Distribution of Nicotinamide Mononucleotide after Intravenous Injection in Normal and Ischemic Stroke Mice.

OBJECTIVE: This study determined for the first time the distribution of intravenous nicotinamide mononucleotide (NMN) and its metabolite nicotinamide adenine dinucleotide (NAD) in normal and ischemic stroke mice, examined the therapeutic effect of NMN on ischemic brain infarction, and evaluated acute toxicity of NMN after intravenous injection of NMN. METHODS: NMN and NAD levels were determined using ultra-high-performance liquid chromatography tandem mass spectrometry in biological samples from mice with or without middle cerebral artery occlusion (MCAO) at different time points post intravenous NMN injection (300 mg/kg). Brain infarction was evaluated 24 h post-MCAO. 2 g/kg NMN was used in the acute toxicity test. RESULTS: Under either normal or MCAO conditions, serum NMN levels sharply increased after intravenous NMN administration and then decreased rapidly within 15 min, while serum NAD levels remained unchanged during 30 min observation. Both substances displayed tissue accumulation over time and stored faster under MCAO conditions, with kidney having the highest concentrations. Particularly, NMN accumulated earlier than NAD in the brain. Moreover, NMN reduced cerebral infarction at 24 h post-MCAO. No acute toxicity was observed for 14 days. NRK1 and SLC12A8 involved in two pathways of NMN uptake exhibited the highest expressions in kidney and colon, respectively, among 11 different tissues. CONCLUSION: NMN distributes to various tissues after intravenous injection and has the ability to enter the brain to boost NAD levels, and exhibits safety and therapeutic effect on acute ischemic stroke injury. High renal distribution of NMN indicates its importance in the kidney.

Nur77-activated lncRNA WFDC21P attenuates hepatocarcinogenesis via modulating glycolysis.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related mortality worldwide. Orphan nuclear receptor Nur77, which is low expressed in HCC, functions as a tumor suppressor to suppress HCC. However, the detailed mechanism is still not well understood. Here, we demonstrate that Nur77 could inhibit HCC development via transcriptional activation of the lncRNA WAP four-disulfide core domain 21 pseudogene (WFDC21P). Nur77 binds to its response elements on the WFDC21P promoter to directly induce WFDC21P transcription, which inhibits HCC cell proliferation, tumor growth, and tumor metastasis both in vitro and in vivo. In clinical HCC samples, WFDC21P expression positively correlated with that of Nur77, and the loss of WFDC21P is associated with worse prognosis. Mechanistically, WFDC21P could inhibit glycolysis by simultaneously interacting with PFKP and PKM2, two key enzymes in glycolysis. These interactions not only abrogate the tetramer formation of PFKP to impede its catalytic activity but also prevent the nuclear translocation of PKM2 to suppress its function as a transcriptional coactivator. Cytosporone-B (Csn-B), an agonist for Nur77, could stimulate WFDC21P expression and suppress HCC in a WFDC21P-dependent manner. Therefore, our study reveals a new HCC suppressor and connects the glycolytic remodeling of HCC with the Nur77-WFDC21P-PFKP/PKM2 axis.

Nuclear Receptor Nur77 Facilitates Melanoma Cell Survival under Metabolic Stress by Protecting Fatty Acid Oxidation.

Fatty acid oxidation (FAO) is crucial for cells to overcome metabolic stress by providing ATP and NADPH. However, the mechanism by which FAO is regulated in tumors remains elusive. Here we show that Nur77 is required for the metabolic adaptation of melanoma cells by protecting FAO. Glucose deprivation activates ERK2 to phosphorylate and induce Nur77 translocation to the mitochondria, where Nur77 binds to TPß, a rate-limiting enzyme in FAO. Although TPß activity is normally inhibited by oxidation under glucose deprivation, the Nur77-TPß association results in Nur77 self-sacrifice to protect TPß from oxidation. FAO is therefore able to maintain NADPH and ATP levels and prevent ROS increase and cell death. The Nur77-TPß interaction further promotes melanoma metastasis by facilitating circulating melanoma cell survival. This study demonstrates a novel regulatory function of Nur77 with linkage of the FAO-NADPH-ROS pathway during metabolic stress, suggesting Nur77 as a potential therapeutic target in melanoma.

Nicotinamide mononucleotide attenuates brain injury after intracerebral hemorrhage by activating Nrf2/HO-1 signaling pathway.

Replenishment of NAD+ has been shown to protect against brain disorders such as amyotrophic lateral sclerosis and ischemic stroke. However, whether this intervention has therapeutic effects in intracerebral hemorrhage (ICH) is unknown. In this study, we sought to determine the potential therapeutic value of replenishment of NAD+ in ICH. In a collagenase-induced ICH (cICH) mouse model, nicotinamide mononucleotide (NMN), a key intermediate of nicotinamide adenine dinucleotide (NAD+) biosynthesis, was administrated at 30 minutes post cICH from tail vein to replenish NAD+. NMN treatment did not decrease hematoma volume and hemoglobin content. However, NMN treatment significantly reduced brain edema, brain cell death, oxidative stress, neuroinflammation, intercellular adhesion molecule-1 expression, microglia activation and neutrophil infiltration in brain hemorrhagic area. Mechanistically, NMN enhanced the expression of two cytoprotective proteins: heme oxygenase 1 (HO-1) and nuclear factor-like 2 (Nrf2). Moreover, NMN increased the nuclear translocation of Nrf2 for its activation. Finally, a prolonged NMN treatment for 7 days markedly promoted the recovery of body weight and neurological function. These results demonstrate that NMN treats brain injury in ICH by suppressing neuroinflammation/oxidative stress. The activation of Nrf2/HO-1 signaling pathway may contribute to the neuroprotection of NMN in ICH.

Neuroprotective Efficacy of an Aminopropyl Carbazole Derivative P7C3-A20 in Ischemic Stroke.

AIM: NAMPT is a novel therapeutic target of ischemic stroke. The aim of this study was to investigate the effect of a potential NAMPT activator, P7C3-A20, an aminopropyl carbazole derivative, on ischemic stroke. METHODS: In vitro study, neuron protection effect of P7C3-A20 was investigated by co-incubation with primary neurons subjected to oxygen-glucose deprivation (OGD) or oxygen-glucose deprivation/reperfusion (OGD/R) injury. In vivo experiment, P7C3-A20 was administrated in middle cerebral artery occlusion (MCAO) rats and infarct volume was examined. Lastly, the brain tissue nicotinamide adenine dinucleotide (NAD) levels were detected in P7C3-A20 treated normal or MCAO mice. RESULTS: Cell viability, morphology, and Tuj-1 staining confirmed the neuroprotective effect of P7C3-A20 in OGD or OGD/R model. P7C3-A20 administration significantly reduced cerebral infarction in MCAO rats. Moreover, brain NAD levels were elevated both in normal and MCAO mice after P7C3-A20 treatment. CONCLUSIONS: P7C3-A20 has neuroprotective effect in cerebral ischemia. The study contributes to the development of NAMPT activators against ischemic stroke and expands the horizon of the neuroprotective effect of aminopropyl carbazole chemicals.

Hepatic NAD(+) deficiency as a therapeutic target for non-alcoholic fatty liver disease in ageing.

BACKGROUND AND PURPOSE: Ageing is an important risk factor of non-alcoholic fatty liver disease (NAFLD). Here, we investigated whether the deficiency of nicotinamide adenine dinucleotide (NAD(+) ), a ubiquitous coenzyme, links ageing with NAFLD. EXPERIMENTAL APPROACH: Hepatic concentrations of NAD(+) , protein levels of nicotinamide phosphoribosyltransferase (NAMPT) and several other critical enzymes regulating NAD(+) biosynthesis, were compared in middle-aged and aged mice or patients. The influences of NAD(+) decline on the steatosis and steatohepatitis were evaluated in wild-type and H247A dominant-negative, enzymically-inactive NAMPT transgenic mice (DN-NAMPT) given normal or high-fat diet (HFD). KEY RESULTS: Hepatic NAD(+) level decreased in aged mice and humans. NAMPT-controlled NAD(+) salvage, but not de novo biosynthesis pathway, was compromised in liver of elderly mice and humans. Given normal chow, middle-age DN-NAMPT mice displayed systemic NAD(+) reduction and had moderate NAFLD phenotypes, including lipid accumulation, enhanced oxidative stress, triggered inflammation and impaired insulin sensitivity in liver. All these NAFLD phenotypes, especially release of pro-inflammatory factors, Kupffer cell accumulation, monocytes infiltration, NLRP3 inflammasome pathway and hepatic fibrosis (Masson's staining and α-SMA staining), deteriorated further under HFD challenge. Oral administration of nicotinamide riboside, a natural NAD(+) precursor, completely corrected these NAFLD phenotypes induced by NAD(+) deficiency alone or HFD, whereas adenovirus-mediated SIRT1 overexpression only partially rescued these phenotypes. CONCLUSIONS AND IMPLICATIONS: These results provide the first evidence that ageing-associated NAD(+) deficiency is a critical risk factor for NAFLD, and suggest that supplementation with NAD(+) substrates may be a promising therapeutic strategy to prevent and treat NAFLD.

Depletion of NAD pool contributes to impairment of endothelial progenitor cell mobilization in diabetes.

OBJECTIVE: The impaired mobilization of endothelial progenitor cells (EPCs) from bone marrow (BM) critically contributes to the diabetes-associated vascular complications. Here, we investigated the relationship between the nicotinamide phosphoribosyltransferase (NAMPT)-controlled nicotinamide adenine dinucleotide (NAD) metabolism and the impaired mobilization of BM-derived EPCs in diabetic condition. METHODS: The NAMPT-NAD pool in BM and BM-derived EPCs in wild-type (WT) and diabetic db/db mice was determined. Nicotinamide, a natural substrate for NAD biosynthesis, was administrated for 2weeks in db/db mice to examine the influence of enhancing NAD pool on BM and blood EPCs number. The modulations of stromal cell-derived factor-1α (SDF-1α) and endothelial nitric oxide synthase (eNOS) protein in BM were measured using immunoblotting. The EPCs intracellular NAMPT level and NAD concentration, as well as the blood EPCs number, were compared between 9 healthy people and 16 patients with type 2 diabetes mellitus (T2DM). The T2DM patients were treated with nicotinamide for two weeks and then the blood EPCs number was determined. Moreover, the association between blood EPCs numbers and EPCs intracellular NAD(+)/NAMPT protein levels in 21 healthy individuals was determined. RESULTS: We found that NAD concentration and NAMPT expression in BM and BM-derived EPCs of db/db mice were significantly lower than those in WT mice BM. Enhancing NAD pool not only increased the EPCs intracellular NAD concentration and blood EPCs number, but also improved post-ischemic wound healing and blood reperfusion in db/db mice with hind-limb ischemia model. Enhancing NAD pool rescued the impaired modulations of stromal cell-derived factor-1α (SDF-1α) and endothelial nitric oxide synthase (eNOS) protein levels in db/db mice BM upon hind-limb ischemia. In addition, enhancing NAD pool significantly inhibited PARP and caspase-3 activates in db/db mice BM. The intracellular NAMPT-NAD pool was positively associated with blood EPCs number in healthy individuals. At last, we found that the EPC intracellular NAMPT and NAD(+) levels were reduced in T2DM patients and enhancing NAD pool elevated the circulating blood EPCs number in T2DM patients. CONCLUSION: Our results indicate that the depletion of NAD pool may contribute to the impairment of EPCs mobilization in diabetic condition, and imply the potential therapeutic value of nicotinamide in the prevention and treatment for cardiovascular complications of diabetes.

Adipocyte Metrnl Antagonizes Insulin Resistance Through PPARγ Signaling.

Adipokines play important roles in metabolic homeostasis and disease. We have recently identified a novel adipokine Metrnl, also known as Subfatin, for its high expression in subcutaneous fat. Here, we demonstrate a prodifferentiation action of Metrnl in white adipocytes. Adipocyte-specific knockout of Metrnl exacerbates insulin resistance induced by high-fat diet (HFD), whereas adipocyte-specific transgenic overexpression of Metrnl prevents insulin resistance induced by HFD or leptin deletion. Body weight and adipose content are not changed by adipocyte Metrnl. Consistently, no correlation is found between serum Metrnl level and BMI in humans. Metrnl promotes white adipocyte differentiation, expandability, and lipid metabolism and inhibits adipose inflammation to form functional fat, which contributes to its activity against insulin resistance. The insulin sensitization of Metrnl is blocked by PPARγ inhibitors or knockdown. However, Metrnl does not drive white adipose browning. Acute intravenous injection of recombinant Metrnl has no hypoglycemic effect, and 1-week intravenous administration of Metrnl is unable to rescue insulin resistance exacerbated by adipocyte Metrnl deficiency. Our results suggest adipocyte Metrnl controls insulin sensitivity at least via its local autocrine/paracrine action through the PPARγ pathway. Adipocyte Metrnl is an inherent insulin sensitizer and may become a therapeutic target for insulin resistance.

Discovery of Novel Inhibitors and Fluorescent Probe Targeting NAMPT.

Nicotinamide phosphoribosyltransferase (NAMPT) is a promising antitumor target. Novel NAMPT inhibitors with diverse chemotypes are highly desirable for development of antitumor agents. Using high throughput screening system targeting NAMPT on a chemical library of 30000 small-molecules, we found a non-fluorescent compound F671-0003 and a fluorescent compound M049-0244 with excellent in vitro activity (IC50: 85 nM and 170 nM respectively) and anti-proliferative activity against HepG2 cells. These two compounds significantly depleted cellular NAD levels. Exogenous NMN rescued their anti-proliferative activity against HepG2 cells. Structure-activity relationship study proposed a binding mode for NAMPT inhibitor F671-0003 and highlighted the importance of hydrogen bonding, hydrophobic and π-π interactions in inhibitor binding. Imaging study provided the evidence that fluorescent compound M049-0244 (3 µM) significantly stained living HepG2 cells. Cellular fluorescence was further verified to be NAMPT dependent by using RNA interference and NAMPT over expression transgenic mice. Our findings provide novel antitumor lead compounds and a "first-in-class" fluorescent probe for imaging NAMPT.

Regenerative Neurogenesis After Ischemic Stroke Promoted by Nicotinamide Phosphoribosyltransferase-Nicotinamide Adenine Dinucleotide Cascade.

BACKGROUND AND PURPOSE: Nicotinamide adenine dinucleotide (NAD) is a ubiquitous fundamental metabolite. Nicotinamide phosphoribosyltransferase (Nampt) is the rate-limiting enzyme for mammalian NAD salvage synthesis and has been shown to protect against acute ischemic stroke. In this study, we investigated the role of Nampt-NAD cascade in brain regeneration after ischemic stroke. METHODS: Nampt transgenic (Nampt-Tg) mice and H247A mutant enzymatic-dead Nampt transgenic (ΔNampt-Tg) mice were subjected with experimental cerebral ischemia by middle cerebral artery occlusion. Activation of neural stem cells, neurogenesis, and neurological function recovery were measured. Besides, nicotinamide mononucleotide and NAD, two chemical enzymatic product of Nampt, were administrated in vivo and in vitro. RESULTS: Compared with wild-type mice, Nampt-Tg mice showed enhanced number of neural stem cells, improved neural functional recovery, increased survival rate, and accelerated body weight gain after middle cerebral artery occlusion, which were not observed in ΔNampt-Tg mice. A delayed nicotinamide mononucleotide administration for 7 days with the first dose at 12 hours post middle cerebral artery occlusion did not protect acute brain infarction and neuronal deficit; however, it still improved postischemic regenerative neurogenesis. Nicotinamide mononucleotide and NAD(+) promoted proliferation and differentiation of neural stem cells in vitro. Knockdown of NAD-dependent deacetylase sirtuin 1 (SIRT1) and SIRT2 inhibited the progrowth action of Nampt-NAD axis, whereas knockdown of SIRT1, SIRT2, and SIRT6 compromised the prodifferentiation effect of Nampt-NAD axis. CONCLUSIONS: Our data demonstrate that the Nampt-NAD cascade may act as a centralizing switch in postischemic regeneration through controlling different sirtuins and therefore represent a promising therapeutic target for long-term recovery of ischemic stroke.

Chronic nicotine treatment enhances vascular smooth muscle relaxation in rats.

AIM: To investigate the effect of chronic nicotine treatment on vascular function and to identify the underlying mechanisms. METHODS: Adult rats were treated with nicotine (3 mg·kg(-1)·d(-1), sc) for 6 weeks. After the rats were sacrificed, aortic rings were prepared for detecting vascular reactivity, and thoracic aorta and periaortic fat samples were collected for histological and molecular biology studies. RESULTS: Chronic nicotine treatment significantly reduced periaortic fat, and specifically enhanced smooth muscle relaxation without altering the aortic adventitial fat and endothelium function. Pretreatment with the soluble guanylyl cyclase inhibitor ODQ (3 µmol/L) or PKG inhibitor Rp-8-Br-PET-cGMP (30 µmol/L) abolished the nicotine-induced enhancement of smooth muscle relaxation, whereas the cGMP analogue 8-Br-cGMP could mimic the nicotine-induced enhancement of smooth muscle relaxation. However, the chronic nicotine treatment did not alter PKG protein expression and activity in aortic media. CONCLUSION: Chronic nicotine treatment enhances vascular smooth muscle relaxation of rats via activation of PKG pathway.

ARRB1/ß-arrestin-1 mediates neuroprotection through coordination of BECN1-dependent autophagy in cerebral ischemia.

Autophagy, a highly conserved process conferring cytoprotection against stress, contributes to the progression of cerebral ischemia. ß-arrestins are multifunctional proteins that mediate receptor desensitization and serve as important signaling scaffolds involved in numerous physiopathological processes. Here, we show that both ARRB1 (arrestin, ß 1) and ARRB2 (arrestin, ß 2) were upregulated by cerebral ischemic stress. Knockout of Arrb1, but not Arrb2, aggravated the mortality, brain infarction, and neurological deficit in a mouse model of cerebral ischemia. Accordingly, Arrb1-deficient neurons exhibited enhanced cell injury upon oxygen-glucose deprivation (OGD), an in vitro model of ischemia. Deletion of Arrb1 did not affect the cerebral ischemia-induced inflammation, oxidative stress, and nicotinamide phosphoribosyltransferase upregulation, but markedly suppressed autophagy and induced neuronal apoptosis/necrosis in vivo and in vitro. Additionally, we found that ARRB1 interacted with BECN1/Beclin 1 and PIK3C3/Vps34, 2 major components of the BECN1 autophagic core complex, under the OGD condition but not normal conditions in neurons. Finally, deletion of Arrb1 impaired the interaction between BECN1 and PIK3C3, which is a critical event for autophagosome formation upon ischemic stress, and markedly reduced the kinase activity of PIK3C3. These findings reveal a neuroprotective role for ARRB1, in the context of cerebral ischemia, centered on the regulation of BECN1-dependent autophagosome formation.

Extracellular visfatin has nicotinamide phosphoribosyltransferase enzymatic activity and is neuroprotective against ischemic injury.

AIM: Visfatin, a novel adipokine, is predominantly produced by visceral adipose tissue and exists in intracellular and extracellular compartments. The intracellular form of visfatin is proved to be nicotinamide phosphoribosyltransferase (NAMPT) and exhibits neuroprotection through maintaining intracellular NAD(+) pool. However, whether extracellular form of visfatin has NAMPT activity and the effect of extracellular visfatin in cerebral ischemia are unknown. METHODS AND RESULTS: Plasma concentrations of visfatin, NAD(+) , and ATP were increased in mice upon cerebral ischemia. Cultured glia, but not neuron, was able to secrete visfatin. Oxygen-glucose deprivation (OGD) stress increased the secretion of visfatin from glia. Extracellular recombinant mouse wild-type visfatin, but not mouse H247A-mutant enzymatic-dead visfatin, had NAMPT enzymatic function in vitro. Treatment of wild-type visfatin, but not H247A-mutant enzymatic-dead visfatin, significantly attenuated detrimental effect of OGD on the cell viability and apoptosis in both cultured mouse neuron and glia. Treatment of neutralizing antibody, abolished the protective effect of extracellular visfatin on cell viability, but failed to block the antiapoptotic effect of extracellular visfatin. At last, we observed that plasma visfatin concentrations decreased in 6-month-old but not 3-month-old SHR-SP compared with that in age-matched Wistar-Kyoto rats. Inhibition of NAMPT enzymatic function of visfatin (by FK866) accelerated the occurrence of stroke in SHR-SP. CONCLUSIONS: Extracellular visfatin has NAMPT enzymatic activity and maybe be neuroprotective just as intracellular visfatin in cerebral ischemic injury.

Vascular smooth muscle cell apoptosis is an early trigger for hypothyroid atherosclerosis.

AIMS: Endothelial dysfunction is an initial and vascular smooth muscle cell (VSMC) apoptosis, a later step of atherosclerosis. Hypothyroidism accelerates atherosclerosis. However, the early events responsible for this pro-atherosclerotic effect are unclear. METHODS AND RESULTS: Rats were resistant to induction of atherosclerosis by high cholesterol diet alone, but became susceptible in hypothyroid state achieved by administration of propylthiouracil (PTU) for 6 weeks. VSMC dysfunction and apoptosis were obvious within 1 week after PTU treatment, without signs of endothelial dysfunction. This early VSMC damage was caused by hypothyroidism but not the high cholesterol diet. In ApoE knockout mice, PTU-induced hypothyroidism triggered early VSMC apoptosis, increased oxidative stress, and accelerated atherosclerosis development. Thyroid hormone supplementation (T4, 10, or 50 µg/kg) prevented atherogenic phenotypes in hypothyroid rats and mice. In rats, thyroidectomy caused severe hypothyroidism 5 days after operation, which also led to rapid VSMC dysfunction and apoptosis. In vitro studies did not show a direct toxic effect of PTU on VSMCs. In contrast, thyroid hormone (T3, 0.75 µg/L plus T4, 50 nmol/L) exerted a direct protection against VSMC apoptosis, which was reduced by knockdown of TRα1, rather than TRß1 and TRß2 receptors. TRα1-mediated inhibition of apoptotic signalling of JNKs and caspase-3 contributed to the anti-apoptotic action of thyroid hormone. CONCLUSION: These findings provide an in vivo example for VSMC apoptosis as an early trigger of hypothyroidism-associated atherosclerosis, and reveal activation of TRα1 receptors to prevent VSMC apoptosis as a therapeutic strategy in this disease.

Subfatin is a novel adipokine and unlike Meteorin in adipose and brain expression.

AIMS: Adipose tissue releases adipokines that play important roles in metabolic and cardio-cerebro-vascular homeostasis. This study was to discover novel adipokines using caloric restriction model. METHODS: Adipokine candidates were captured by gene array and bioinformatics analysis and verified by preparation of recombinant protein and antibody. RESULTS: We established a potential secreted protein database containing 208 genes and identified a novel adipokine, Subfatin, that was the highest expressed in subcutaneous fat of both rodents and humans among 15 detected tissues. The secreted mammalian Subfatin was a glycosylated protein. Subfatin was located diffusely throughout the adipose tissue except lipid droplets, with comparable expression between adipocytes and stromal cells, but much lower expression in macrophages than adipocytes. Subfatin was downregulated in white adipose tissue of caloric restriction rats, whereas dramatically upregulated during white adipocyte differentiation as well as in white adipose tissue of diet-induced obese mice. Subfatin was annotated as Meteorin-like (Metrnl) in public databases, a similar transcript of Meteorin (Metrn, also known as glial cell differentiation regulator). Meteorin displayed a brain-specific expression and was scarce in various adipose tissues, in contrast to the tissue expression patterns of Subfatin. CONCLUSIONS: Subfatin is a novel adipokine regulated by adipogenesis and obesity, with tissue distribution different from its homologue Meteorin.

Nicotinamide phosphoribosyltransferase is required for the calorie restriction-mediated improvements in oxidative stress, mitochondrial biogenesis, and metabolic adaptation.

Calorie restriction (CR) is one of the most reproducible treatments for weight loss and slowing aging. However, how CR induces these metabolic alterations is not fully understood. In this work, we studied whether nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme for nicotinamide adenine dinucleotide biosynthesis, plays a role in CR-induced beneficial metabolic effects using a specific inhibitor of NAMPT (FK866). CR upregulated NAMPT mRNA and protein levels in rat skeletal muscle and white adipose tissue. Inhibition of NAMPT activity by FK866 in rats did not affect the SIRT1 upregulation by CR but suppressed the CR-induced SIRT1 activity and deacetylation of Forkhead box protein O1/peroxisome proliferator-activated receptor γ coactivator-1α. Inhibition of NAMPT activity by FK866 also attenuated the CR-induced SIRT3 activity, evidenced by deacetylation of superoxide dismutase-2. Furthermore, FK866 not only weakened the CR-induced decrease of oxidative stress (dichlorofluorescin signal, superoxide , and malondialdehyde levels), but also greatly attenuated the CR-induced improvements of antioxidative activity (total superoxide dismutase, glutathione, and glutathione/oxidized glutathione ratio) and mitochondrial biogenesis (mRNA levels of nuclear respiratory factor 1, cytochrome c oxidase IV, peroxisome proliferator-activated receptor-γ coactivator-1α, and transcription factor A, mitochondrial and citrate synthase activity). At last, FK866 blocked the CR-induced insulin sensitizing, Akt signaling activation, and endothelial nitric oxide synthase phosphorylation. Collectively, our data provide the first evidence that the CR-induced beneficial effects in oxidative stress, mitochondrial biogenesis, and metabolic adaptation require NAMPT.

Telemetric ambulatory arterial stiffness index, a predictor of cardio-cerebro-vascular mortality, is associated with aortic stiffness-determining factors.

BACKGROUND: Ambulatory arterial stiffness index (AASI) has been proposed as a new measure of arterial stiffness for predicting cardio-cerebro-vascular morbidity and mortality. However, there has been no research on the direct relationships between AASI and arterial stiffness-determining factors. METHODS: We utilized beat-to-beat intra-aortic blood pressure (BP) telemetry to characterize AASI in Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR). By determination of aortic structural components and analysis of their correlations with AASI, we provided the first direct evidence for the associations between AASI and arterial stiffness-determining factors including the collagen content and collagen/elastin. RESULTS: Ambulatory arterial stiffness index was positively correlated with pulse pressure in both WKY and SHR, less dependent on BP and BP variability than pulse pressure, and relatively stable, especially the number of BP readings not less than ~36. The correlations between AASI and aortic components were comparable for various AASI values derived from BP readings not less than ~36. Not only AASI but also BP variability and pulse pressure demonstrated a direct relationship with arterial stiffness. CONCLUSIONS: These findings indicate AASI may become a routine measure in human arterial stiffness assessment. It is recommended to use a cluster of parameters such as AASI, BP variability, and pulse pressure for evaluating arterial stiffness.

Protein tyrosine phosphatase 1B inhibits adipocyte differentiation and mediates TNFα action in obesity.

Protein tyrosine phosphatase 1B (PTP1B) is a negative regulator of systemic glucose and insulin homeostasis; however, its exact role in adipocytes is poorly understood. This study was to elucidate the role of PTP1B in adipocyte differentiation and its implication in obesity. During differentiation of 3T3-L1 white preadipocytes, PTP1B decreased progressively with adipocyte maturation. Lentivirus-mediated PTP1B overexpression in preadipocytes delayed adipocyte differentiation, shown as lack of mature adipocytes, low level of lipid accumulation, and down-regulation of main markers (PPARγ2, SREBP-1c, FAS and LPL). In contrast, lentivirus-mediated PTP1B knockdown accelerated adipocyte differentiation, demonstrated as full of mature adipocytes, high level of lipid accumulation, and up-regulation of main markers. Dominant-negative inhibition on endogenous PTP1B by lentivirus-mediated overexpression of PTP1B double mutant in Tyr-46 and Asp-181 residues (LV-D/A-Y/F) also stimulated adipogenesis, more efficient than PTP1B knockdown. Diet-induced obesity mice exhibited an up-regulation of PTP1B and TNFα accompanied by a down-regulation of PPARγ2 in white adipose tissue. TNFα recombinant protein impeded PTP1B reduction and inhibited adipocyte differentiation in vitro; this inhibitory effect was prevented by LV-D/A-Y/F. Moreover, PTP1B inhibitor treatment improved adipogenesis and suppressed TNFα in adipose tissue of obese mice. All together, PTP1B negatively regulates adipocyte development and may mediate TNFα action to impair adipocyte differentiation in obesity. Our study provides novel evidence for the importance of PTP1B in obesity and for the potential application of PTP1B inhibitors.