Hypoxia causes pancreatic ß-cell dysfunction and impairs insulin secretion by activating the transcriptional repressor BHLHE40.

Hypoxia can occur in pancreatic ß-cells in type 2 diabetes. Although hypoxia exerts deleterious effects on ß-cell function, the associated mechanisms are largely unknown. Here, we show that the transcriptional repressor basic helix-loop-helix family member e40 (BHLHE40) is highly induced in hypoxic mouse and human ß-cells and suppresses insulin secretion. Conversely, BHLHE40 deficiency in hypoxic MIN6 cells or ß-cells of ob/ob mice reverses defects in insulin secretion. Mechanistically, BHLHE40 represses the expression of Mafa, encoding the transcription factor musculoaponeurotic fibrosarcoma oncogene family A (MAFA), by attenuating the binding of pancreas/duodenum homeobox protein 1 (PDX1) to its enhancer region. Impaired insulin secretion in hypoxic ß-cells was recovered by MAFA re-expression. Collectively, our work identifies BHLHE40 as a key hypoxia-induced transcriptional repressor in ß-cells that inhibit insulin secretion by suppressing MAFA expression.

Insulin signaling in osteoblasts integrates bone remodeling and energy metabolism.

The broad expression of the insulin receptor suggests that the spectrum of insulin function has not been fully described. A cell type expressing this receptor is the osteoblast, a bone-specific cell favoring glucose metabolism through a hormone, osteocalcin, that becomes active once uncarboxylated. We show here that insulin signaling in osteoblasts is necessary for whole-body glucose homeostasis because it increases osteocalcin activity. To achieve this function insulin signaling in osteoblasts takes advantage of the regulation of osteoclastic bone resorption exerted by osteoblasts. Indeed, since bone resorption occurs at a pH acidic enough to decarboxylate proteins, osteoclasts determine the carboxylation status and function of osteocalcin. Accordingly, increasing or decreasing insulin signaling in osteoblasts promotes or hampers glucose metabolism in a bone resorption-dependent manner in mice and humans. Hence, in a feed-forward loop, insulin signals in osteoblasts activate a hormone, osteocalcin, that promotes glucose metabolism.

Selective Inhibitor Design for Kinase Homologs Using Multiobjective Monte Carlo Tree Search.

Designing highly selective molecules for a drug target protein is a challenging task in drug discovery. This task can be regarded as a multiobjective problem that simultaneously satisfies criteria for various objectives, such as selectivity for a target protein, pharmacokinetic endpoints, and drug-like indices. Recent breakthroughs in artificial intelligence have accelerated the development of molecular structure generation methods, and various researchers have applied them to computational drug designs and successfully proposed promising drug candidates. However, designing efficient selective inhibitors with releasing activities against various homologs of a target protein remains a difficult issue. In this study, we developed a de novo structure generator based on reinforcement learning that is capable of simultaneously optimizing multiobjective problems. Our structure generator successfully proposed selective inhibitors for tyrosine kinases while optimizing 18 objectives consisting of inhibitory activities against 9 tyrosine kinases, 3 pharmacokinetics endpoints, and 6 other important properties. These results show that our structure generator and optimization strategy for selective inhibitors will contribute to the further development of practical structure generators for drug designs.

SIRT7 Deficiency Protects against Aß42-Induced Apoptosis through the Regulation of NOX4-Derived Reactive Oxygen Species Production in SH-SY5Y Cells.

Alzheimer's disease (AD) is an age-related neurodegenerative disease that is characterized by irreversible memory loss and cognitive decline. The deposition of amyloid-ß (Aß), especially aggregation-prone Aß42, is considered to be an early event preceding neurodegeneration in AD. Sirtuins (SIRT1-7 in mammals) are nicotinamide adenine dinucleotide-dependent lysine deacetylases/deacylases, and several sirtuins play important roles in AD. However, the involvement of SIRT7 in AD pathogenesis is not known. Here, we demonstrate that SIRT7 mRNA expression is increased in the cortex, entorhinal cortex, and prefrontal cortex of AD patients. We also found that Aß42 treatment rapidly increased NADPH oxidase 4 (NOX4) expression at the post-transcriptional level, and induced reactive oxygen species (ROS) production and apoptosis in neuronal SH-SY5Y cells. In contrast, SIRT7 knockdown inhibited Aß42-induced ROS production and apoptosis by suppressing the upregulation of NOX4. Collectively, these findings suggest that the inhibition of SIRT7 may play a beneficial role in AD pathogenesis through the regulation of ROS production.

Sirt7 Deficiency Attenuates Neointimal Formation Following Vascular Injury by Modulating Vascular Smooth Muscle Cell Proliferation.

BACKGROUND: Sirt7 is a recently identified sirtuin and has important roles in various pathological conditions, including cancer progression and metabolic disorders. It has previously been reported that Sirt7 is a key molecule in acute myocardial wound healing and pressure overload-induced cardiac hypertrophy. In this study, the role of Sirt7 in neointimal formation after vascular injury is investigated.Methods and Results:Systemic (Sirt7-/-) and smooth muscle cell-specific Sirt7-deficient mice were subjected to femoral artery wire injury. Primary vascular smooth muscle cells (VSMCs) were isolated from the aorta of wild type (WT) and Sirt7-/-mice and their capacity for cell proliferation and migration was compared. Sirt7 expression was increased in vascular tissue at the sites of injury. Sirt7-/-mice demonstrated significant reduction in neointimal formation compared to WT mice. In vitro, Sirt7 deficiency attenuated the proliferation of serum-induced VSMCs. Serum stimulation-induced upregulation of cyclins and cyclin-dependent-kinase 2 (CDK2) was significantly attenuated in VSMCs of Sirt7-/-compared with WT mice. These changes were accompanied by enhanced expression of the microRNA 290-295 cluster, the translational negative regulator of CDK2, in VSMCs of Sirt7-/-mice. It was confirmed that smooth muscle cell-specific Sirt7-deficient mice showed significant reduction in neointima compared with control mice. CONCLUSIONS: Sirt7 deficiency attenuates neointimal formation after vascular injury. Given the predominant role in vascular neointimal formation, Sirt7 is a potentially suitable target for treatment of vascular diseases.

Brown adipocyte-derived exosomal miR-132-3p suppress hepatic Srebf1 expression and thereby attenuate expression of lipogenic genes.

Recent evidence has revealed a novel signaling mechanism through which brown adipose tissue (BAT)-derived exosomal microRNAs (miRNAs) influence hepatic gene expression. Here, we uncover neuronal control of these miRNAs and identify exosomal miR-132-3p as a regulator of hepatic lipogenesis under cold stress conditions. Norepinephrine, a sympathetic nervous system neurotransmitter mediating cold-induced BAT activation, altered the composition of brown adipocyte (BAC)-derived exosomal miRNAs; among them, miR-132-3p was significantly induced. The isolated BAC-derived exosomes suppressed expression of hepatic Srebf1, a predicted target of miR-132-3p. In an indirect co-culture system, BACs suppressed expression of hepatic Srebf1 and its target lipogenic genes; this effect was not seen with miR-132-3p-inhibited BACs. Srebf1 was experimentally validated as an miR-132-3p target. Cold stimuli consistently induced miR-132-3p expression in BAT and attenuated Srebf1 expression in the liver. Our results suggest that BAT-derived exosomal miR-132-3p acts as an endocrine factor that regulates hepatic lipogenesis for cold adaptation.

Age-dependent increase in angiopoietin-like protein 2 accelerates skeletal muscle loss in mice.

Skeletal muscle atrophy, or sarcopenia, is commonly observed in older individuals and in those with chronic disease and is associated with decreased quality of life. There is recent medical and broad concern that sarcopenia is rapidly increasing worldwide as populations age. At present, strength training is the only effective intervention for preventing sarcopenia development, but it is not known how this exercise regimen counteracts this condition. Here, we report that expression of the inflammatory mediator angiopoietin-like protein 2 (ANGPTL2) increases in skeletal muscle of aging mice. Moreover, in addition to exhibiting increased inflammation and accumulation of reactive oxygen species (ROS), denervated atrophic skeletal muscles in a mouse model of denervation-induced muscle atrophy had increased ANGPTL2 expression. Interestingly, mice with a skeletal myocyte-specific Angptl2 knockout had attenuated inflammation and ROS accumulation in denervated skeletal muscle, accompanied by increased satellite cell activity and inhibition of muscular atrophy compared with mice harboring wildtype Angptl2 Moreover, consistent with these phenotypes, wildtype mice undergoing exercise training displayed decreased ANGPTL2 expression in skeletal muscle. In conclusion, ANGPTL2 up-regulation in skeletal myocytes accelerates muscle atrophy, and exercise-induced attenuation of ANGPTL2 expression in those tissues may partially explain how exercise training prevents sarcopenia.

Hypoxia reduces HNF4α/MODY1 protein expression in pancreatic ß-cells by activating AMP-activated protein kinase.

Hypoxia plays a role in the deterioration of ß-cell function. Hepatocyte nuclear factor 4α (HNF4α) has an important role in pancreatic ß-cells, and mutations of the human HNF4A gene cause a type of maturity-onset diabetes of the young (MODY1). However, it remains unclear whether hypoxia affects the expression of HNF4α in ß-cells. Here, we report that hypoxia reduces HNF4α protein expression in ß-cells. Hypoxia-inducible factor was not involved in the down-regulation of HNF4α under hypoxic conditions. The down-regulation of HNF4α was dependent on the activation of AMP-activated protein kinase (AMPK), and the reduction of HNF4α protein expression by metformin, an AMPK activator, and hypoxia was inhibited by the overexpression of a kinase-dead (KD) form of AMPKα2. In addition, hypoxia decreased the stability of the HNF4α protein, and the down-regulation of HNF4α was sensitive to proteasome inhibitors. Adenovirus-mediated overexpression of KD-AMPKα2 improved insulin secretion in metformin-treated islets, hypoxic islets, and ob/ob mouse islets. These results suggest that down-regulation of HNF4α could be of importance in ß-cell dysfunction by hypoxia.

Sirtuin 7 is involved in the consolidation of fear memory in mice.

Sirtuin 7 (SIRT7) is an NAD+-dependent deacetylase/deacylase, and is involved in a variety of biological processes relevant to the transcription of rRNA, the DNA damage response, tumorigenesis, and metabolism. SIRT7 mRNA is expressed ubiquitously, including in the brain, but there is no detailed information about the anatomical distribution and functional role of SIRT7 in the brain. Here, we demonstrated that SIRT7 is widely expressed in the mouse brain, including in the cortex, striatum, thalamus, hippocampus, and amygdala. Behavioral examinations revealed that Sirt7 knockout (KO) and control mice showed similar levels of freezing behavior immediately after a fear response, but a significant decrease of freezing behavior at 24 h after fear conditioning was observed in Sirt7 KO mice. Histological analysis revealed that there is no apparent structural abnormality of the amygdala and hippocampus, which are regions involved in fear memory consolidation, in Sirt7 KO mice. Our results indicate that SIRT7 is involved in the consolidation of fear memory.

SIRT7 is an important regulator of cartilage homeostasis and osteoarthritis development.

Sirtuins (SIRT1-7) are NAD+-dependent deacetylase/deacylases that regulate a wide variety of biological functions. Although the roles of sirtuins in cartilage homeostasis and cartilage diseases have been well studied, there is no information on the contribution of SIRT7 to cartilage homeostasis and osteoarthritis (OA) pathologies. Here, we demonstrate that Sirt7 knockout mice are resistant to the development of aging-associated OA and forced exercise-induced OA. Attenuation of Sirt7 in the murine chondrogenic cell line ATDC5 increased the deposition of a glycosaminoglycan-rich extracellular matrix and the mRNA expression of extracellular matrix components such as Col2a1 and Acan. Mechanistically, we found that SIRT7 suppressed the transcriptional activity of SOX9, which is an important transcription factor in chondrocytes, and that the enzymatic activity of SIRT7 was required for its function. Our results indicate that SIRT7 is a novel important regulator of cartilage homeostasis and OA development.

Sirtuin 7-dependent deacetylation of DDB1 regulates the expression of nuclear receptor TR4.

Sirtuin 7 (SIRT7) is an NAD+-dependent deacetylase/deacylase, but only a limited number of SIRT7 substrates have been identified. Recently, we found that Sirt7 knockout mice are resistant to high-fat diet-induced fatty liver, and that SIRT7 positively regulates the protein level of TR4, a nuclear receptor involved in lipid metabolism, by inhibiting the CUL4B/DDB1/DCAF1 E3 ubiquitin ligase complex. However, the mechanism by which SIRT7 inhibits the E3 ubiquitin ligase complex was not identified. Here, we demonstrate that SIRT7 binds directly to DDB1 and deacetylates DDB1 at Lys1121. K1121R-DDB1 (a deacetylation-mimicking mutant) displayed reduced binding with DCAF1. The expression of TR4 protein and TR4 target genes, including Cd36, Cidea, Cidec and Pparg1, was increased in K1121R-DDB1-overexpressing Hepa1-6 cells compared to WT-DDB1-overexpressing cells. Our results indicate that the SIRT7-mediated deacetylation of K1121 attenuates the activity of the CUL4B/DDB1/DCAF1 E3 ubiquitin ligase complex by reducing binding between DDB1 and DCAF1, leading to the increased expression of TR4.

Sirt7 Contributes to Myocardial Tissue Repair by Maintaining Transforming Growth Factor-ß Signaling Pathway.

BACKGROUND: Sirt7, 1 of the 7 members of the mammalian sirtuin family, promotes oncogenic transformation. Tumor growth and metastasis require fibrotic and angiogenic responses. Here, we investigated the role of Sirt7 in cardiovascular tissue repair process. METHODS AND RESULTS: In wild-type mice, Sirt7 expression increased in response to acute cardiovascular injury, including myocardial infarction and hind-limb ischemia, particularly at the active wound healing site. Compared with wild-type mice, homozygous Sirt7-deficient (Sirt7(-/-)) mice showed susceptibility to cardiac rupture after myocardial infarction, delayed blood flow recovery after hind-limb ischemia, and impaired wound healing after skin injury. Histological analysis showed reduced fibrosis, fibroblast differentiation, and inflammatory cell infiltration in the border zone of infarction in Sirt7(-/-) mice. In vitro, Sirt7(-/-) mouse-derived or Sirt7 siRNA-treated cardiac fibroblasts showed reduced transforming growth factor-ß signal activation and low expression levels of fibrosis-related genes compared with wild-type mice-derived or control siRNA-treated cells. These changes were accompanied by reduction in transforming growth factor receptor I protein. Loss of Sirt7 activated autophagy in cardiac fibroblasts. Transforming growth factor-ß receptor I downregulation induced by loss of Sirt7 was blocked by autophagy inhibitor, and interaction of Sirt7 with protein interacting with protein kinase-Cα was involved in this process. CONCLUSION: Sirt7 maintains transforming growth factor receptor I by modulating autophagy and is involved in the tissue repair process.

[Osteocalcin and osteonetwork].

Nearly nine years ago, new mechanism that bone secretes a hormone called osteocalcin and regulates glucose/energy metabolism was discovered. To date the study of osteocalcin as the bone hormone was progressed well, and the new roles in various tissues, such as glucose metabolism, male fertility, and development of the brain, are demonstrated. On the other hand, signaling pathway of osteocalcin has not yet been fully understood, though its receptor was reported. This review focuses on the diverse roles of osteocalcin and also on the future task that should be solved.

Roles of hepatic glucokinase in intertissue metabolic communication: Examination of novel liver-specific glucokinase knockout mice.

Glucokinase is expressed principally in pancreatic ß-cells and hepatocytes, and catalyzes the phosphorylation of glucose to glucose-6-phosphate, a rate-limiting step of glycolysis. To better understand the roles of hepatic glucokinase, we generated Gck knockout mice by ablating liver-specific exon 1b. The knockout mice exhibited impaired glucose tolerance, decreased hepatic glycogen content, and reduced Pklr and Fas gene expression in the liver, indicating that hepatic glucokinase plays important roles in glucose metabolism. It has also been reported that hepatic glucokinase regulates the expression of thermogenesis-related genes in brown adipose tissue (BAT) and insulin secretion in response to glucose. However, the liver-specific Gck knockout mice displayed neither altered expression of thermogenesis-related genes in BAT nor impaired insulin secretion by ß-cells under a normal chow diet. These results suggest that chronic suppression of hepatic glucokinase has a small influence on intertissue (liver-to-BAT as well as liver-to-ß-cell) metabolic communication.

Anks4b, a novel target of HNF4α protein, interacts with GRP78 protein and regulates endoplasmic reticulum stress-induced apoptosis in pancreatic ß-cells.

Mutations of the HNF4A gene cause a form of maturity-onset diabetes of the young (MODY1) that is characterized by impairment of pancreatic ß-cell function. HNF4α is a transcription factor belonging to the nuclear receptor superfamily (NR2A1), but its target genes in pancreatic ß-cells are largely unknown. Here, we report that ankyrin repeat and sterile α motif domain containing 4b (Anks4b) is a target of HNF4α in pancreatic ß-cells. Expression of Anks4b was decreased in both ßHNF4α KO islets and HNF4α knockdown MIN6 ß-cells, and HNF4α activated Anks4b promoter activity. Anks4b bound to glucose-regulated protein 78 (GRP78), a major endoplasmic reticulum (ER) chaperone protein, and overexpression of Anks4b enhanced the ER stress response and ER stress-associated apoptosis of MIN6 cells. Conversely, suppression of Anks4b reduced ß-cell susceptibility to ER stress-induced apoptosis. These results indicate that Anks4b is a HNF4α target gene that regulates ER stress in ß-cells by interacting with GRP78, thus suggesting that HNF4α is involved in maintenance of the ER.

Inhibition of H3K18 deacetylation of Sirt7 by Myb-binding protein 1a (Mybbp1a).

Sirt7 localizes in the nucleus (enriched in the nucleolus) and is an NAD(+)-dependent deacetylase with high selectivity for the acetylated lysine 18 of histone H3 (H3K18Ac). It has been reported that Sirt7 is necessary for maintaining the fundamental properties of the cancer cell phenotype and stabilizing the tumorigenicity of human cancer via deacetylation of H3K18Ac. However, the regulators of Sirt7 deacetylase activity are unknown. Myb-binding protein 1a (Mybbp1a) is reported to interact with and regulate the function of a number of transcription factors. In the present study, we demonstrated that Mybbp1a binds to Sirt7 in vitro and in vivo. Serial deletion studies indicated that N- and C-terminal regions of Sirt7 and C-terminal region of Mybbp1a are important for the binding. Furthermore, transfection experiments showed that Mybbp1a is capable of inhibiting the deacetylation activity of H3K18Ac by Sirt7. Our findings demonstrate that Mybbp1a is a novel negative regulator of Sirt7.

The Emerging Role of SIRT7 in Glucose and Lipid Metabolism.

Sirtuins (SIRT1-7 in mammals) are a family of NAD+-dependent lysine deacetylases and deacylases that regulate diverse biological processes, including metabolism, stress responses, and aging. SIRT7 is the least well-studied member of the sirtuins, but accumulating evidence has shown that SIRT7 plays critical roles in the regulation of glucose and lipid metabolism by modulating many target proteins in white adipose tissue, brown adipose tissue, and liver tissue. This review focuses on the emerging roles of SIRT7 in glucose and lipid metabolism in comparison with SIRT1 and SIRT6. We also discuss the possible implications of SIRT7 inhibition in the treatment of metabolic diseases such as type 2 diabetes and obesity.

Identification of hepatocyte growth factor activator (Hgfac) gene as a target of HNF1α in mouse ß-cells.

HNF1α is a transcription factor that is expressed in pancreatic ß-cells and mutations of the HNF1α gene cause a form of monogenic diabetes. To understand the role of HNF1α in pancreatic ß-cells, we established the MIN6 ß-cell line that stably expressed HNF1α-specific shRNA. Expression of the gene encoding hepatocyte growth factor (HGF) activator (Hgfac), a serine protease that efficiently activates HGF, was decreased in HNF1α KD-MIN6 cells. Down-regulation of Hgfac expression was also found in the islets of HNF1α (+/-) mice. Reporter gene analysis and the chromatin immunoprecipitation assay indicated that HNF1α directly regulates the expression of Hgfac in ß-cells. It has been reported that HGF has an important influence on ß-cell mass and ß-cell function. Thus, HNF1α might regulate ß-cell mass or function at least partly by modulating Hgfac expression.

[Diabetes mellitus and osteoporosis. Insulin signaling and bone/glucose/energy metabolism].

Recently, we reported that insulin signaling in osteoblasts was a positive regulator of bone acquisition, but also of bone resorption. Interestingly, insulin signaling in osteoblasts activated osteocalcin embedded in bone ECM by stimulating resorption activity in osteoclast. It has been demonstrated that activated osteocalcin acts as hormone and regulates glucose metabolism through increasing insulin secretion from pancreas. The interaction of bone and glucose metabolism established by these studies will be important to a study of the two scientific fields in the future, particularly a clinical field.

Hemispheric asymmetry of chromatic motion perception.

The present study investigated hemispheric symmetry of cortical functions, in terms of the chromatic motion mechanism. A series of experiments examined the visual sensitivities to chromatic and achromatic stimuli with or without motion, presented in either of the two (left or right) visual hemifields. Experiment 1 measured, individually, the subjective isoluminance of red/green stimuli for each visual field. Experiment 2 examined the visual field differences of the detection thresholds for static stimuli with the isoluminant color contrast and achromatic luminance contrast. Subsequent experiments measured contrast thresholds for motion detection (Experiment 3) and motion direction discrimination (Experiment 4) with both chromatic and achromatic stimuli. No visual field differences between thresholds were found in Experiments 1 and 2, whereas in Experiments 3 and 4, thresholds for the chromatic conditions were found to be lower in the left than in the right visual field, suggesting functional lateralization of the early motion mechanism with chromatic information in motion detection and direction discrimination.