accumulation and fibrosis in murine hepatocytes and in mice fed a high fructose diet in the presence and absence of CoPP, an inducer of HO-1, and SnMP, an inhibitor of HO activity. Fructose increased oxidative stress markers and decreased HO-1 and SIRT1 levels in hepatocytes (p<0.05). Further fructose supplementation increased FAS, PPARα, pAMPK and triglycerides levels; CoPP negated this increase. Concurrent treatment with CoPP and SIRT1 siRNA in hepatocytes increased FAS, PPARα, pAMPK and triglycerides levels suggesting that HO-1 is upstream of SIRT1 and suppression of SIRT1 attenuates the beneficial effects of HO-1. A high fructose diet increased insulin resistance, blood pressure, markers of oxidative stress and lipogenesis along with fibrotic markers in mice (p<0.05). Increased levels of HO-1 increased SIRT1 levels and ameliorated fructose-mediated lipid accumulation and fibrosis in liver along with decreasing vascular dysfunction (p<0.05 vs. fructose). These beneficial effects of CoPP were reversed by SnMP.

Conclusion: Taken together, our study demonstrates, for the first time, that HO-1 induction attenuates fructose-induced hepatic lipid deposition, prevents the development of hepatic fibrosis and abates NAFLD-associated vascular dysfunction; effects that are mediated by activation of SIRT1 gene expression.

No MeSH data available.


Related in: MedlinePlus

Effect of CoPP with and without SIRT1-siRNA, and with and without SIRT plasmid on pAMPK, PPARα, FAS expression and triglyceride levels in fructose (Fr)-treated hepatocytes.(A) pAMPK/AMPK expression by western blot analysis. (B) PPARα mRNA levels. (C) FAS mRNA levels measured by RT-PCR in hepatocytes. Results are mean±SE, n = 4/group. * p<0.05 vs CTR; # p<0.05 vs HFr, + p<0.05 vs HFr+CoPP, $ vs Fr+CoPP+SIRT Plasmid. (D) Triglyceride levels measured by RT-PCR in hepatocytes. Results are mean±SE, n = 4/group. * p<0.05 vs CTR; # p<0.05 vs HFr, + p<0.05 vs HFr+CoPP, $ vs Fr+CoPP+SIRT Plasmid.
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pone.0128648.g002: Effect of CoPP with and without SIRT1-siRNA, and with and without SIRT plasmid on pAMPK, PPARα, FAS expression and triglyceride levels in fructose (Fr)-treated hepatocytes.(A) pAMPK/AMPK expression by western blot analysis. (B) PPARα mRNA levels. (C) FAS mRNA levels measured by RT-PCR in hepatocytes. Results are mean±SE, n = 4/group. * p<0.05 vs CTR; # p<0.05 vs HFr, + p<0.05 vs HFr+CoPP, $ vs Fr+CoPP+SIRT Plasmid. (D) Triglyceride levels measured by RT-PCR in hepatocytes. Results are mean±SE, n = 4/group. * p<0.05 vs CTR; # p<0.05 vs HFr, + p<0.05 vs HFr+CoPP, $ vs Fr+CoPP+SIRT Plasmid.

Mentions: To assess whether HO-1 requires the participation of SIRT1 to mediate and/or amplify its actions, we studied the effect of SIRT1 siRNA and SIRT plasmid in hepatocytes treated with fructose. Our results showed that fructose decreased pAMPK and PPARα levels and increased the expression of FAS (Fig 2A, 2B and 2C respectively); this effect of fructose treatment was negated by treatment with CoPP. Interestingly, concurrent treatment with CoPP and SIRT1 siRNA decreased pAMPK and PPARα and increased FAS levels suggesting that HO-1 is upstream of SIRT1 and that suppression of SIRT1 attenuates the beneficial effects of increased levels of HO-1. We also utilized plasmid SIRT1 to assess if increased expression of SIRT1 (in the absence of HO-1 up-regulation) is sufficient to prevent the detrimental effects of HFr on lipid accumulation and metabolic imbalance. Treatment of hepatocytes with fructose, SnMP and SIRT plasmid decreased pAMPK and PPARα and increased FAS levels as compared to hepatocytes treated with fructose, CoPP and SIRT plasmid (Fig 2A, 2B and 2C respectively; p<0.05). In agreement with our hypothesis, our results further showed that hepatocytes treated with fructose, CoPP and SIRT plasmid did not significantly decrease pAMPK, PPARα and FAS levels as compared to cells treated with fructose and CoPP alone indicating a HO-1 dependent activation of SIRT1 expression.


Fructose Mediated Non-Alcoholic Fatty Liver Is Attenuated by HO-1-SIRT1 Module in Murine Hepatocytes and Mice Fed a High Fructose Diet.

Sodhi K, Puri N, Favero G, Stevens S, Meadows C, Abraham NG, Rezzani R, Ansinelli H, Lebovics E, Shapiro JI - PLoS ONE (2015)

Effect of CoPP with and without SIRT1-siRNA, and with and without SIRT plasmid on pAMPK, PPARα, FAS expression and triglyceride levels in fructose (Fr)-treated hepatocytes.(A) pAMPK/AMPK expression by western blot analysis. (B) PPARα mRNA levels. (C) FAS mRNA levels measured by RT-PCR in hepatocytes. Results are mean±SE, n = 4/group. * p<0.05 vs CTR; # p<0.05 vs HFr, + p<0.05 vs HFr+CoPP, $ vs Fr+CoPP+SIRT Plasmid. (D) Triglyceride levels measured by RT-PCR in hepatocytes. Results are mean±SE, n = 4/group. * p<0.05 vs CTR; # p<0.05 vs HFr, + p<0.05 vs HFr+CoPP, $ vs Fr+CoPP+SIRT Plasmid.
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getmorefigures.php?uid=PMC4476565&req=5

pone.0128648.g002: Effect of CoPP with and without SIRT1-siRNA, and with and without SIRT plasmid on pAMPK, PPARα, FAS expression and triglyceride levels in fructose (Fr)-treated hepatocytes.(A) pAMPK/AMPK expression by western blot analysis. (B) PPARα mRNA levels. (C) FAS mRNA levels measured by RT-PCR in hepatocytes. Results are mean±SE, n = 4/group. * p<0.05 vs CTR; # p<0.05 vs HFr, + p<0.05 vs HFr+CoPP, $ vs Fr+CoPP+SIRT Plasmid. (D) Triglyceride levels measured by RT-PCR in hepatocytes. Results are mean±SE, n = 4/group. * p<0.05 vs CTR; # p<0.05 vs HFr, + p<0.05 vs HFr+CoPP, $ vs Fr+CoPP+SIRT Plasmid.
Mentions: To assess whether HO-1 requires the participation of SIRT1 to mediate and/or amplify its actions, we studied the effect of SIRT1 siRNA and SIRT plasmid in hepatocytes treated with fructose. Our results showed that fructose decreased pAMPK and PPARα levels and increased the expression of FAS (Fig 2A, 2B and 2C respectively); this effect of fructose treatment was negated by treatment with CoPP. Interestingly, concurrent treatment with CoPP and SIRT1 siRNA decreased pAMPK and PPARα and increased FAS levels suggesting that HO-1 is upstream of SIRT1 and that suppression of SIRT1 attenuates the beneficial effects of increased levels of HO-1. We also utilized plasmid SIRT1 to assess if increased expression of SIRT1 (in the absence of HO-1 up-regulation) is sufficient to prevent the detrimental effects of HFr on lipid accumulation and metabolic imbalance. Treatment of hepatocytes with fructose, SnMP and SIRT plasmid decreased pAMPK and PPARα and increased FAS levels as compared to hepatocytes treated with fructose, CoPP and SIRT plasmid (Fig 2A, 2B and 2C respectively; p<0.05). In agreement with our hypothesis, our results further showed that hepatocytes treated with fructose, CoPP and SIRT plasmid did not significantly decrease pAMPK, PPARα and FAS levels as compared to cells treated with fructose and CoPP alone indicating a HO-1 dependent activation of SIRT1 expression.

Bottom Line: Fructose increased oxidative stress markers and decreased HO-1 and SIRT1 levels in hepatocytes (p<0.05).Increased levels of HO-1 increased SIRT1 levels and ameliorated fructose-mediated lipid accumulation and fibrosis in liver along with decreasing vascular dysfunction (p<0.05 vs. fructose).Taken together, our study demonstrates, for the first time, that HO-1 induction attenuates fructose-induced hepatic lipid deposition, prevents the development of hepatic fibrosis and abates NAFLD-associated vascular dysfunction; effects that are mediated by activation of SIRT1 gene expression.

View Article: PubMed Central - PubMed

Affiliation: Departments of Medicine and Surgery, Joan C. Edwards School of Medicine, Marshall University, Huntington, West Virginia, United States of America.

ABSTRACT

Background: Oxidative stress underlies the etiopathogenesis of nonalcoholic fatty liver disease (NAFLD), obesity and cardiovascular disease (CVD). Heme Oxygenase-1 (HO-1) is a potent endogenous antioxidant gene that plays a key role in decreasing oxidative stress. Sirtuin1 (SIRT1) belongs to the family of NAD-dependent de-acyetylases and is modulated by cellular redox.

Hypothesis: We hypothesize that fructose-induced obesity creates an inflammatory and oxidative environment conducive to the development of NAFLD and metabolic syndrome. The aim of this study is to determine whether HO-1 acts through SIRT1 to form a functional module within hepatocytes to attenuate steatohepatitis, hepatic fibrosis and cardiovascular dysfunction.

Methods and results: We examined the effect of fructose, on hepatocyte lipid accumulation and fibrosis in murine hepatocytes and in mice fed a high fructose diet in the presence and absence of CoPP, an inducer of HO-1, and SnMP, an inhibitor of HO activity. Fructose increased oxidative stress markers and decreased HO-1 and SIRT1 levels in hepatocytes (p<0.05). Further fructose supplementation increased FAS, PPARα, pAMPK and triglycerides levels; CoPP negated this increase. Concurrent treatment with CoPP and SIRT1 siRNA in hepatocytes increased FAS, PPARα, pAMPK and triglycerides levels suggesting that HO-1 is upstream of SIRT1 and suppression of SIRT1 attenuates the beneficial effects of HO-1. A high fructose diet increased insulin resistance, blood pressure, markers of oxidative stress and lipogenesis along with fibrotic markers in mice (p<0.05). Increased levels of HO-1 increased SIRT1 levels and ameliorated fructose-mediated lipid accumulation and fibrosis in liver along with decreasing vascular dysfunction (p<0.05 vs. fructose). These beneficial effects of CoPP were reversed by SnMP.

Conclusion: Taken together, our study demonstrates, for the first time, that HO-1 induction attenuates fructose-induced hepatic lipid deposition, prevents the development of hepatic fibrosis and abates NAFLD-associated vascular dysfunction; effects that are mediated by activation of SIRT1 gene expression.

No MeSH data available.


Related in: MedlinePlus