SIRT6 is an epigenetic repressor of thoracic aortic aneurysms via inhibiting inflammation and senescence.

Thoracic aortic aneurysms (TAAs) develop asymptomatically and are characterized by dilatation of the aorta. This is considered a life-threating vascular disease due to the risk of aortic rupture and without effective treatments. The current understanding of the pathogenesis of TAA is still limited, especially for sporadic TAAs without known genetic mutation. Sirtuin 6 (SIRT6) expression was significantly decreased in the tunica media of sporadic human TAA tissues. Genetic knockout of Sirt6 in mouse vascular smooth muscle cells accelerated TAA formation and rupture, reduced survival, and increased vascular inflammation and senescence after angiotensin II infusion. Transcriptome analysis identified interleukin (IL)-1ß as a pivotal target of SIRT6, and increased IL-1ß levels correlated with vascular inflammation and senescence in human and mouse TAA samples. Chromatin immunoprecipitation revealed that SIRT6 bound to the Il1b promoter to repress expression partly by reducing the H3K9 and H3K56 acetylation. Genetic knockout of Il1b or pharmacological inhibition of IL-1ß signaling with the receptor antagonist anakinra rescued Sirt6 deficiency mediated aggravation of vascular inflammation, senescence, TAA formation and survival in mice. The findings reveal that SIRT6 protects against TAA by epigenetically inhibiting vascular inflammation and senescence, providing insight into potential epigenetic strategies for TAA treatment.

Vascular Transcriptome Profiling Reveals Aging-Related Genes in Angiotensin â ¡-Induced Hypertensive Mouse Aortas.

Objective Angiotensin Ⅱ (Ang Ⅱ)-induced vascular damage is a major risk of hypertension. However, the underlying molecular mechanism of AngⅡ-induced vascular damage is still unclear. In this study, we explored the novel mechanism associated with Ang II-induced hypertension. Methods We treated 8- to 12-week-old C57BL/6J male mice with saline and Ang Ⅱ(0.72 mg/kg·d) for 28 days, respectively. Then the RNA of the media from the collected mice aortas was extracted for transcriptome sequencing. Principal component analysis was applied to show a clear separation of different samples and the distribution of differentially expressed genes was manifested by Volcano plot. Functional annotations including Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway were performed to reveal the molecular mechanism of Ang Ⅱ-induced hypertension. Finally, the differentially expressed genes were validated by using quantitative real-time PCR. Results The result revealed that a total of 773 genes, including 599 up-regulated genes and 174 down-regulated genes, were differentially expressed in the aorta of Ang Ⅱ-induced hypertension mice model. Functional analysis of differentially expressed genes manifested that various cellular processes may be involved in the Ang Ⅱ-induced hypertension, including some pathways associated with hypertension such as extracellular matrix, inflammation and immune response. Interestingly, we also found that the differentially expressed genes were enriched in vascular aging pathway, and further validated that the expression levels of insulin-like growth factor 1 and adiponectin were significantly increased (P<0.05). Conclusion We identify that vascular aging is involved in Ang Ⅱ-induced hypertension, and insulin-like growth factor 1 and adiponectin may be important candidate genes leading to vascular aging.

Caloric Restriction Induces MicroRNAs to Improve Mitochondrial Proteostasis.

Both caloric restriction (CR) and mitochondrial proteostasis are linked to longevity, but how CR maintains mitochondrial proteostasis in mammals remains elusive. MicroRNAs (miRNAs) are well known for gene silencing in cytoplasm and have recently been identified in mitochondria, but knowledge regarding their influence on mitochondrial function is limited. Here, we report that CR increases miRNAs, which are required for the CR-induced activation of mitochondrial translation, in mouse liver. The ablation of miR-122, the most abundant miRNA induced by CR, or the retardation of miRNA biogenesis via Drosha knockdown significantly reduces the CR-induced activation of mitochondrial translation. Importantly, CR-induced miRNAs cause the overproduction of mtDNA-encoded proteins, which induces the mitochondrial unfolded protein response (UPRmt), and consequently improves mitochondrial proteostasis and function. These findings establish a physiological role of miRNA-enhanced mitochondrial function during CR and reveal miRNAs as critical mediators of CR in inducing UPRmt to improve mitochondrial proteostasis.

Sirt6 regulates efficiency of mouse somatic reprogramming and maintenance of pluripotency.

BACKGROUND: Mouse somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs) by defined factors known to regulate pluripotency, including Oct4, Sox2, Klf4, and c-Myc. It has been reported that Sirtuin 6 (Sirt6), a member of the sirtuin family of NAD+-dependent protein deacetylases, is involved in embryonic stem cell differentiation. However, whether and how Sirt6 influences epigenetic reprogramming remains unknown. METHODS: Mouse embryonic fibroblasts isolated from transgenic Oct4-GFP reporter mice with or without Sirt6 were used for reprogramming by Yamanaka factors. Alkaline phosphatase-positive and OCT4-GFP-positive colony were counted to calculate reprogramming efficiency. OP9 feeder cell co-culture system was used to measure the hematopoietic differentiation from mouse ES and iPS cells. RNA sequencing was measured to identify the differential expressed genes due to loss of Sirt6 in somatic and pluripotent cells. RESULTS: In this study, we provide evidence that Sirt6 is involved in mouse somatic reprogramming. We found that loss of function of Sirt6 could significantly decrease reprogramming efficiency. Furthermore, we showed that Sirt6-null iPS-like cell line has intrinsically a differentiation defect even though the establishment of normal self-renewal. Particularly, by performing transcriptome analysis, we observed that several pluripotent transcriptional factors increase in knockout cell line, which explains the underlying loss of pluripotency in Sirt6-null iPS-like cell line. CONCLUSIONS: Taken together, we have identified a new regulatory role of Sirt6 in reprogramming and maintenance of pluripotency.

Applications of Virus Vector-Mediated Gene Therapy in China.

Due to the increased safety and efficiency of virus vectors, virus vector-mediated gene therapy is now widely used for various diseases, including monogenic diseases, complex disorders, and infectious diseases. Recent gene therapy trials have shown significant therapeutic benefits, and Chinese researchers have contributed significantly to this progress. This review highlights disease applications and strategies for virus vector-mediated gene therapy in preclinical studies and clinical trials in China.

SIRT2 Acts as a Cardioprotective Deacetylase in Pathological Cardiac Hypertrophy.

BACKGROUND: Pathological cardiac hypertrophy induced by stresses such as aging and neurohumoral activation is an independent risk factor for heart failure and is considered a target for the treatment of heart failure. However, the mechanisms underlying pathological cardiac hypertrophy remain largely unknown. We aimed to investigate the roles of SIRT2 in aging-related and angiotensin II (Ang II)-induced pathological cardiac hypertrophy. METHODS: Male C57BL/6J wild-type and Sirt2 knockout mice were subjected to the investigation of aging-related cardiac hypertrophy. Cardiac hypertrophy was also induced by Ang II (1.3 mg/kg/d for 4 weeks) in male C57BL/6J Sirt2 knockout mice, cardiac-specific SIRT2 transgenic (SIRT2-Tg) mice, and their respective littermates (8 to ≈12 weeks old). Metformin (200 mg/kg/d) was used to treat wild-type and Sirt2 knockout mice infused with Ang II. Cardiac hypertrophy, fibrosis, and cardiac function were examined in these mice. RESULTS: SIRT2 protein expression levels were downregulated in hypertrophic hearts from mice. Sirt2 knockout markedly exaggerated cardiac hypertrophy and fibrosis and decreased cardiac ejection fraction and fractional shortening in aged (24-month-old) mice and Ang II-infused mice. Conversely, cardiac-specific SIRT2 overexpression protected the hearts against Ang II-induced cardiac hypertrophy and fibrosis and rescued cardiac function. Mechanistically, SIRT2 maintained the activity of AMP-activated protein kinase (AMPK) in aged and Ang II-induced hypertrophic hearts in vivo as well as in cardiomyocytes in vitro. We identified the liver kinase B1 (LKB1), the major upstream kinase of AMPK, as the direct target of SIRT2. SIRT2 bound to LKB1 and deacetylated it at lysine 48, which promoted the phosphorylation of LKB1 and the subsequent activation of LKB1-AMPK signaling. Remarkably, the loss of SIRT2 blunted the response of AMPK to metformin treatment in mice infused with Ang II and repressed the metformin-mediated reduction of cardiac hypertrophy and protection of cardiac function. CONCLUSIONS: SIRT2 promotes AMPK activation by deacetylating the kinase LKB1. Loss of SIRT2 reduces AMPK activation, promotes aging-related and Ang II-induced cardiac hypertrophy, and blunts metformin-mediated cardioprotective effects. These findings indicate that SIRT2 will be a potential target for therapeutic interventions in aging- and stress-induced cardiac hypertrophy.

Epigenetic regulation in cell senescence.

Cell senescence, which is an irreversible state of cell proliferative arrest, has emerged as a potentially important contributor to tissue dysfunction and organismal ageing. Cell senescence is triggered by a variety of senescence stressors, which affect gene expression and multiple signalling pathways that give rise to various senescence phenotypes. Epigenetic mechanisms, as critical regulators of chromosomal architecture and gene expression, have added an extra dimension to the molecular mechanisms of cell senescence. Cell senescence is accompanied by changes in DNA methylation, histone-associated epigenetic processes, chromatin remodelling and ncRNA expression. Those senescence-associated epigenetic alterations interact with the senescence regulatory programme networks and lead to various cell senescence phenotypes. This review provides a comprehensive overview of epigenetic changes and their effects on cell senescence. The differences in epigenetic alterations among different types of senescence are also discussed. Furthermore, we summarise the interactions among different epigenetic mechanisms during cell senescence and analyse the possibility of using epigenetic signatures as biomarkers and therapeutic targets for the treatment of senescence-associated diseases.

Long noncoding RNA LINC00305 promotes inflammation by activating the AHRR-NF-κB pathway in human monocytes.

Accumulating data from genome-wide association studies (GWAS) have provided a collection of novel candidate genes associated with complex diseases, such as atherosclerosis. We identified an atherosclerosis-associated single-nucleotide polymorphism (SNP) located in the intron of the long noncoding RNA (lncRNA) LINC00305 by searching the GWAS database. Although the function of LINC00305 is unknown, we found that LINC00305 expression is enriched in atherosclerotic plaques and monocytes. Overexpression of LINC00305 promoted the expression of inflammation-associated genes in THP-1 cells and reduced the expression of contractile markers in co-cultured human aortic smooth muscle cells (HASMCs). We showed that overexpression of LINC00305 activated nuclear factor-kappa beta (NF-κB) and that inhibition of NF-κB abolished LINC00305-mediated activation of cytokine expression. Mechanistically, LINC00305 interacted with lipocalin-1 interacting membrane receptor (LIMR), enhanced the interaction of LIMR and aryl-hydrocarbon receptor repressor (AHRR), and promoted protein expression as well as nuclear localization of AHRR. Moreover, LINC00305 activated NF-κB exclusively in the presence of LIMR and AHRR. In light of these findings, we propose that LINC00305 promotes monocyte inflammation by facilitating LIMR and AHRR cooperation and the AHRR activation, which eventually activates NF-κB, thereby inducing HASMC phenotype switching.

The Paraoxonase Gene Cluster Protects Against Abdominal Aortic Aneurysm Formation.

OBJECTIVE: Abdominal aortic aneurysm (AAA) is a life-threatening vascular pathology, the pathogenesis of which is closely related to oxidative stress. However, an effective pharmaceutical treatment is lacking because the exact cause of AAA remains unknown. Here, we aimed at delineating the role of the paraoxonases (PONs) gene cluster (PC), which prevents atherosclerosis through the detoxification of oxidized substrates, in AAA formation. APPROACH AND RESULTS: PC transgenic (Tg) mice were crossed to an Apoe-/- background, and an angiotensin II-induced AAA mouse model was used to analyze the effect of the PC on AAA formation. Four weeks after angiotensin II infusion, PC-Tg Apoe-/- mice had a lower AAA incidence, smaller maximal abdominal aortic external diameter, and less medial elastin degradation than Apoe-/- mice. Importantly, PC-Tg Apoe-/- mice exhibited lower aortic reactive oxidative species production and oxidative stress than did the Apoe-/- control mice. As a consequence, the PC transgene alleviated angiotensin II-induced arterial inflammation and suppressed arterial extracellular matrix degradation. Specifically, on angiotensin II stimulation, PC-Tg vascular smooth muscle cells exhibited lower levels of reactive oxidative species production and a decrease in the activities and expression levels of matrix metalloproteinase-2 and matrix metalloproteinase-9. Moreover, PC-Tg serum also enhanced vascular smooth muscle cell oxidative stress resistance and further decreased the expression levels of matrix metalloproteinase-2 and matrix metalloproteinase-9, indicating that circulatory and vascular smooth muscle cell PC members suppress oxidative stress in a synergistic manner. CONCLUSIONS: Our findings reveal, for the first time, a protective role of the PC in AAA formation and suggest PONs as promising targets for AAA prevention.

SIRT1 deacetylates the cardiac transcription factor Nkx2.5 and inhibits its transcriptional activity.

The homeodomain transcription factor Nkx2.5/Csx is critically essential for heart specification, morphogenesis, and homeostasis. Acetylation/deacetylation is important for the localization, stability and activation of transcription factors. It remains unknown how Nkx2.5 is deacetylated and how Nkx2.5 acetylation determines its activity. In this study, we provide evidence that the NAD+-dependent class III protein deacetylase SIRT1 deacetylates Nkx2.5 in cardiomyocytes and represses the transcriptional activity of Nkx2.5. We show that SIRT1 interacts with the C-terminus of Nkx2.5 and deacetylates Nkx2.5 at lysine 182 in the homeodomain. The mutation of Nkx2.5 at lysine 182 reduces its transcriptional activity. Furthermore, SIRT1 inhibits the transcriptional activity of Nkx2.5 and represses the expression of its target genes partly by reducing Nkx2.5 binding to its co-factors, including SRF and TBX5. Taken together, these findings demonstrate that SIRT1 deacetylates Nkx2.5 and inhibits the transcriptional activity of Nkx2.5.

Age-Associated Sirtuin 1 Reduction in Vascular Smooth Muscle Links Vascular Senescence and Inflammation to Abdominal Aortic Aneurysm.

RATIONALE: Uncontrolled growth of abdominal aortic aneurysms (AAAs) is a life-threatening vascular disease without an effective pharmaceutical treatment. AAA incidence dramatically increases with advancing age in men. However, the molecular mechanisms by which aging predisposes individuals to AAAs remain unknown. OBJECTIVE: In this study, we investigated the role of SIRT1 (Sirtuin 1), a class III histone deacetylase, in AAA formation and the underlying mechanisms linking vascular senescence and inflammation. METHODS AND RESULTS: The expression and activity of SIRT1 were significantly decreased in human AAA samples. SIRT1 in vascular smooth muscle cells was remarkably downregulated in the suprarenal aortas of aged mice, in which AAAs induced by angiotensin II infusion were significantly elevated. Moreover, vascular smooth muscle cell-specific knockout of SIRT1 accelerated angiotensin II-induced formation and rupture of AAAs and AAA-related pathological changes, whereas vascular smooth muscle cell-specific overexpression of SIRT1 suppressed angiotensin II-induced AAA formation and progression in Apoe-/- mice. Furthermore, the inhibitory effect of SIRT1 on AAA formation was also proved in a calcium chloride (CaCl2)-induced AAA model. Mechanistically, the reduction of SIRT1 was shown to increase vascular cell senescence and upregulate p21 expression, as well as enhance vascular inflammation. Notably, inhibition of p21-dependent vascular cell senescence by SIRT1 blocked angiotensin II-induced nuclear factor-κB binding on the promoter of monocyte chemoattractant protein-1 and inhibited its expression. CONCLUSIONS: These findings provide evidence that SIRT1 reduction links vascular senescence and inflammation to AAAs and that SIRT1 in vascular smooth muscle cells provides a therapeutic target for the prevention of AAA formation.

Human paraoxonase gene cluster overexpression alleviates angiotensin II-induced cardiac hypertrophy in mice.

Cardiac hypertrophy is the strongest predictor of the development of heart failure, and anti-hypertrophic treatment holds the key to improving the clinical syndrome and increasing the survival rates for heart failure. The paraoxonase (PON) gene cluster (PC) protects against atherosclerosis and coronary artery diseases. However, the role of PC in the heart is largely unknown. To evaluate the roles of PC in cardiac hypertrophy, transgenic mice carrying the intact human PON1, PON2, and PON3 genes and their flanking sequences were studied. We demonstrated that the PC transgene (PC-Tg) protected mice from cardiac hypertrophy induced by Ang II; these mice had reduced heart weight/body weight ratios, decreased left ventricular wall thicknesses and increased fractional shortening compared with wild-type (WT) control. The same protective tendency was also observed with an Apoe -/- background. Mechanically, PC-Tg normalized the disequilibrium of matrix metalloproteinases (MMPs)/tissue inhibitors of MMPs (TIMPs) in hypertrophic hearts, which might contribute to the protective role of PC-Tg in cardiac fibrosis and, thus, protect against cardiac remodeling. Taken together, our results identify a novel anti-hypertrophic role for the PON gene cluster, suggesting a possible strategy for the treatment of cardiac hypertrophy through elevating the levels of the PON gene family.

Epigenetic regulation of NKG2D ligands is involved in exacerbated atherosclerosis development in Sirt6 heterozygous mice.

Sirt6 is a member of the class III histone deacetylase family which is associated with aging and longevity. Sirt6 deficient mice show an aging-like phenotype, while male transgenic mice of Sirt6 show increased longevity. Sirt6 acts as a tumor suppressor and deficiency of Sirt6 leads to cardiac hypertrophy and heart failure. Whether Sirt6 is involved in atherosclerosis development, the major cause of cardiovascular diseases, is unknown. We found that the expression of Sirt6 is lower in human atherosclerotic plaques than that in controls. When Sirt6(+/-)ApoE(-/-) and ApoE(-/-) mice are fed with high fat diet for 16 weeks, Sirt6(+/-)ApoE(-/-) mice show increased plaque fromation and exhibit feature of plaque instability. Furthermore, Sirt6 downregulation increases expression of NKG2D ligands, which leads to increased cytokine expression. Blocking NKG2D ligand almost completely blocks this effect. Mechanistically, Sirt6 binds to promoters of NKG2D ligand genes and regulates the H3K9 and H3K56 acetylation levels.

Suppression of Mic60 compromises mitochondrial transcription and oxidative phosphorylation.

Precise regulation of mtDNA transcription and oxidative phosphorylation (OXPHOS) is crucial for human health. As a component of mitochondrial contact site and cristae organizing system (MICOS), Mic60 plays a central role in mitochondrial morphology. However, it remains unclear whether Mic60 affects mitochondrial transcription. Here, we report that Mic60 interacts with mitochondrial transcription factors TFAM and TFB2M. Furthermore, we found that Mic60 knockdown compromises mitochondrial transcription and OXPHOS activities. Importantly, Mic60 deficiency decreased TFAM binding and mitochondrial RNA polymerase (POLRMT) recruitment to the mtDNA promoters. In addition, through mtDNA immunoprecipitation (mIP)-chromatin conformation capture (3C) assays, we found that Mic60 interacted with mtDNA and was involved in the architecture of mtDNA D-loop region. Taken together, our findings reveal a previously unrecognized important role of Mic60 in mtDNA transcription.

SIRT1-mediated epigenetic downregulation of plasminogen activator inhibitor-1 prevents vascular endothelial replicative senescence.

The inactivation of plasminogen activator inhibitor-1 (PAI-1) has been shown to exert beneficial effects in age-related vascular diseases. Limited information is available on the molecular mechanisms regarding the negatively regulated expression of PAI-1 in the vascular system. In this study, we observed an inverse correlation between SIRT1, a class III histone deacetylase, and PAI-1 expression in human atherosclerotic plaques and the aortas of old mice, suggesting that internal negative regulation exists between SIRT1 and PAI-1. SIRT1 overexpression reversed the increased PAI-1 expression in senescent human umbilical vein endothelial cells (HUVECs) and aortas of old mice, accompanied by decreased SA-ß-gal activity in vitro and improved endothelial function and reduced arterial stiffness in vivo. Moreover, the SIRT1-mediated inhibition of PAI-1 expression exerted an antisenescence effect in HUVECs. Furthermore, we demonstrated that SIRT1 is able to bind to the PAI-1 promoter, resulting in a decrease in the acetylation of histone H4 lysine 16 (H4K16) on the PAI-1 promoter region. Thus, our findings suggest that the SIRT1-mediated epigenetic inhibition of PAI-1 expression exerts a protective effect in vascular endothelial senescence.

The involvement of NFAT transcriptional activity suppression in SIRT1-mediated inhibition of COX-2 expression induced by PMA/Ionomycin.

SIRT1, a class III histone deacetylase, acts as a negative regulator for many transcription factors, and plays protective roles in inflammation and atherosclerosis. Transcription factor nuclear factor of activated T cells (NFAT) has been previously shown to play pro-inflammatory roles in endothelial cells. Inhibition of NFAT signaling may be an attractive target to regulate inflammation in atherosclerosis. However, whether NFAT transcriptional activity is suppressed by SIRT1 remains unknown. In this study, we found that SIRT1 suppressed NFAT-mediated transcriptional activity. SIRT1 interacted with NFAT, and the NHR and RHR domains of NFAT mediated the interaction with SIRT1. Moreover, we found that SIRT1 primarily deacetylated NFATc3. Adenoviral over-expression of SIRT1 suppressed PMA and calcium ionophore Ionomycin (PMA/Io)-induced COX-2 expression in human umbilical vein endothelial cells (HUVECs), while SIRT1 RNAi reversed the effects in HUVECs. Moreover, inhibition of COX-2 expression by SIRT1 in PMA/Io-treated HUVECs was largely abrogated by inhibiting NFAT activation. Furthermore, SIRT1 inhibited NFAT-induced COX-2 promoter activity, and reduced NFAT binding to the COX-2 promoter in PMA/Io-treated HUVECs. These results suggest that suppression of NFAT transcriptional activity is involved in SIRT1-mediated inhibition of COX-2 expression induced by PMA/Io, and that the negative regulatory mechanisms of NFAT by SIRT1 may contribute to its anti-inflammatory effects in atherosclerosis.

SIRT1 mediates the protective function of Nkx2.5 during stress in cardiomyocytes.

Nkx2.5 plays protective roles in cardiac homeostasis and survival in the postnatal hearts. However, the underlying molecular mechanisms that mediate the protective functions of Nkx2.5 remain unknown. Here, we showed that Nkx2.5 was downregulated in response to various stresses and was required for protection against the stress-induced apoptosis of cardiomyocytes. SIRT1, a member of the sirtuin family of proteins, was found to be a direct transcriptional target of Nkx2.5 and was required for the Nkx2.5-mediated protection of cardiomyocytes from doxorubicin (DOX)-induced apoptosis. Moreover, using chromatin immunoprecipitation assays, we found that Nkx2.5 was able to bind to the SIRT1 promoter and that this binding was significantly decreased in DOX-treated mouse hearts. Furthermore, the cardiac-specific overexpression of SIRT1 decreased the DOX-induced apoptosis of cardiomyocytes in SIRT1 transgenic mouse hearts compared with the hearts of their wild-type littermates. These findings demonstrate that SIRT1 acts as a direct transcriptional target of Nkx2.5 that maintains cardiomyocyte homeostasis and survival.

Up-regulation of Fas ligand expression by sirtuin 1 in both flow-restricted vessels and serum-stimulated vascular smooth muscle cells.

OBJECTIVE: To study the role of sirtuin 1 (SIRT1) in Fas ligand (FasL) expression regulation during vascular lesion formation and to elucidate the potential mechanisms. METHODS: SIRT1 and FasL protein levels were detected by Western blotting in either mouse arteries extract or the whole rat aortic vascular smooth muscle cell (VSMC) lysate. Smooth muscle cell (SMC)-specific human SIRT1 transgenic (Tg) C57BL/6 mice and their littermate wild-type (WT) controls underwent complete carotid artery ligation (ligation groups) or the ligation-excluded operation (sham groups). The carotid arteries were collected 1 day after operation. Reverse transcription-polymerase chain reaction was performed to detect the mRNA levels of SIRT1 and FasL. Luciferase reporter assays were performed to detect the effect of WT-SIRT1, a dominant-negative form of SIRT1 (SIRT1H363Y), and GATA-6 on the promoter activity of FasL. Flow cytometry assay was applied to measure the hypodiploid DNA content of VSMC so as to monitor cellular apoptosis. RESULTS: SIRT1 was expressed in both rat aortic VSMCs and mouse arteries. Forced SIRT1 expression increased FasL expression both in injured mouse carotid arteries 1 day after ligation (P<0.001) and VSMCs treated with serum (P<0.05 at the transcriptional level, P<0.001 at the protein level). No notable apoptosis was observed. Furthermore, transcription factor GATA-6 increased the promoter activity of FasL (P<0.001). The induction of FasL promoter activity by GATA-6 was enhanced by WT-SIRT1 (P<0.001), while SIRT1H363Y significantly relieved the enhancing effect of WT-SIRT1 on GATA-6 (P<0.001). CONCLUSIONS: Overexpression of SIRT1 up-regulates FasL expression in both flow-restricted mouse carotid arteries and serum-stimulated VSMCs. The transcription factor GATA-6 participates in the transcriptional regulation of FasL expression by SIRT1.