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.

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.

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.

Non-alcoholic fatty liver disease (NAFLD) is a worldwide public health problem with no established pharmacological therapy. Here, we explored the potential benefit of P7C3-A20, a novel aminopropyl carbazole compound with neuroprotective activity, in a NAFLD model, induced in mice by a high-fat diet (HFD).

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.

This study was to investigate the effect of nicotine on insulin sensitivity and explore the underlying mechanisms. Treatment of Sprague-Dawley rats with nicotine (3 mg/kg/day) for 6 weeks reduced 43% body weight gain and 65% blood insulin level, but had no effect on blood glucose level. Both insulin tolerance test and glucose tolerance test demonstrated that nicotine treatment enhanced insulin sensitivity. Pretreatment of rats with hexamethonium (20 mg/kg/day) to antagonize peripheral nicotinic receptors except for α7 nicotinic acetylcholine receptor (α7-nAChR) had no effect on the insulin sensitizing effect of nicotine. However, the insulin sensitizing effect but not the bodyweight reducing effect of nicotine was abrogated in α7-nAChR knockout mice. Further, chronic treatment with PNU-282987 (0.53 mg/kg/day), a selective α7-nAChR agonist, significantly enhanced insulin sensitivity without apparently modifying bodyweight not only in normal mice but also in AMP-activated kinase-α2 knockout mice, an animal model of insulin resistance with no sign of inflammation. Moreover, PNU-282987 treatment enhanced phosphorylation of signal transducer and activator of transcription 3 (STAT3) in skeletal muscle, adipose tissue and liver in normal mice. PNU-282987 treatment also increased glucose uptake by 25% in C2C12 myotubes and this effect was total abrogated by STAT3 inhibitor, S3I-201. All together, these findings demonstrated that nicotine enhanced insulin sensitivity in animals with or without insulin resistance, at least in part via stimulating α7-nAChR-STAT3 pathway independent of inflammation. Our results contribute not only to the understanding of the pharmacological effects of nicotine, but also to the identifying of new therapeutic targets against insulin resistance.

Stroke is the second leading cause of death and main cause of disability worldwide, but with few effective therapies. Although stem cell-based therapy has been proposed as an exciting regenerative medicine strategy for brain injury, there are limitations. The developed cerebral organoids (COs) represent a promising transplantation source for stroke that remains to be answered. Here, we transplanted COs at 55 days and explored the feasibility in the rat middle cerebral artery occlusion (MCAO) model of stroke. COs transplantation at 6 h or even 24 h after MCAO significantly reduces brain infarct volume and improves neurological motor function. Transplanted COs show the potential of multilineage differentiation to mimic in vivo cortical development, support motor cortex region-specific reconstruction, form neurotransmitter-related neurons, and achieve synaptic connection with host brain via in situ differentiation and cell replacement in stroke. Cells from transplanted COs show extensive migration into different brain regions along corpus callosum. The mechanisms underlying COs transplantation therapy are also associated with enhanced neurogenesis, synaptic reconstruction, axonal regeneration and angiogenesis, and decreased neural apoptosis with more survival neurons after stroke. Moreover, COs transplantation promotes predominantly exogenous neurogenesis in the transplantation periphery of ipsilateral cortex and predominantly endogenous neurogenesis in the hippocampus and subventricular zone. Together, we demonstrate the efficacy and underlying mechanisms of COs transplantation in stroke. This preliminary but promising study provides first-hand preclinical evidence for COs transplantation as a potential and effective intervention for stroke treatment.

Nicotinamide phosphoribosyltransferase (Nampt) is the rate-limiting enzyme in the salvage pathway of nicotinamide adenine dinucleotide (NAD) biosynthesis. Thus far, hepatic Nampt has not been extensively explored in terms of its effects on serum lipid stability and liver lipids metabolism. In this study, hepatocyte-specific Nampt knockout (HC-Nampt-/-) mice were generated by Cre/loxP system. Nampt mRNA expression was reduced in the liver, but not in other tissues, in HC-Nampt-/- mice compared with wild-type (WT) mice. Hepatic Nampt deficiency had no effect on body weight and fasting blood glucose, and it did not induce atherosclerosis in mice under both normal chow diet (NCD) and high fat diet (HFD). At baseline state under NCD, hepatic Nampt deficiency also did not affect liver weight, liver function index, including alanine aminotransferase, aspartate aminotransferase, albumin and alkaline phosphatase, and serum levels of lipids, including triglycerides (TG), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and non-esterified fatty acids (NEFA). However, under HFD, deficiency of hepatic Nampt resulted in increased liver weight, liver function index, and serum levels of TG, TC, HDL-C, and NEFA. Meanwhile, histopathological examination showed increased fat accumulation and fibrosis in the liver of HC-Nampt-/- mice compared with WT mice. Taken together, our results show that hepatic Nampt deficiency aggravates dyslipidemia and liver damage in HFD fed mice. Hepatocyte Nampt can be a protective target against dyslipidemia and fatty liver.

Whether the well-known metabolic switch AMP-activated protein kinase (AMPK) is involved in the insulin-sensitizing effect of calorie restriction (CR) is unclear. In this study, we investigated the role of AMPK in the insulin-sensitizing effect of CR in skeletal muscle. Wild-type (WT) and AMPK-α2(-/-) mice received ad libitum (AL) or CR (8 weeks at 60% of AL) feeding. CR increased the protein level of AMPK-α2 and phosphorylation of AMPK-α2. In WT and AMPK-α2(-/-) mice, CR induced comparable changes of body weight, fat pad weight, serum triglycerides, serum nonesterified fatty acids, and serum leptin levels. However, decreasing levels of fasting/fed insulin and fed glucose were observed in WT mice but not in AMPK-α2(-/-) mice. Moreover, CR-induced improvements of whole-body insulin sensitivity (evidenced by glucose tolerance test/insulin tolerance test assays) and glucose uptake in skeletal muscle tissues were abolished in AMPK-α2(-/-) mice. Furthermore, CR-induced activation of Akt-TBC1D1/TBC1D4 signaling, inhibition of mammalian target of rapamycin-S6K1-insulin receptor substrate-1 pathway, and induction of nicotinamide phosphoribosyltransferase-NAD(+)-sirtuin-1 cascade were remarkably impaired in AMPK-α2(-/-) mice. CR serum increased stability of AMPK-α2 protein via inhibiting the X chromosome-linked ubiquitin-specific protease 9-mediated ubiquitylation of AMPK-α2. Our results suggest that AMPK may be modulated by CR in a ubiquitylation-dependent manner and acts as a chief dictator for the insulin-sensitizing effects of CR in skeletal muscle.

Combinations therapy is often used in hypertensive patients whether combination therapy is necessary for preventing end-organ damage is not known. The objective of this study was to determine in four different hypertensive animal models the necessity of adding the calcium channel blocker amlodipine to therapy with the ss-blocker atenolol to modulate end-organ damage. Spontaneously hypertensive rats, DOCA-salt hypertensive rats, two-kidney, one-clip renovascular hypertensive rats and Lyon genetically hypertensive rats were used to study this objective. These animal models have different sensitivities to atenolol and amlodipine. The dosages of therapy employed were 10 mg/kg atenolol alone, 1 mg/kg amlodipine, 10 mg atenolol + 1 mg/kg amlodipine and 5 mg/kg atenolol+0.5 mg/kg amlodipine. BP was continuously recorded in all animals. After determination of baroreflex sensitivity, rats were sacrificed for end-organ damage evaluation. The combination of amlodipine and atenolol had a synergistic inhibitory effect on blood pressure and blood pressure variability, and end-organ damage as compared with monotherapy with atenolol or amlodipine in all animal models. Baroreflex sensitivity also improved with the combination therapy more than with monotherapy. In conclusion, atenolol and amlodipine combination exerts a superior effect on blood pressure, blood pressure variability, baroreflex sensitivity and end-organ damage. The superior effect of the combination was observed in all four models of hypertension.

Metrnl is a newly identified secreted protein highly expressed in the intestinal epithelium. This study aimed to explore the role and mechanism of intestinal epithelial Metrnl in ulcerative colitis. Metrnl-/- (intestinal epithelial cell-specific Metrnl knockout) mice did not display any phenotypes of colitis under basal conditions. However, under administration of 3% dextran sodium sulfate (DSS) drinking water, colitis was more severe in Metrnl-/- mice than in WT mice, as indicated by comparisons of body weight loss, the presence of occult or gross blood per rectum, stool consistency, shrinkage in the colon, intestinal damage, and serum levels of inflammatory factors. DSS-induced colitis activated autophagy in the colon. This activation was partially inhibited by intestinal epithelial Metrnl deficiency, as indicated by a decrease in Beclin-1 and LC3-II/I and an increase in p62 in DSS-treated Metrnl-/- mice compared with WT mice. These phenomena were further confirmed by observation of autophagosomes and immunofluorescence staining for LC3 in epithelial cells. The autophagy-related AMPK-mTOR-p70S6K pathway was also activated in DSS-induced colitis, and this pathway was partially blocked by intestinal epithelial Metrnl deficiency, as indicated by a decrease in AMPK phosphorylation and an increase in mTOR and p70S6K phosphorylation in DSS-treated Metrnl-/- mice compared with WT mice. Therefore, Metrnl deficiency deteriorated ulcerative colitis at least partially through inhibition of autophagy via the AMPK-mTOR-p70S6K pathway, suggesting that Metrnl is a therapeutic target for ulcerative colitis.

Psychosocial stress is a critical inducing factor of inflammatory bowel diseases (IBD), while autophagy is a novel central issue of IBD development. The present study investigated the potential role of autophagy in stress-related IBD in patients and animal model. The correlation between psychosocial stress and intestinal autophagy was determined in 23 patients with IBD. Corticotropin-releasing hormone (CRH), a well-established inducer of psychosocial stress, was administrated in dextran sulfate sodium (DSS)-induced IBD mice and lipopolysaccharide (LPS)-stimulated bone marrow-derived macrophages (BMDM). In IBD patients, the autophagy markers beclin-1, LC3-II/I ratio, Atg16L1, and Atg4B were significantly enhanced. The psychosocial stress score was positively associated with the levels of beclin-1 and the LC3II/I ratio in intestinal biopsy specimens. In IBD mouse model, CRH significantly aggravated intestinal inflammation, increased Paneth cell metaplasia, and enhanced intestinal autophagy (beclin-1, Atg16L1, PIK3R4, and Atg4B upregulation; GAA, CTSD, and PPKAA1 downregulation). Additionally, the CRH-induced gut microbial dysbiosis was evidenced by a marked increase in the number of detrimental bacteria. In LPS-stimulated BMDM, CRH substantially increased M1/M2 polarization and thus promoted inflammation. In both IBD mice and LPS-treated BMDM, blockade of autophagy by chloroquine abrogated the unbeneficial effects of CRH, whereas autophagy inducer rapamycin resulted in a pronounced protective effect against IBD lesion. Our data demonstrate that psychosocial stress may link the enhanced intestinal autophagy by modulating gut microbiota and inflammation to aggravate IBD. These data indicate autophagy as a promising therapeutic target for psychosocial stress-related IBD.

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 TPβ, a rate-limiting enzyme in FAO. Although TPβ activity is normally inhibited by oxidation under glucose deprivation, the Nur77-TPβ association results in Nur77 self-sacrifice to protect TPβ from oxidation. FAO is therefore able to maintain NADPH and ATP levels and prevent ROS increase and cell death. The Nur77-TPβ 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.

Tissue plasminogen activator (tPA) is the only approved pharmacological therapy for acute brain ischaemia; however, a major limitation of tPA is the haemorrhagic transformation that follows tPA treatment. Here, we determined whether nicotinamide mononucleotide (NMN), a key intermediate of nicotinamide adenine dinucleotide biosynthesis, affects tPA-induced haemorrhagic transformation.

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.

Cerebral organoids (COs) have been used for studying brain development, neural disorders, and species-specific drug pharmacology and toxicology, but the potential of COs transplantation therapy for brain injury remains to be answered.