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AAV8-mediated Sirt1 gene transfer to the liver prevents high carbohydrate diet-induced nonalcoholic fatty liver disease.

Vilà L, Elias I, Roca C, Ribera A, Ferré T, Casellas A, Lage R, Franckhauser S, Bosch F - Mol Ther Methods Clin Dev (2014)

Bottom Line: Long-term hepatic SIRT1 overexpression led to upregulation of key hepatic genes involved in β-oxidation, prevented HC diet-induced lipid accumulation and reduced liver inflammation.AAV8-Sirt1-treated mice showed improved insulin sensitivity, increased oxidative capacity in skeletal muscle and reduced white adipose tissue inflammation.Moreover, HC feeding induced leptin resistance, which was also attenuated in AAV8-Sirt1-treated mice.

View Article: PubMed Central - PubMed

Affiliation: Center of Animal Biotechnology and Gene Therapy, School of Veterinary Medicine, Universitat Autònoma de Barcelona , Bellaterra, Spain ; Department of Biochemistry and Molecular Biology, School of Veterinary Medicine, Universitat Autònoma de Barcelona , Bellaterra, Spain ; Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM) , Barcelona, Spain.

ABSTRACT
Nonalcoholic fatty liver disease (NAFLD) is the most common hepatic disease worldwide, and evidence suggests that it promotes insulin resistance and type 2 diabetes. Caloric restriction (CR) is the only available strategy for NAFLD treatment. The protein deacetylase Sirtuin1 (SIRT1), which is activated by CR, increases catabolic metabolism and decreases lipogenesis and inflammation, both involved in the development of NAFLD. Here we show that adeno-associated viral vectors of serotype 8 (AAV8)-mediated liver-specific Sirt1 gene transfer prevents the development of NAFLD induced by a high carbohydrate (HC) diet. Long-term hepatic SIRT1 overexpression led to upregulation of key hepatic genes involved in β-oxidation, prevented HC diet-induced lipid accumulation and reduced liver inflammation. AAV8-Sirt1-treated mice showed improved insulin sensitivity, increased oxidative capacity in skeletal muscle and reduced white adipose tissue inflammation. Moreover, HC feeding induced leptin resistance, which was also attenuated in AAV8-Sirt1-treated mice. Therefore, AAV-mediated gene transfer to overexpress SIRT1 specifically in the liver may represent a new gene therapy strategy to counteract NAFLD and related diseases such as type 2 diabetes.

No MeSH data available.


Related in: MedlinePlus

AAV8-Sirt1–treated mice showed increased β-oxidation capacity in skeletal muscle. (a) Triglyceride content in skeletal muscle. (b) Expression levels of Mitochondrial uncoupling protein 2 (Ucp2), Mitochondrial uncoupling protein 3 (Ucp3), Peroxisome proliferator activated receptor α (Pparα), Peroxisome proliferator activated receptor δ (Ppard), Carnitinepalmitoyltransferase 2 (Cpt2), Carnitinepalmitoyltransferase 1 (mCpt1), Peroxisome proliferative activated receptor, gamma, coactivator 1 α (Ppargc1α), and Pyruvate deshydrogenase kinase isoenzyme 4 (Pdk4). (c) Representative western blots and quantifications are shown for total AMP protein kinase (AMPKtot), phosphorylated AMPK (AMPK-P), phosphorylated Acetyl-CoA Carboxylase (ACC-P) and tubulin protein levels in skeletal muscle of AAV8-Null and AAV8-Sirt1 mice. Tubulin was used as a loading control. All analyses were performed after 15 weeks on HC diet. Data represent the mean ± SEM of at least four animals per group. *P < 0.05 and **P < 0.01 versus AAV8-Null. a.u., arbitrary units.
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fig5: AAV8-Sirt1–treated mice showed increased β-oxidation capacity in skeletal muscle. (a) Triglyceride content in skeletal muscle. (b) Expression levels of Mitochondrial uncoupling protein 2 (Ucp2), Mitochondrial uncoupling protein 3 (Ucp3), Peroxisome proliferator activated receptor α (Pparα), Peroxisome proliferator activated receptor δ (Ppard), Carnitinepalmitoyltransferase 2 (Cpt2), Carnitinepalmitoyltransferase 1 (mCpt1), Peroxisome proliferative activated receptor, gamma, coactivator 1 α (Ppargc1α), and Pyruvate deshydrogenase kinase isoenzyme 4 (Pdk4). (c) Representative western blots and quantifications are shown for total AMP protein kinase (AMPKtot), phosphorylated AMPK (AMPK-P), phosphorylated Acetyl-CoA Carboxylase (ACC-P) and tubulin protein levels in skeletal muscle of AAV8-Null and AAV8-Sirt1 mice. Tubulin was used as a loading control. All analyses were performed after 15 weeks on HC diet. Data represent the mean ± SEM of at least four animals per group. *P < 0.05 and **P < 0.01 versus AAV8-Null. a.u., arbitrary units.

Mentions: AAV8-Sirt1 mice showed a trend to accumulate less triglycerides in skeletal muscle than AAV8-Null mice (Figure 5a), suggesting that β-oxidation could be upregulated in this tissue following the treatment. In agreement with this hypothesis, the expression of key genes involved in lipid oxidation such as Mitochondrial uncoupling protein 2 (Ucp2), Peroxisome proliferator activated receptor δ (Ppard) and Pyruvate dehydrogenase kinase isoenzyme 4 (Pdk4) was increased in AAV8-Sirt1–treated mice (Figure 5b). In addition, the levels of phosphorylated AMP activated kinase (AMPK) were approximately 50% higher in mice receiving AAV8-Sirt1 vectors versus those receiving AAV8-Null (Figure 5c). The phosphorylation of acetyl-CoA carboxylase, one of the targets of AMPK, was also augmented in AAV8-Sirt1–treated mice (Figure 5c). Thus, these results suggested that treatment with AAV8-Sirt1 vectors increased β-oxidation in the skeletal muscle.


AAV8-mediated Sirt1 gene transfer to the liver prevents high carbohydrate diet-induced nonalcoholic fatty liver disease.

Vilà L, Elias I, Roca C, Ribera A, Ferré T, Casellas A, Lage R, Franckhauser S, Bosch F - Mol Ther Methods Clin Dev (2014)

AAV8-Sirt1–treated mice showed increased β-oxidation capacity in skeletal muscle. (a) Triglyceride content in skeletal muscle. (b) Expression levels of Mitochondrial uncoupling protein 2 (Ucp2), Mitochondrial uncoupling protein 3 (Ucp3), Peroxisome proliferator activated receptor α (Pparα), Peroxisome proliferator activated receptor δ (Ppard), Carnitinepalmitoyltransferase 2 (Cpt2), Carnitinepalmitoyltransferase 1 (mCpt1), Peroxisome proliferative activated receptor, gamma, coactivator 1 α (Ppargc1α), and Pyruvate deshydrogenase kinase isoenzyme 4 (Pdk4). (c) Representative western blots and quantifications are shown for total AMP protein kinase (AMPKtot), phosphorylated AMPK (AMPK-P), phosphorylated Acetyl-CoA Carboxylase (ACC-P) and tubulin protein levels in skeletal muscle of AAV8-Null and AAV8-Sirt1 mice. Tubulin was used as a loading control. All analyses were performed after 15 weeks on HC diet. Data represent the mean ± SEM of at least four animals per group. *P < 0.05 and **P < 0.01 versus AAV8-Null. a.u., arbitrary units.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4362360&req=5

fig5: AAV8-Sirt1–treated mice showed increased β-oxidation capacity in skeletal muscle. (a) Triglyceride content in skeletal muscle. (b) Expression levels of Mitochondrial uncoupling protein 2 (Ucp2), Mitochondrial uncoupling protein 3 (Ucp3), Peroxisome proliferator activated receptor α (Pparα), Peroxisome proliferator activated receptor δ (Ppard), Carnitinepalmitoyltransferase 2 (Cpt2), Carnitinepalmitoyltransferase 1 (mCpt1), Peroxisome proliferative activated receptor, gamma, coactivator 1 α (Ppargc1α), and Pyruvate deshydrogenase kinase isoenzyme 4 (Pdk4). (c) Representative western blots and quantifications are shown for total AMP protein kinase (AMPKtot), phosphorylated AMPK (AMPK-P), phosphorylated Acetyl-CoA Carboxylase (ACC-P) and tubulin protein levels in skeletal muscle of AAV8-Null and AAV8-Sirt1 mice. Tubulin was used as a loading control. All analyses were performed after 15 weeks on HC diet. Data represent the mean ± SEM of at least four animals per group. *P < 0.05 and **P < 0.01 versus AAV8-Null. a.u., arbitrary units.
Mentions: AAV8-Sirt1 mice showed a trend to accumulate less triglycerides in skeletal muscle than AAV8-Null mice (Figure 5a), suggesting that β-oxidation could be upregulated in this tissue following the treatment. In agreement with this hypothesis, the expression of key genes involved in lipid oxidation such as Mitochondrial uncoupling protein 2 (Ucp2), Peroxisome proliferator activated receptor δ (Ppard) and Pyruvate dehydrogenase kinase isoenzyme 4 (Pdk4) was increased in AAV8-Sirt1–treated mice (Figure 5b). In addition, the levels of phosphorylated AMP activated kinase (AMPK) were approximately 50% higher in mice receiving AAV8-Sirt1 vectors versus those receiving AAV8-Null (Figure 5c). The phosphorylation of acetyl-CoA carboxylase, one of the targets of AMPK, was also augmented in AAV8-Sirt1–treated mice (Figure 5c). Thus, these results suggested that treatment with AAV8-Sirt1 vectors increased β-oxidation in the skeletal muscle.

Bottom Line: Long-term hepatic SIRT1 overexpression led to upregulation of key hepatic genes involved in β-oxidation, prevented HC diet-induced lipid accumulation and reduced liver inflammation.AAV8-Sirt1-treated mice showed improved insulin sensitivity, increased oxidative capacity in skeletal muscle and reduced white adipose tissue inflammation.Moreover, HC feeding induced leptin resistance, which was also attenuated in AAV8-Sirt1-treated mice.

View Article: PubMed Central - PubMed

Affiliation: Center of Animal Biotechnology and Gene Therapy, School of Veterinary Medicine, Universitat Autònoma de Barcelona , Bellaterra, Spain ; Department of Biochemistry and Molecular Biology, School of Veterinary Medicine, Universitat Autònoma de Barcelona , Bellaterra, Spain ; Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERDEM) , Barcelona, Spain.

ABSTRACT
Nonalcoholic fatty liver disease (NAFLD) is the most common hepatic disease worldwide, and evidence suggests that it promotes insulin resistance and type 2 diabetes. Caloric restriction (CR) is the only available strategy for NAFLD treatment. The protein deacetylase Sirtuin1 (SIRT1), which is activated by CR, increases catabolic metabolism and decreases lipogenesis and inflammation, both involved in the development of NAFLD. Here we show that adeno-associated viral vectors of serotype 8 (AAV8)-mediated liver-specific Sirt1 gene transfer prevents the development of NAFLD induced by a high carbohydrate (HC) diet. Long-term hepatic SIRT1 overexpression led to upregulation of key hepatic genes involved in β-oxidation, prevented HC diet-induced lipid accumulation and reduced liver inflammation. AAV8-Sirt1-treated mice showed improved insulin sensitivity, increased oxidative capacity in skeletal muscle and reduced white adipose tissue inflammation. Moreover, HC feeding induced leptin resistance, which was also attenuated in AAV8-Sirt1-treated mice. Therefore, AAV-mediated gene transfer to overexpress SIRT1 specifically in the liver may represent a new gene therapy strategy to counteract NAFLD and related diseases such as type 2 diabetes.

No MeSH data available.


Related in: MedlinePlus