SIRT3 protects kidneys from ischemia-reperfusion injury by modulating the DRP1 pathway to induce mitochondrial autophagy.

Renal ischemia-reperfusion injury (IRI) is a leading cause of acute kidney injury (AKI) and may influence renal graft survival. In this study, we investigate the involvement of SIRT3 and DRP1 in mitochondrial autophagy and AKI in a mouse model of IRI. Autophagy was detected in the absence of SIRT3, and hypoxic reoxygenation (H/R) experiments using renal tubular epithelial cells NRK52E were performed in vitro to validate these results. We found that autophagosomes increased following IRI and that the expression of autophagy-related genes was up-regulated. The inhibition of autophagy with 3-methyladenine exacerbated IRI, whereas the DRP1 inhibitor Mdivi-1 reversed this inhibition. Mdivi-1 did not reverse the inhibition of autophagy in the absence of SIRT3. During IRI, Mdivi-1 reduced autophagy and DRP1 expression, whereas SIRT3 overexpression attenuated this condition. Rescue experiment showed that autophagy was increased when both SIRT3 or DRP1 were over- or under-expressed or just DRP1 was under-expressed but expression was reduced when just SIRT3 was under-expressed. However, the expression of DRP1-related molecules was reduced when SIRT3 was overexpressed and when DRP1 was under-expressed. Taken together, these findings indicate that SIRT3 protects against kidney damage from IRI by modulating the DRP1 pathway to induce mitochondrial autophagy.

Toll-Like Receptor 4 Mediated Oxidized Low-Density Lipoprotein-Induced Foam Cell Formation in Vascular Smooth Muscle Cells via Src and Sirt1/3 Pathway.

Oxidized low-density lipoprotein (oxLDL) induced a foam-cell-like phenotype of the vascular smooth muscle cells (VSMCs), leading to the inflammatory responses incorporating Toll-like receptor- (Tlr-) mediated cellular alterations. However, the role of Tlr4 in foam cell formation and underlying molecular pathways has not been comprehensively elucidated. To further investigate the mechanism, VSMCs were incubated with different doses of oxLDL, and then, the lipid, reactive oxygen species (ROS) accumulation, Tlr family genes, and the foam cell phenotype were explored. We observed that oxLDL induced foam cell-like phenotype in VSMCs and led to lipid and ROS accumulation in a dose-dependent manner. Furthermore, in the Tlr family, Tlr4 demonstrated the strongest upregulation under oxLDL stimulation. Simultaneously, oxLDL induced activation of Src, higher expression of Nox2, and lower expression of Mnsod, Sirt1, and Sirt3. By interfering the TLR4 expression, the phenotype alteration, lipid accumulation in VSMCs, and Src kinase activation induced by oxLDL were abolished. After interfering Src activation, the oxLDL-induced lipid accumulation and foam cell phenotype in VSMCs were also alleviated. Furthermore, the ROS accumulation, upregulated Nox2 expression, downregulated Sirt1, Sirt3, and Mnsod expression in VSMCs under oxLDL stimulation were also relieved after the knockdown of Tlr4. Additionally, overexpression of Sirt1 and Sirt3 ameliorated the ROS accumulation and foam cell-like marker expression in VSMCs. These results demonstrated that beyond its familiar role in regulating inflammation response, Tlr4 is a critical regulator in oxLDL-induced foam cell formation in VSMCs via regulating Src kinase activation as well as Sirt1 and Sirt3 expression.

SIRT3 consolidates heterochromatin and counteracts senescence.

Sirtuin 3 (SIRT3) is an NAD+-dependent deacetylase linked to a broad range of physiological and pathological processes, including aging and aging-related diseases. However, the role of SIRT3 in regulating human stem cell homeostasis remains unclear. Here we found that SIRT3 expression was downregulated in senescent human mesenchymal stem cells (hMSCs). CRISPR/Cas9-mediated depletion of SIRT3 led to compromised nuclear integrity, loss of heterochromatin and accelerated senescence in hMSCs. Further analysis indicated that SIRT3 interacted with nuclear envelope proteins and heterochromatin-associated proteins. SIRT3 deficiency resulted in the detachment of genomic lamina-associated domains (LADs) from the nuclear lamina, increased chromatin accessibility and aberrant repetitive sequence transcription. The re-introduction of SIRT3 rescued the disorganized heterochromatin and the senescence phenotypes. Taken together, our study reveals a novel role for SIRT3 in stabilizing heterochromatin and counteracting hMSC senescence, providing new potential therapeutic targets to ameliorate aging-related diseases.

Sirtuin-3 mediates sex differences in kidney ischemia-reperfusion injury.

Studies suggest that biological sex influences susceptibility to kidney diseases with males demonstrating greater risk for developing ischemic acute kidney injury (AKI). Sex-related differences in mitochondrial function and homeostasis exist, likely contributing to sexual dimorphism in kidney injury, but the mechanisms are not well characterized. Our observations reveal lower baseline expression of Sirtuin-3 (Sirt3, a major mitochondrial acetyltransferase) in the kidneys of male mice versus females. We tested the hypothesis that differential expression of kidney Sirt3 may mediate sexual dimorphism in AKI using a bilateral kidney ischemia-reperfusion injury (IRI) model and three transgenic mouse models: (1) mice with global transgenic overexpression of Sirt3; (2) mice with inducible, kidney tubule-specific Sirt3 knockdown (iKD); and (3) mice with global Sirt3 knockout. Low mitochondrial Sirt3 (mtSirt3) in males versus females is associated with development of kidney tubular epithelium vacuoles, increased mitochondrial ROS and susceptibility to IRI. Transgenic overexpression of Sirt3 in males protects against kidney IRI and development of tubular epithelium vacuoles. In both sexes, mice with partial kidney tubular epithelium-specific Sirt3 knockdown display intermediate - while global Sirt3 knockout mice display the highest susceptibility to IRI. Female Sirt3 iKD mice demonstrate decreased survival and kidney function after IRI indistinguishable from control males, abolishing the protective effects observed in females. Mechanistically, observed differences in kidney mtSirt3 are sex hormone-dependent; estradiol increases - while testosterone decreases mtSirt3 protein. Our results demonstrate that Sirt3 is an important contributor to the observed sex-related differences in IRI susceptibility, and a potential therapeutic target in the clinical management of AKI.

SIRT3 ameliorates osteoarthritis via regulating chondrocyte autophagy and apoptosis through the PI3K/Akt/mTOR pathway.

Osteoarthritis (OA) is the most common form of joint disease. The aim of this study was to explore the functions of SIRT3 on OA pathophysiology and the mechanism involved. Rat chondrocytes and destabilized medial meniscus (DMM) rat OA model were used as in vitro and in vivo models. In addition, lentivirus and plasmid were used to overexpress SIRT3, while siRNA was applied to establish SIRT3 knockdown. IL-1ß induced inflammation, apoptosis, mitochondrial dysfunction, and chondrocyte degeneration were inhibited by SIRT3 overexpression, which were enhanced in SIRT3-knockdown rat chondrocytes. Furthermore, overexpression of SIRT3 could restore IL-1ß-induced autophagy inhibition. We also found that IL-1ß-induced PI3K/Akt/mTOR signaling pathway activation was inhibited by SIRT3 overexpression, which was enhanced by SIRT3 knockdown. Last, intra-articular SIRT3 overexpression alleviated the severity of OA-induced rat joint damage. Our results demonstrated that SIRT3 is an important protective agent against OA pathophysiology via inhibiting PI3K/Akt/mTOR signaling.

Sirtuin 3 Downregulation in Mycobacterium tuberculosis-Infected Macrophages Reprograms Mitochondrial Metabolism and Promotes Cell Death.

Mycobacterium tuberculosis induces metabolic reprogramming in macrophages like the Warburg effect. This enhances antimicrobial performance at the expense of increased inflammation, which may promote a pathogen-permissive host environment. Since the NAD+-dependent protein deacetylase Sirtuin 3 (SIRT3) is an important regulator of mitochondrial metabolism and cellular redox homeostasis, we hypothesized that SIRT3 modulation mediates M. tuberculosis-induced metabolic reprogramming. Infection of immortalized and primary murine macrophages resulted in reduced levels of SIRT3 mRNA and protein and perturbation of SIRT3-regulated enzymes in the tricarboxylic acid cycle, electron transport chain, and glycolytic pathway. These changes were associated with increased reactive oxygen species and reduced antioxidant scavenging, thereby triggering mitochondrial stress and macrophage cell death. Relevance to tuberculosis disease in vivo was indicated by greater bacterial burden and immune pathology in M. tuberculosis-infected Sirt3-/- mice. CD11b+ lung leukocytes isolated from infected Sirt3-/- mice showed decreased levels of enzymes involved in central mitochondrial metabolic pathways, along with increased reactive oxygen species. Bacterial burden was also greater in lungs of LysMcreSirt3L2/L2 mice, demonstrating the importance of macrophage-specific SIRT3 after infection. These results support the model of SIRT3 as a major upstream regulatory factor, leading to metabolic reprogramming in macrophages by M. tuberculosisIMPORTANCE Tuberculosis, the disease caused by the bacterium M. tuberculosis, remains one of the top 10 causes of death worldwide. Macrophages, the first cells to encounter M. tuberculosis and critical for defense against infection, are hijacked by M. tuberculosis as a protected growth niche. M. tuberculosis-infected macrophages undergo metabolic reprogramming where key mitochondrial pathways are modulated, but the mechanisms driving this metabolic shift is unknown. Our study demonstrates that M. tuberculosis downregulates Sirtuin 3 (SIRT3), an important regulator of mitochondrial metabolism, leading to SIRT3-dependent transcriptional downregulation of mitochondrial metabolic proteins, which is followed by oxidative stress and macrophage necrosis. This study identifies SIRT3 modulation as a key event in M. tuberculosis-induced metabolic reprograming in macrophages that defend against tuberculosis.

Role of histone deacetylase Sirt3 in the development and regression of atherosclerosis.

Atherosclerosis (AS) is the most common cause of death in cardiovascular diseases and poses severe challenges to human life and safety. Epigenetics plays a vital role in every single link of AS. Whereas, how epigenetics regulates its development and regression is still unknown. Sirt3, a recognized histone deacetylase, having been reported to be involved in other acylation processes in recent years, is broadening its role in epigenetic modifications. Sirt3 is an important factor in the normal physiology of blood vessels through deacetylation of mitochondrial proteins and participates in various metabolic activities. Besides, medical research targeting Sirt3 is in full swing as well. This review combining histone deacetylase Sirt3 with AS, aims to clarify the latest progress in the significant role of Sirt3 in the development and regression of AS and to provide a novel prospect for a new regulatory factor and potential intervention target for AS.

A Sex-Specific Role of Endothelial Sirtuin 3 on Blood Pressure and Diastolic Dysfunction in Female Mice.

BACKGROUND: Heart failure with preserved ejection fraction (HFpEF) is characterized by a diastolic dysfunction and is highly prevalent in aged women. Our study showed that ablation of endothelial Sirtuin 3 (SIRT3) led to diastolic dysfunction in male mice. However, the sex-specific role of endothelial SIRT3 deficiency on blood pressure and diastolic function in female mice remains to be investigated. METHODS AND RESULTS: In this study, we demonstrate that the ablation of endothelial SIRT3 in females elevated blood pressure as compared with control female mice. Diastolic function measurement also showed that the isovolumic relaxation time (IVRT) and myocardial performance index (MPI) were significantly increased, whereas the E' velocity/A' velocity (E'/A') ratio was reduced in the endothelial-specific SIRT3 knockout (SIRT3 ECKO) female mice. To further investigate the regulatory role of endothelial SIRT3 on blood pressure and diastolic dysfunction in metabolic stress, SIRT3 ECKO female mice were fed a normal diet and high-fat diet (HFD) for 20 weeks. The knockout of endothelial SIRT3 resulted in an increased blood pressure in female mice fed with an HFD. Intriguingly, SIRT3 ECKO female mice + HFD exhibited impaired coronary flow reserve (CFR) and more severe diastolic dysfunction as evidenced by an elevated IVRT as compared with control female mice + HFD. In addition, female SIRT3 ECKO mice had higher blood pressure and diastolic dysfunction as compared to male SIRT3 ECKO mice. Moreover, female SIRT3 ECKO mice + HFD had an impaired CFR and diastolic dysfunction as compared to male SIRT3 ECKO mice + HFD. CONCLUSIONS: These results implicate a sex-specific role of endothelial SIRT3 in regulating blood pressure and diastolic function in mice. Deficiency of endothelial SIRT3 may be responsible for a diastolic dysfunction in aged female.

Histone Acetyltransferase p300 Inhibitor Improves Coronary Flow Reserve in SIRT3 (Sirtuin 3) Knockout Mice.

Background Coronary microvascular dysfunction is common in patients of myocardial infarction with non-obstructive coronary artery disease. Coronary flow reserve (CFR) reflects coronary microvascular function and is a powerful independent index of coronary microvascular dysfunction and heart failure. Our previous studies showed that knockout of SIRT3 (Sirtuin 3) decreased CFR and caused a diastolic dysfunction. Few studies focus on the treatment of impaired CFR and heart failure. In the present study, we explored the role of C646, a histone acetyltransferase p300 inhibitor, in regulating CFR and cardiac remodeling in SIRT3 knockout (SIRT3KO) mice. Methods and Results After treating with C646 for 14 days, CFR, pulse-wave velocity, and cardiac function were measured in SIRT3KO mice. SIRT3KO mice treated with C646 showed a significant improvement of CFR, pulse-wave velocity, ejection fraction, and fractional shortening. Treatment with C646 reversed pre-existing cardiac fibrosis, hypertrophy, and capillary rarefaction in SIRT3KO mice. Mechanistically, knockout of Sirtuin 3 resulted in significant increases in p300 expression and H3K56 acetylation. Treatment with C646 significantly reduced levels of p300 and H3K56 acetylation in SIRT3KO mice. Furthermore, treatment with C646 increased endothelial nitric oxide synthase expression and reduced arginase II expression and activity. The expression of NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) and VCAM-1 (vascular cell adhesion molecule 1) was also significantly suppressed by C646 treatment in SIRT3KO mice. Conclusions C646 treatment attenuated p300 and H3K56 acetylation and improved arterial stiffness and CFR via improvement of endothelial cell (EC) dysfunction and suppression of NF-κB.

Combination of sirtuin 3 and hyperoxia diminishes tumorigenic properties of MDA-MB-231 cells.

AIMS: Since the role of the major mitochondrial NAD+-dependent deacetylase, sirtuin 3 (Sirt3), is differential in cancer, opposite to the well-known tumor-suppressing effect of hyperoxia, this study aimed to investigate the role of Sirt3 in triple-negative breast cancer (TNBC) cell line MDA-MB-231 upon hyperoxic (95% O2) conditions. MAIN METHODS: MDA-MB-231 cells were stably transfected with Flag-tagged Sirt-3 or empty plasmid. Western blot and real-time PCR were used to monitor the expression of proteins or genes involved in mitochondrial biogenesis, metabolic regulation and antioxidant defense. Immunocytochemistry and confocal microscopy were used to confirm the cellular localization and abundance of proteins. Flow cytometry was used to analyze mitochondrial mass, potential and ROS production, and MTT test as a measure of metabolic activity. Mitotic index analysis, colony-forming unit assay, DNA damage and Annexin V-FITC analyses were used to assess the differences in the growth and apoptosis rate. KEY FINDINGS: Although Sirt3 seemed to improve mitochondrial properties by increasing mitochondrial mass and potential, metabolic activity (Warburg effect) and antioxidative defense (SOD2, Cat), it also increased mitochondrial ROS, induced DNA damage, timp-1 expression, formation of multinucleated cells and apoptosis, and finally markedly reduced the proliferation of MDA-MB-231 cells. All these effects were even more evident upon the hyperoxic treatment, thus pointing towards combined negative effect of Sirt3 and hyperoxia on MDA-MB-231 cells. SIGNIFICANCE: Both Sirt3 and hyperoxia, alone or in combination, have the potential to negatively affect the malignant properties of the MDA-MB-231 cells and should be further explored as a possible therapy for TNBC.

Sirt3 is dispensable for oocyte quality and female fertility in lean and obese mice.

Mammalian oocytes rely heavily on mitochondrial oxidative phosphorylation (OXPHOS) for generating ATP. However, mitochondria are also the primary source of damaging reactive oxygen species (ROS). Mitochondrial de-regulation, therefore, underpins poor oocyte quality associated with conditions such as obesity and aging. The mitochondrial sirtuin, Sirt3, is critical for mitochondrial respiration and redox regulation. Interestingly, however, Sirt3 knockout (Sirt3-/- ) mice do not exhibit systemic compromise under basal conditions, only doing so under stressed conditions such as high-fat diet (HFD)-induced obesity. Mouse oocytes depleted of Sirt3 exhibit increased ROS in vitro, but it is unknown whether Sirt3 is necessary for female fertility in vivo. Here, we test this for the first time by investigating ovarian follicular reserve, oocyte maturation (including detailed spindle assembly and chromosome segregation), and female fertility in Sirt3-/- females. We find that under basal conditions, young Sirt3-/- females exhibit no defects in any parameters. Surprisingly, all parameters also remain intact following HFD-induced obesity. Despite markedly increased ROS levels in HFD Sirt3-/- oocytes, ATP levels nevertheless remain normal. Our data support that ATP is sustained in vivo through increased mitochondrial mass possibly secondary to compensatory upregulation of another sirtuin, Sirt1, which has overlapping functions with Sirt3.

SIRT3 promotes lipophagy and chaperon-mediated autophagy to protect hepatocytes against lipotoxicity.

Lipophagy is a lysosomal lipolytic pathway that complements the actions of cytosolic neutral lipases. Chaperon-mediated autophagy (CMA) triggers lipid droplets (LDs) breakdown, to initiate lipolysis via either cytosolic lipases or macroautophagy. SIRT3, a mitochondrial NAD+-dependent deacetylase, regulates the acetylation status and activity of many substrates involving in energy metabolism. However, the role of SIRT3 in regulating lipophagy is controversial. The current study showed that SIRT3 expression was decreased and the macroautophagy flux was blocked in the primary hepatocytes from high-fat diet fed mice and P/O (palmitic acid and oleic acid mixture) treated AML12 mouse hepatocytes, compared with the corresponding controls. SIRT3 overexpression promoted macroautophagy in LDs from P/O-treated hepatocytes through activating AMP-activated protein kinase (AMPK) and unc-51-like kinase 1, to boost LDs digestion. Gain of SIRT3 expression stimulated the formation of lysosome-associated membrane protein 2A (LAMP-2A)-heat shock cognate 71 kDa protein (HSC70)-perilipin-2 (PLN2) complex, to promote CMA process and reduce the stability of LDs in hepatocytes. Moreover, SIRT3 reduced the expression of stearoyl-CoA desaturase 1, to suppress lipogenesis. In addition, SIRT3 overexpression promoted LDs dispersion on detyrosinated microtubules, and directly deacetylated long-chain acyl-CoA dehydrogenase to enhance mitochondrial energetics. Taken together, SIRT3 ameliorates lipotoxicity in hepatocytes, which might be a potential target for the treatment of nonalcoholic fatty liver disease.

The yin and yang faces of the mitochondrial deacetylase sirtuin 3 in age-related disorders.

Aging, the most important risk factor for many of the chronic diseases affecting Western society, is associated with a decline in mitochondrial function and dynamics. Sirtuin 3 (SIRT3) is a mitochondrial deacetylase that has emerged as a key regulator of fundamental processes which are frequently dysregulated in aging and related disorders. This review highlights recent advances and controversies regarding the yin and yang functions of SIRT3 in metabolic, cardiovascular and neurodegenerative diseases, as well as the use of SIRT3 modulators as a therapeutic strategy against those disorders. Although most studies point to a protective role upon SIRT3 activation, there are conflicting findings that need a better elucidation. The discovery of novel SIRT3 modulators with higher selectivity together with the assessment of the relative importance of different SIRT3 enzymatic activities and the relevance of crosstalk between distinct sirtuin isoforms will be pivotal to validate SIRT3 as a useful drug target for the prevention and treatment of age-related diseases.

SIRT3 retards intervertebral disc degeneration by anti-oxidative stress by activating the SIRT3/FOXO3/SOD2 signaling pathway.

OBJECTIVE: The objective of this paper is to determine whether SIRT3 could retard intervertebral disc degeneration and study the mechanism. MATERIALS AND METHODS: We chose the 3-month mice to establish intervertebral disc degeneration model and study the effect of SIRT3 on the intervertebral disc by Western blotting, quantitative Real Time-Polymerase Chain Reaction (qRT-PCR), immunohistochemistry. Mouse nucleus pulposus cells were cultured to study the exact mechanism. RESULTS: The expression of SIRT3 was decreased in degenerated human nucleus pulposus. Intervertebral discs of mice treated with theacrine expressed more collagen II and less collagen X. In addition, nucleus pulposus cells stimulated with interleukin-1ß (IL-1ß) expressed less SIRT3 than that in the control group and nucleus pulposus cells with SIRT3 overexpress vectors expressed more collagen II FOXO3a and superoxide dismutase 2 (SOD2), indicating that SIRT3 could improve the intervertebral disc degeneration by anti-oxidative stress. CONCLUSIONS: SIRT3 is a protective factor for intervertebral discs and can reduce oxidative stress in the intervertebral disc.

Melatonin ameliorates cerebral ischemia/reperfusion injury through SIRT3 activation.

AIMS: Previous literature has shown that melatonin plays a critical role in protecting against cerebral ischemia/reperfusion (I/R) injury. Sirtuin3(SIRT3), as one member of the sirtuin family, protects against oxidative stress-related diseases. However, the association between melatonin and SIRT3 in cerebral I/R injury is not well understood. Our experiment was planned to investigate whether melatonin protects against cerebral I/R injury through SIRT3 activation. MAIN METHODS: We selected transient middle cerebral artery occlusion (tMCAO) mice as the model of cerebral I/R injury. Male C57/BL6 mice were pre-treated with or without a selective SIRT3 inhibitor and then subjected to tMCAO surgery. Melatonin (20 mg/kg) was given to mice by intraperitoneal injection after ischemia and before reperfusion. Then, we observed the changes in the SIRT3 and downstream relative proteins, infarction volume, neurological score, Nissl, H&E and TUNEL staining, and the expression of apoptosis proteins after tMCAO. KEY FINDINGS: Melatonin upregulated the expression of SIRT3 after tMCAO, and alleviated the neurological dysfunction and cell apoptosis through SIRT3 activation. SIGNIFICANCE: Our research proved that melatonin promoted SIRT3 expression after tMCAO and alleviated cerebral I/R injury by activating the SIRT3 signaling pathway. This study provides novel therapeutic targets and mechanisms for the treatment of ischemic stroke in the clinic, especially during cerebrovascular reperfusion.

Update on the role of Sirtuin 3 in cell differentiation: A major metabolic target that can be pharmacologically controlled.

Cell differentiation is a fundamental biological event in which a precursor stem cell is turning into a specialized somatic cell. It is thus crucial for the development, tissue turnover and regeneration in mammals. Among the numerous changes taking place in a cell during a differentiation programme, the biology of mitochondria, the central organelle mainly responsible for energy homeostasis and stress adaptation, is deeply modified. These modifications are now well recognized as taking an active part to the completion of the differentiation programme. Indeed, mitochondrial biogenesis and metabolic shift are observed during cell differentiation, adapting many syntheses, calcium homeostasis, ATP and reactive oxygen species production, to the needs. These mitochondrial functions are substantially regulated by the post-translational modifications of the mitochondrial proteins among which lysine acetylation is essential. This mitoacetylome is then globally controlled by the balance between spontaneous/enzymatically-catalysed protein acetylation and the NAD+-dependent deacetylation mediated by Sirtuin 3. This enzyme is now considered as a major regulator of the function of the organelle. Regarding the requirement of these mitochondrial adaptations, the subsequent growing interest for this enzyme recently extended to the investigation of the mechanisms driving cell differentiation. This review summarizes the currently available information about the significance of SIRT3 in cell differentiation in physio-pathological contexts. We also suggest a control of the differentiation-activated autophagy by SIRT3, a hypothesis supported by recent findings establishing a causal link between SIRT3 and autophagy. Eventually, an update on the present pharmacological modulators of SIRT3 in a context of cell differentiation is discussed.

Inhibition of Mitochondrial Oxidative Damage Improves Reendothelialization Capacity of Endothelial Progenitor Cells via SIRT3 (Sirtuin 3)-Enhanced SOD2 (Superoxide Dismutase 2) Deacetylation in Hypertension.

OBJECTIVE: Dysfunction of endothelial progenitor cells (EPCs) leads to impaired endothelial repair capacity in patients with hypertension, but the mechanisms remain incompletely understood. Mitochondrial oxidative stress is involved in endothelial injury in hypertension. In this study, we aim to investigate the role of mitochondrial oxidative stress in the deficient endothelial reparative capacity of EPCs and identify enhanced SIRT3 (sirtuin 3)-mediated SOD2 (superoxide dismutase 2) deacetylation as a novel endothelial protective mechanism in hypertension. Approach and Results: Hypertension-EPCs displayed increased mitochondrial reactive oxygen species and mitochondrial damage, including loss of mitochondrial membrane potential, abnormal mitochondrial ultrastructure, and mtDNA oxidative injury, which was coincided with impaired in vitro function and in vivo reendothelialization capacity. The harmful effects of hypertension on mitochondrial function of EPCs were in vitro mimicked by angiotensin II coincubation. Scavenging of mitochondrial reactive oxygen species with mitoTEMPO attenuated mitochondrial oxidative damage and rescued reendothelialization capacity. Enzymatic activity and deacetylation level of SOD2 were significantly reduced in hypertension-EPCs, which was accompanied with decreased SIRT3 expression. Knockdown of SIRT3 in EPCs resulted in mitochondrial oxidative damage, hyperacetylation of SOD2, and suppression of reendothelialization capacity. SIRT3 physically interacted with SOD2 and eliminated excess mitochondrial reactive oxygen species, restored mitochondrial function through enhancing SOD2 activity by deacetylation of K68. Upregulation of SIRT3/SOD2 signaling improved reendothelialization capability of EPCs. CONCLUSIONS: The present study demonstrated for the first time that mitochondrial oxidative damage because of deficient SIRT3/SOD2 signaling contributes to the decline in reendothelialization capacity of EPCs in hypertension. Maintenance of mitochondrial redox homeostasis in EPCs may be a novel therapeutic target for endothelial injury.

SIRT3 deacetylase activity confers chemoresistance in AML via regulation of mitochondrial oxidative phosphorylation.

Acute myeloid leukaemia (AML) cells possess metabolism profiles, such as higher rates of oxidative phosphorylation and dependence on fatty acid oxidation for survival, and are dependent on the sophisticated regulation of reactive oxygen species (ROS) generation for survival, drug resistance and stemness maintenance. We found that sensitivity of primary AML cells to cytarabine correlated with SOD2 acetylation and the ability of the drug to induce mitochondrial ROS. The SOD2 deacetylase, SIRT3, protected AML cells from chemotherapy as shown by inhibited apoptosis via inhibited drug-induced production of mitochondrial ROS. SIRT3 significantly decreased nicotinamide adenine dinucleotide phosphate (NADP)/reduced NADP ratio and increased reduced glutathione/oxidized glutathione ratio. Furthermore, SIRT3 enhanced oxidative phosphorylation (OxPhos) in AML cells under both basic and cytarabine-treated conditions. A xenograft mouse model showed that SIRT3 overexpressing AML cells and patient-derived xenograft mice bearing high SIRT3 deacetylase activity were more resistant to chemotherapy in vivo. SIRT3 inhibitor displayed synergy with cytarabine to ablate AML cells in vitro and in mouse models. Taken together, our study showed that SIRT3 is capable of reprograming mitochondrial metabolism towards OxPhos and downregulating ROS generation, which contribute to the chemoresistance of AML cells. SIRT3 can be utilized as a potential therapeutic target to improve the anti-leukaemic efficacy of standard chemotherapeutic agents for AML.

Short-term high salt intake impairs hepatic mitochondrial bioenergetics and biosynthesis in SIRT3 knockout mice.

High salt intake (HS) is an important factor in the development of many metabolic diseases. The liver is the metabolic center in the body. However, the effect of short-term HS on the liver mitochondria and its mechanism are still unclear. In this study, we investigated the effects of short-term HS on liver mitochondrial function. We found that HS reduced Sirtuin3 (SIRT3) protein level, increasing protein carbonylation in mice liver. HS intake decreased ATP production, mitochondrial transcription factor A (TFAM), and complex I level. SIRT3 knockout (SKO) mice exhibited similar results with HS-treated wild-type mice but with a less extent of carbonylation and ATP reduction. Our study shows that short-term HS led to increased hepatic oxidative state, impaired mitochondrial biosynthesis, and bioenergetics. HS-treated mice could still maintain hepatic glucose homeostasis by compensatory activation of Adenosine 5'-monophosphate-activated protein kinase (AMPK). However, in HS-treated SKO mice, AMPK was not activated, instead, the glycogen synthase activity increased, which caused an exceptionally increased glycogen accumulation. This study provides evidence that short-term HS intake could cause the early hepatic metabolic changes, highlighting the importance of controlling salt intake especially in those patients with defects in SIRT3. Highlights High salt intake down-regulates SIRT3 protein level and increases oxidation. High salt intake activates AMPK via AMP-dependent pathway. High salt intake impairs energy metabolism. High salt combined with SIRT3 knockout results in glycogen accumulation.

Berberine alleviates nonalcoholic fatty liver induced by a high-fat diet in mice by activating SIRT3.

Berberine (BBR) shows promising effects in the treatment of nonalcoholic fatty liver disease (NAFLD) by influencing various metabolic aspects. Inhibition of mitochondrial ß-oxidation (ß-OX) participates in the pathogenesis of NAFLD. Silent mating-type information regulation 2 homolog 3 (SIRT3) has been reported to regulate mitochondrial ß-OX by deacetylating its substrate, long-chain acyl-coenzyme A dehydrogenase (LCAD). This study aimed to explore whether BBR can promote mitochondrial ß-OX and the role of SIRT3 as well as the mechanisms underlying the effects of BBR on hepatic lipid metabolism in mice fed a high-fat diet (HFD). BBR can significantly improve systematic and hepatic lipid metabolism in HFD-fed mice. Metabolomics analysis revealed that ß-OX was inhibited in HFD-induced mice, as indicated by the reduced production of short and medium carbon chain acyl-carnitines, the activated form of free fatty acids, via ß-OX, which was reversed by BBR intervention. Exploration of the mechanism found that BBR intervention reversed the down-regulation of SIRT3 and decreased the LCAD hyperacetylation level in HFD-fed mice. SIRT3 knockout (KO) mice were used to identify the role of SIRT3 in the BBR's influence of ß-OX. The beneficial effects of BBR on systemic and hepatic metabolism were profoundly attenuated in KO mice. Moreover, the promotive effect of BBR on ß-OX in HFD-induced mice was partially abolished in KO mice. These results suggested that BBR alleviates HFD-induced inhibition of fatty acid ß-OX partly through SIRT3-mediated LCAD deacetylation, which may provide a novel mechanism and support BBR as a promising therapeutic for NAFLD.-Xu, X., Zhu, X.-P., Bai, J.-Y., Xia, P., Li, Y., Lu, Y., Li, X.-Y., Gao, X. Berberine alleviates nonalcoholic fatty liver induced by a high-fat diet in mice by activating SIRT3.