Role of liraglutide in brain repair promotion through Sirt1-mediated mitochondrial improvement in stroke

Brain repair, especially axonal sprouting, is critical to restore motor function in disabled stroke patients. Liraglutide (LG) is a new kind of long-acting analogue of glucagon-like peptide-1 (GLP-1) and has potential protective effects in stroke. The mitochondria participate in brain repair after cerebral injury. However, the mechanism of the effect of LG on brain repair and its potential influence on mitochondria in stroke remains obscure. Here, in focal cerebral cortical ischemic mice model, LG improved the motor functional recovery and promoted axonal sprouting by restoring the activities of isocitrate dehydrogenase, α-ketoglutarate dehydrogenase, and succinate dehydrogenase. Moreover, LG remarkably increased the cell survival rate and revived the NeuN and GAP-43 levels in cortical neurons under hydrogen peroxide (H₂O₂) exposure. It was also observed that LG reduced the generation of reactive oxygen species, stabilized the mitochondrial membrane potential, enhanced the levels of adenosine triphosphate, enhanced activities of mitochondrial complex-I, and decreased protein expression levels of fission-1 in H₂O₂-injured cortical neurons. Additionally, LG suppressed the expressions of sirtuin 1 (Sirt1) in cortical neurons exposed to H₂O₂. Furthermore, knockdown of Sirt1 by short interfering RNA facilitated the LG-mediated mitochondrial protection in cortical neurons under H₂O₂. Collectively, this data from the present study illustrated that LG exerted a promoting influence on brain repair, after cerebral ischemic injury, through Sirt1-mediated mitochondrial improvement.     

Sirt1 and the Mitochondria

Sirt1 is the most prominent and extensively studied member of sirtuins, the family of mammalian class III histone deacetylases heavily implicated in health span and longevity. Although primarily a nuclear protein, Sirt1’s deacetylation of Peroxisome proliferator-activated receptor Gamma Coactivator-1α (PGC-1α) has been extensively implicated in metabolic control and mitochondrial biogenesis, which was proposed to partially underlie Sirt1’s role in caloric restriction and impacts on longevity. The notion of Sirt1’s regulation of PGC-1α activity and its role in mitochondrial biogenesis has, however, been controversial. Interestingly, Sirt1 also appears to be important for the turnover of defective mitochondria by mitophagy. I discuss here evidences for Sirt1’s regulation of mitochondrial biogenesis and turnover, in relation to PGC-1α deacetylation and various aspects of cellular physiology and disease.     

Interaction of Sirt3 with OGG1 contributes to repair of mitochondrial DNA and protects from apoptotic cell death under oxidative stress

Sirtuin 3 (Sirt3), a major mitochondrial NAD⁺-dependent deacetylase, targets various mitochondrial proteins for lysine deacetylation and regulates important cellular functions such as energy metabolism, aging, and stress response. In this study, we identified the human 8-oxoguanine-DNA glycosylase 1 (OGG1), a DNA repair enzyme that excises 7,8-dihydro-8-oxoguanine (8-oxoG) from damaged genome, as a new target protein for Sirt3. We found that Sirt3 physically associated with OGG1 and deacetylated this DNA glycosylase and that deacetylation by Sirt3 prevented the degradation of the OGG1 protein and controlled its incision activity. We further showed that regulation of the acetylation and turnover of OGG1 by Sirt3 played a critical role in repairing mitochondrial DNA (mtDNA) damage, protecting mitochondrial integrity, and preventing apoptotic cell death under oxidative stress. We observed that following ionizing radiation, human tumor cells with silencing of Sirt3 expression exhibited deteriorated oxidative damage of mtDNA, as measured by the accumulation of 8-oxoG and 4977 common deletion, and showed more severe mitochondrial dysfunction and underwent greater apoptosis in comparison with the cells without silencing of Sirt3 expression. The results reported here not only reveal a new function and mechanism for Sirt3 in defending the mitochondrial genome against oxidative damage and protecting from the genotoxic stress-induced apoptotic cell death but also provide evidence supporting a new mtDNA repair pathway.     

Sirt7 inhibits Sirt1-mediated activation of Suv39h1

Sirtuins regulate a variety of cellular processes through protein deacetylation. The best-known member of mammalian sirtuin family, Sirt1, plays important roles in the maintenance of cellular homeostasis by regulating cell metabolism, differentiation and stress responses, among others. Sirt1 activity requires tight regulation to meet specific cellular requirements, which is achieved at different levels and by specific mechanisms. Recently, a regulatory loop between Sirt1 and another sirtuin, Sirt7, was identified. Sirt7 inhibits Sirt1 autodeacetylation at K230 and activation thereby preventing Sirt1-mediated repression of adipocyte differentiation by inhibition of the PPARγ gene. Here, we extend the regulatory complexity of Sirt7-dependent restriction of Sirt1 activity by demonstrating that Sirt7 reduces activation of a previously described prominent Sirt1 target, the histone methyltransferase Suv39h1. We show that removal of the acetyl-group at K230 in Sirt1 due to the absence of Sirt7 leads to hyperactivation of Sirt1 and thereby to constantly increased activity of Suv39h1.     

Sirt1–hypoxia‐inducible factor‐1α interaction is a key mediator of tubulointerstitial damage in the aged kidney

Although it is known that the expression and activity of sirtuin 1 (Sirt1) decrease in the aged kidney, the role of interaction between Sirt1 and hypoxia‐inducible factor (HIF)‐1α is largely unknown. In this study, we investigated whether HIF‐1α could be a deacetylation target of Sirt1 and the effect of their interaction on age‐associated renal injury. Five‐week‐old (young) and 24‐month‐old (old) C57Bl/6J mice were assessed for their age‐associated changes. Kidneys from aged mice showed increased infiltration of CD68‐positive macrophages, higher expression of extracellular matrix (ECM) proteins, and more apoptosis than young controls. They also showed decreased Sirt1 expression along with increased acetylated HIF‐1α. The level of Bcl‐2/adenovirus E1B‐interacting protein 3, carbonic anhydrase 9, Snail, and transforming growth factor‐β1, which are regulated by HIF‐1α, was significantly higher in aged mice suggesting that HIF‐1α activity was increased. In HK‐2 cells, Sirt1 inhibitor sirtinol and siRNA‐mediated knockdown of Sirt1 enhanced apoptosis and ECM accumulation. During hypoxia, Sirt1 was down‐regulated, which allowed the acetylation and activation of HIF‐1α. Resveratrol, a Sirt1 activator, effectively prevented hypoxia‐induced production of ECM proteins, mitochondrial damage, reactive oxygen species generation, and apoptosis. The inhibition of HIF‐1α activity by Sirt1‐induced deacetylation of HIF‐1α was confirmed by Sirt1 overexpression under hypoxic conditions and by resveratrol treatment or Sirt1 overexpression in HIF‐1α‐transfected HK‐2 cells. Finally, we confirmed that chronic activation of HIF‐1α promoted apoptosis and fibrosis, using tubular cell‐specific HIF‐1α transgenic mice. Taken together, our data suggest that Sirt1‐induced deacetylation of HIF‐1α may have protective effects against tubulointerstitial damage in aged kidney.     

Resveratrol prevents doxorubicin cardiotoxicity through mitochondrial stabilization and the Sirt1 pathway

Doxorubicin (DOX) is one of the most effective chemotherapeutic drugs; however, its incidence of cardiotoxicity compromises its therapeutic index. DOX-induced heart failure is thought to be caused by reduction/oxidation cycling of DOX to generate oxidative stress and cardiomyocyte cell death. Resveratrol (RV), a stilbene found in red wine, has been reported to play a cardioprotective role in diseases associated with oxidative stress. The objective of this study was to test the ability of RV to protect against DOX-induced cardiomyocyte death. We hypothesized that RV protects cardiomyocytes from DOX-induced oxidative stress and subsequent cell death through changes in mitochondrial function. DOX induced a rapid increase in reactive oxygen species (ROS) production in cardiac cell mitochondria, which was inhibited by pretreatment with RV, most likely owing to an increase in MnSOD activity. This effect of RV caused additional polarization of the mitochondria in the absence and presence of DOX to increase mitochondrial function. RV pretreatment also prevented DOX-induced cardiomyocyte death. The protective ability of RV against DOX was abolished when Sirt1 was inhibited by nicotinamide. Our data suggest that RV protects against DOX-induced oxidative stress through changes in mitochondrial function, specifically the Sirt1 pathway leading to cardiac cell survival.     

LKB1及其生物学功能

LKB1基因是一种保守的抑癌基因,其编码产物LKB1即丝氨酸-苏氨酸激酶11(serine/threonine kinase,STK11)。LKB1与细胞极性调节、男性精子形成、肿瘤及细胞代谢等方面有关。本文阐述了近年来LKB1的最新研究进展。     

LKB1 loss of function studied in vivo

Recent developments have placed the serine/threonine kinase LKB1 on the crossroads linking energy metabolism, cell structure and cancer progression and that its deletion can affect tumorigenesis, metastasis, cell adhesion and polarity. LKB1 can regulate a host of different functions which all have potential to impact upon the initiation and progression of neoplastic disease. To understand the phenotypic consequences of LKB1 loss in a range of different settings, a number of animal models of loss of function have been generated and analyzed. In this review we summarize recent data generated from a range of these models, which reveal clear tissue specific differences in LKB1 function in vivo and in the consequences of its loss.     

LKB1抑癌机制研究进展

人LKB1(Liver Kinase B1,或Serine-Threonine Kinase 11,STK11)基因的胚系失活突变可导致癌症易感病皮杰氏综合征(Peutz-Jeghers syndrome,PJS),该病患者多发错构瘤息肉且患癌症风险增加。LKB1基因的体细胞突变还广泛地存在于众多类型的恶性肿瘤中,如肺癌、结肠癌和乳腺癌等,因此,LKB1被普遍认为是抑癌基因。LKB1基因的编码产物LKB1是一种丝氨酸/苏氨酸激酶,调节多种细胞生理病理过程。虽然LKB1的抑癌机制尚不完全清楚,但现有的研究表明,对细胞生长增殖、能量代谢和细胞极性等的调控是其抑制肿瘤发生和发展的重要方面。本文就目前已知的LKB1的抑癌机制作一综述。     

Molecular mechanisms of tumor suppression by LKB1

The LKB1 tumor suppressor gene is frequently mutated in sporadic lung adenocarcinomas and cervical cancers and germline mutations are causative for Peutz-Jeghers syndrome characterized by gastrointestinal polyposis. The intracellular LKB1 kinase is implicated in regulating polarity, metabolism, cell differentiation, and proliferation – all functions potentially contributing to tumor suppression. LKB1 acts as an activating kinase of at least 14 kinases mediating LKB1 functions in a complex signaling network with partial overlaps. Regulation of the LKB1 signaling network is highly context dependent, and spatially organized in various cellular compartments. Also the mechanisms by which LKB1 activity suppresses tumorigenesis is context dependent, where recent observations are providing hints on the molecular mechanisms involved.     

Downregulation of Sirt1 as aging change in advanced heart failure

Background

In congestive heart failure the balance between cell death and cell survival in cardiomyocytes is compromised. Sirtuin 1 (Sirt1) activates cell survival machinery and has been shown to be protective against ischemia/reperfusion injury in murine heart. The role of Sirt1 in heart failure, especially in human hearts is not clear.

Results

The expression of Sirt1 and other (associated) downstream molecules in human cardiomyocytes from patients with advanced heart failure was examined. Sirt1 was down-regulated (54.92% ± 7.80% in advanced heart failure samples compared with healthy control cardiomyocytes). The modulation of molecules involved in cardiomyocyte survival and death in advanced heart failure were also examined. The expression of Mn-superoxide dismutase and thioredoxin1, as well as an antiapoptotic molecule, Bcl-xL, were all significantly reduced in advanced heart failure cardiomyoctes (0.71 ± 0.02-fold, 0.61 ± 0.05-fold, and 0.53 ± 0.08-fold vs. control, respectively); whereas the expression of proapoptotic molecule Bax was significantly increased (1.62 ± 0.18-fold vs. control). Increased TUNEL-positive number of cardiomyocytes and oxidative stress, confirmed by 8-hydorxydeoxyguanosine staining, were associated with advanced heart failure. The AMPK-Nampt-Sirt1 axis also showed inhibition in advanced heart failure in addition to severely impaired AMPK activation. Increased p53 (acetyl form) and decreased FoxO1 translocation in the nucleus may be the mechanism of down-regulation of antioxidants and up-regulation of proapoptotic molecules due to low expression of Sirt1.

Conclusion

In advanced heart failure, low Sirt1 expression, like aging change may be a significant contributing factor in the downregulation of antioxidants and upregulation of proapoptotic molecules through the p53, FoxO1, and oxidative stress pathways.     

Disruption of Igfbp1 fails to rescue the phenotype of Sirt1−/− mice

Sirtuin 1 (SIRT1) is an NAD-dependent histone deacetylase (HDAC) whose activity is thought to forestall the onset of a variety of age-related diseases. Mice carrying null mutations of the Sirt1 gene suffer high rates of neonatal lethality and those that survive are sterile, growth retarded, lean and their livers express high levels of insulin-like growth factor binding protein-1 (IGFBP1). IGFBP1 binds and regulates the bioavailability of Igfs. Interestingly, Igfbp1 transgenic mice largely phenocopy Sirt1−/− mice, suggesting the possibility that the over-expression of IGFBP1 in Sirt1−/− mice might be responsible for many of their phenotypes. We interbred Sirt1 heterozygote mice to Igfbp1-deficient mice to test the hypothesis that the disruption of one or both alleles of Igfbp1 would rescue the phenotype of Sirt1−/− mice. We report that mono- or bi-allelic disruption of the Igfbp1 gene had no effect on the embryonic and neonatal lethality of Sirt1−/− mice. However, we show that mice lacking at least one allele of both Sirt1 and Igfbp1 genes have a much higher incidence of malocclusion.     

Sirt3 modulate renal ischemia-reperfusion injury through enhancing mitochondrial fusion and activating the ERK-OPA1 signaling pathway

Mitochondrial fusion is linked to heart and liver ischemia-reperfusion (IR) insult. Unfortunately, there is no report to elucidate the detailed influence of mitochondrial fusion in renal IR injury. This study principally investigated the mechanism by which mitochondrial fusion protected kidney against IR injury. Our results indicated that sirtuin 3 (Sirt3) was inhibited after renal IR injury in vivo and in vitro. Overexpression of Sirt3 improved kidney function, modulated oxidative injury, repressed inflammatory damage, and reduced tubular epithelial cell apoptosis. The molecular investigation found that Sirt3 overexpression attenuated IR-induced mitochondrial damage in renal tubular epithelial cells, as evidenced by decreased reactive oxygen species production, increased antioxidants sustained mitochondrial membrane potential, and inactivated mitochondria-initiated death signaling. In addition, our information also illuminated that Sirt3 maintained mitochondrial homeostasis against IR injury by enhancing optic atrophy 1 (OPA1)-triggered fusion of mitochondrion. Inhibition of OPA1-induced fusion repressed Sirt3 overexpression-induced kidney protection, leading to mitochondrial dysfunction. Further, our study illustrated that OPA1-induced fusion could be affected through ERK; inhibition of ERK abolished the regulatory impacts of Sirt3 on OPA1 expression and mitochondrial fusion, leading to mitochondrial damage and tubular epithelial cell apoptosis. Altogether, our results suggest that renal IR injury is closely associated with Sirt3 downregulation and mitochondrial fusion inhibition. Regaining Sirt3 and/or activating mitochondrial fission by modifying the ERK-OPA1 cascade may represent new therapeutic modalities for renal IR injury.     

The LKB1/AMPK polarity pathway

The LKB1 tumor suppressor kinase is an activator of the AMP-activated protein kinase (AMPK), a metabolic gauge that responds to variations of cellular energetic levels by favoring catabolic versus anabolic processes. Recent studies have provided substantial evidence that LKB1 and AMPK control cell polarity from invertebrates to mammals. This review examines how the LKB1–AMPK pathway, in conjunction with other positional signals, converts energy-sensing information into the activation of Myosin II to maintain epithelial-cell architecture but also to complete cell division. This molecular link between polarity and metabolism may constitute an ancient stress-response protective mechanism that was co-opted for tumor suppression during evolution.     

LKB1IP promotes pathological cardiac hypertrophy by targeting PTEN/Akt signalling pathway

Pathological cardiac hypertrophy represents a leading cause of morbidity and mortality worldwide. Liver kinase B1 interacting protein 1 (LKB1IP) was identified as the binding protein of tumour suppressor LKB1. However, the role of LKB1IP in the development of pathological cardiac hypertrophy has not been explored. The aim of this study was to investigate the function of LKB1IP in cardiac hypertrophy in response to hypertrophic stimuli. We investigated the cardiac level of LKB1IP in samples from patients with heart failure and mice with cardiac hypertrophy induced by isoproterenol (ISO) or transverse aortic constriction (TAC). LKB1IP knockout mice were generated and challenged with ISO injection or TAC surgery. Cardiac function, hypertrophy and fibrosis were then examined. LKB1IP expression was significantly up-regulated on hypertrophic stimuli in both human and mouse cardiac samples. LKB1IP knockout markedly protected mouse hearts against ISO- or TAC-induced cardiac hypertrophy and fibrosis. LKB1IP overexpression aggravated ISO-induced cardiomyocyte hypertrophy, and its inhibition attenuated hypertrophy in vitro. Mechanistically, LKB1IP activated Akt signalling by directly targeting PTEN and then inhibiting its phosphatase activity. In conclusion, LKB1IP may be a potential target for pathological cardiac hypertrophy.     

Src-Tyrosine kinases are major agents in mitochondrial tyrosine phosphorylation

Mitochondrial tyrosine phosphorylation is emerging as an important mechanism in regulating mitochondrial function. This article, aimed at identifying which kinases are the major agents in mitochondrial tyrosine phosphorylation, shows that this role should be attributed to Src family members. Indeed, various members of this family, for example, Fgr, Fyn, Lyn, c-Src, are constitutively present in the internal structure of mitochondria as well as Csk, a key enzyme in the regulation of the activity of this family. By means of different approaches, biochemical fractioning, Western blotting and immunogold analysis "in situ" of phosphotyrosine signaling, evidence is reported on the existence of a signal transduction pathway from plasma membrane to mitochondria, resulting in increasing Src-dependent mitochondrial tyrosine phosphorylation. The activation of Src kinases at mitochondrial level is associated with the proliferative status where several mitochondrial proteins are specifically tyrosine-phosphorylated.     

The deacetylation of Foxk2 by Sirt1 reduces chemosensitivity to cisplatin

In multiple types of cancer, decreased tumour cell apoptosis during chemotherapy is indicative of decreased chemosensitivity. Forkhead box K2 (FOXK2), which is essential for cell fate, regulates cancer cell apoptosis through several post‐translational modifications. However, FOXK2 acetylation has not been extensively studied. Here, we evaluated the effects of sirtiun 1 (SIRT1) on FOXK2 deacetylation. Our findings demonstrated that SIRT1 inhibition increased FOXK2‐induced chemosensitivity to cisplatin and that K223 in FOXK2 was acetylated. Furthermore, FOXK2 K223 deacetylation reduced chemosensitivity to cisplatin in vitro and in vivo. Mechanistically, FOXK2 was acetylated by the acetyltransferase cAMP response element binding protein and deacetylated by SIRT1. Furthermore, cisplatin attenuated the interaction between FOXK2 and SIRT1. Cisplatin or SIRT1 inhibition enhanced FOXK2 acetylation, thereby reducing the nuclear distribution of FOXK2. Additionally, FOXK2 K223 acetylation significantly affected the expression of cell cycle–related and apoptosis‐related genes in cisplatin‐stimulated cancer cells, and FOXK2 K223 hyperacetylation promoted mitotic catastrophe, which enhanced chemosensitivity to cisplatin. Overall, our results provided insights into the mechanisms of SIRT1‐mediated FOXK2 deacetylation, which was involved in chemosensitivity to cisplatin.     

Rapamycin enhances growth inhibition on urothelial carcinoma cells through LKB1 deficiency-mediated mitochondrial dysregulation

Rapamycin, a mammalian target of rapamycin (mTOR) inhibitor, has significant potential for application in the treatment of urothelial carcinoma (URCa) of the bladder. Previous studies have shown that regulation of the AMP-activated serine/threonine protein kinase (AMPK)–mTOR signaling pathway enhances apoptosis by inducing autophagy or mitophagy in bladder cancer. Alteration of liver kinase B1 (LKB1)-AMPK signaling leads to mitochondrial dysfunction and the accumulation of autophagy-related proteins as a result of mitophagy, resulting in enhanced cell sensitivity to drug treatments. Therefore, we hypothesized that LKB1 deficiency in URCa cells could lead to increased sensitivity to rapamycin by inducing mitochondrial defect-mediated mitophagy. To test this, we established stable LKBI-knockdown URCa cells and analyzed the effects of rapamycin on their growth. Rapamycin enhanced growth inhibition and apoptosis in stable LKB1-knockdown URCa cells and in a xenograft mouse model. In spite of the stable downregulation of LKB1 expression, rapamycin induced AMPK activation in URCa cells, causing loss of the mitochondrial membrane potential, ATP depletion, and ROS accumulation, indicating an alteration of mitochondrial biogenesis. Our findings suggest that the absence of LKB1 can be targeted to induce dysregulated mitochondrial biogenesis by rapamycin treatment in the design of novel therapeutic strategies for bladder cancer.