Salivary concentrations of cytokines and other analytes in healthy children.

BACKGROUND: Blood has been the usual biological fluid for measuring analytes, but there is mounting evidence that saliva may be also useful for detecting cytokines in a noninvasive way. Thus, in this study we aimed to determine concentration of cytokines and other analytes in saliva from a population of healthy children. METHODS: We collected un-stimulated whole saliva samples from clinically healthy children, and concentration of 17 cytokines and 12 other analytes were measured in supernatants. All values were adjusted by albumin content and were log-transformed before multivariate statistical analysis. RESULTS: We included 114 children (53.5% females) between 6.0 and 11.9 years old. The highest concentrations (medians, pg/µg albumin) were seen for visfatin (183.70) and adiponectin (162.26) and the lowest for IL-13 and IL-2 (~0.003). Albumin concentration was associated with age (rS = 0.39, p < 0.001). In the multivariate analysis, five analytes (C peptide, ghrelin, GLP-1, glucagon, leptin) inversely correlated with age and positively with height-for-age. Age was also positively associated with PAI-1, while height-for-age was also positively associated with insulin and visfatin. Finally, BMI-for-age had a positive correlation with GM-CSF and insulin. CONCLUSIONS: Herein, we provided concentration values for 29 analytes in saliva from healthy children that may be useful as preliminary reference framework in the clinical research setting.

Optimization and bioactivity verification of porcine recombinant visfatin with high expression and low endotoxin content using pig liver as template.

In order to obtain the porcine recombinant visfatin protein with high expression and low endotoxin content, the current study aims to express and verify the biological activity of the purified porcine recombinant visfatin protein. Firstly, four different expression strains were successfully constructed. Then they were simultaneously induced at 37 °C for 4 h and 16 °C for 16 h. The results showed that Visfatin-pET28a-Transetta was the best strain with high protein expression and purity at 16 °C induction for 16 h. After that, endotoxin was reduced from the recombinant visfatin until the residual endotoxin was less than one endotoxin units per milliliter (EU/mL). Finally, the purified porcine recombinant visfatin protein was incubated with RAW264.7 cells. The results of cell counting kit-8 (CCK-8) showed the survival rate of the cells first increased and then decreased with the increase in visfatin concentration. When the concentration of visfatin was 700 ng/mL, the survival rate of the cells was the highest. Thereafter, control (PBS), Visfatin and Visfatin + PolymyxinB (Ploy.B) groups were incubated with the RAW264.7 cells for 6 h. Real-time quantitative polymerase chain reaction (RT-qPCR) and Enzyme Linked Immuno-Sorbent Assay (ELISA) results showed that, as compared to the control group, the expressions of interleukin (IL)-1ß, tumor necrosis factor (TNF)-α and monocyte chemoattractant protein (MCP)-1 in Visfatin group were significantly increased (P < 0.05). However, there was no significant difference between the Visfatin and Visfatin + Poly.B groups, indicating that porcine recombinant visfatin protein promoted the inflammatory activity of RAW264.7 cells while the residual endotoxin did not play a role, suggesting biological activity of porcine recombinant visfatin protein.

Filifactor alocis and Tumor Necrosis Factor-Alpha Stimulate Synthesis of Visfatin by Human Macrophages.

There is little known about the effect of the periodontopathogen Filifactor alocis on macrophages as key cells of the innate immune defense in the periodontium. Therefore, the aim of the present study was to investigate the effect of F. alocis and additionally of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNFα) on visfatin and other pro-inflammatory and proteolytic molecules associated with periodontitis in human macrophages. The presence of macrophage markers CD14, CD86, CD68, and CD163 was examined in gingival biopsies from healthy individuals and periodontitis patients. Human macrophages were incubated with F. alocis and TNFα for up to 2 d. The effects of both stimulants on macrophages were determined by real-time PCR, ELISA, immunocytochemistry, and immunofluorescence. F. alocis was able to significantly stimulate the synthesis of visfatin by human macrophages using TLR2 and MAPK pathways. In addition to visfatin, F. alocis was also able to increase the synthesis of cyclooxygenase 2, TNFα, and matrix metalloproteinase 1. Like F. alocis, TNFα was also able to stimulate the production of these proinflammatory and proteolytic molecules. Our results highlight the pathogenetic role of F. alocis in periodontal diseases and also underline the involvement of visfatin in the aetiopathogenesis of periodontitis.

Nicotinamide Phosphoribosyltransferase in Smooth Muscle Cells Maintains Genome Integrity, Resists Aortic Medial Degeneration, and Is Suppressed in Human Thoracic Aortic Aneurysm Disease.

RATIONALE: The thoracic aortic wall can degenerate over time with catastrophic consequences. Vascular smooth muscle cells (SMCs) can resist and repair artery damage, but their capacities decline with age and stress. Recently, cellular production of nicotinamide adenine dinucleotide (NAD+) via nicotinamide phosphoribosyltransferase (Nampt) has emerged as a mediator of cell vitality. However, a role for Nampt in aortic SMCs in vivo is unknown. OBJECTIVES: To determine whether a Nampt-NAD+ control system exists within the aortic media and is required for aortic health. METHODS AND RESULTS: Ascending aortas from patients with dilated aortopathy were immunostained for NAMPT, revealing an inverse relationship between SMC NAMPT content and aortic diameter. To determine whether a Nampt-NAD+ control system in SMCs impacts aortic integrity, mice with Nampt-deficient SMCs were generated. SMC-Nampt knockout mice were viable but with mildly dilated aortas that had a 43% reduction in NAD+ in the media. Infusion of angiotensin II led to aortic medial hemorrhage and dissection. SMCs were not apoptotic but displayed senescence associated-ß-galactosidase activity and upregulated p16, indicating premature senescence. Furthermore, there was evidence for oxidized DNA lesions, double-strand DNA strand breaks, and pronounced susceptibility to single-strand breakage. This was linked to suppressed poly(ADP-ribose) polymerase-1 activity and was reversible on resupplying NAD+ with nicotinamide riboside. Remarkably, we discovered unrepaired DNA strand breaks in SMCs within the human ascending aorta, which were specifically enriched in SMCs with low NAMPT. NAMPT promoter analysis revealed CpG hypermethylation within the dilated human thoracic aorta and in SMCs cultured from these tissues, which inversely correlated with NAMPT expression. CONCLUSIONS: The aortic media depends on an intrinsic NAD+ fueling system to protect against DNA damage and premature SMC senescence, with relevance to human thoracic aortopathy.

Oversecretion and Overexpression of Nicotinamide Phosphoribosyltransferase/Pre-B Colony-Enhancing Factor/Visfatin in Inflammatory Bowel Disease Reflects the Disease Activity, Severity of Inflammatory Response and Hypoxia.

Nicotinamide phosphoribosyltransferase's (Nampt) association with inflammatory bowel disease (IBD) is unclear. The study was aimed at unraveling Nampt's clinical and diagnostic relevance. The serum concentration (Luminex-xMAP® technology) was measured in 113 patients with Crohn's disease (CD), 127 with ulcerative colitis (UC) and 60 non-IBD controls: 40 healthy individuals and 20 with irritable bowel syndrome (IBS). The leukocyte (44 CD/37 UC/19 IBS) and bowel expression (186 samples) was also evaluated (RT-qPCR). All were referred to IBD phenotype, activity, treatment, and inflammatory/nutritional/angiogenic/hypoxia indices. Serum-Nampt and leukocyte-Nampt were positively correlated and were more elevated in active-IBD than in IBS, with leukocyte-Nampt being a fair differential marker. Serum-Nampt in UC positively correlated with its clinical and endoscopic activity as well as with pro-inflammatory cytokines. Serum-Nampt ≤1.54 ng/mL was a good indicator of mucosal healing. The expression of Nampt was up-regulated both in inflamed and quiescent colon and reflected, similarly to leukocyte-Nampt, the clinical activity of IBD. Bowel-Nampt was independently associated with IL1B and hypoxia-inducible factor 1α (HIF1A) expression in inflamed bowel but with FGF2 expression in quiescent bowel. In summary, Nampt's elevation in IBD at local and systemic levels, and protein and mRNA levels, reflects IBD activity and is associated with inflammation, hypoxia (active) and tissue repair (inactive disease).

Determinants of Visfatin/NAMPT Serum Concentration and its Leukocyte Expression in Hyperthyroidism.

We aimed to analyze the potential influence of thyroid autoimmunity on visfatin/NAMPT serum concentration and its leukocyte expression in hyperthyroid patients. This is a single-center, cross-sectional study with consecutive enrollment. All patients with newly diagnosed overt hyperthyroidism in a course of Graves' disease or toxic nodular goiter were included in the study. They underwent physical examination, laboratory investigation, body composition analysis, and thyroid ultrasound. NAMPT mRNA leukocyte expressions were measured using RT-qPCR. Of the 173 patients, 95 were enrolled in further analysis [67 patients with Graves' disease (GD) and 28 with toxic nodular goiter (TNG)]. Control group consisted of 43 healthy volunteers adjusted for age, sex, and BMI. Higher NAMPT/visfatin serum concentration was found in patients with GD comparing with patients with TNG (p=0.03855). We found significant NAMPT leukocyte overexpression in GD patients (n=32) as compared to TNG patients (n=18) and euthyroid controls (n=24) (p=0.005965). Simple linear regression analysis revealed that NAMPT/visfatin serum concentration was significantly associated with NAMPT leukocyte expression, thyroid autoimmunity, age, HOMA-IR, and fat mass percentage (FM%). NAMPT leukocyte expression was related to thyroid autoimmunity, age, and TRAb levels. The stepwise multiple regression analysis revealed FM% and HOMA-IR as independent predictors of visfatin/NAMPT serum levels. In a separate stepwise multiple regression analysis, we confirmed the association between NAMPT leukocyte expression and TRAb levels. We found that fat mass percentage together with HOMA-IR are the most significant predictors of visfatin/NAMPT serum elevation in hyperthyroid patients.

Omega-3 polyunsaturated fatty acids suppress the inflammatory responses of lipopolysaccharide-stimulated mouse microglia by activating SIRT1 pathways.

Obesity and diabetes are known risk factors for dementia, and it is speculated that chronic neuroinflammation contributes to this increased risk. Microglia are brain-resident immune cells modulating the neuroinflammatory state. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), the major ω-3 polyunsaturated fatty acids (PUFAs) of fish oil, exhibit various effects, which include shifting microglia to the anti-inflammatory phenotype. To identify the molecular mechanisms involved, we examined the impact of EPA, DHA, and EPA+DHA on the lipopolysaccharide (LPS)-induced cytokine profiles and the associated signaling pathways in the mouse microglial line MG6. Both EPA and DHA suppressed the production of the pro-inflammatory cytokines TNF-α and IL-6 by LPS-stimulated MG6 cells, and this was also observed in LPS-stimulated BV-2 cells, the other microglial line. Moreover, the EPA+DHA mixture activated SIRT1 signaling by enhancing mRNA level of nicotinamide phosphoribosyltransferase (NAMPT), cellular NAD+ level, SIRT1 protein deacetylase activity, and SIRT1 mRNA levels in LPS-stimulated MG6. EPA+DHA also inhibited phosphorylation of the stress-associated transcription factor NF-κB subunit p65 at Ser536, which is known to enhance NF-κB nuclear translocation and transcriptional activity, including cytokine gene activation. Further, EPA+DHA increased the LC3-II/LC3-I ratio, an indicator of autophagy. Suppression of TNF-α and IL-6 production, inhibition of p65 phosphorylation, and autophagy induction were abrogated by a SIRT1 inhibitor. On the other hand, NAMPT inhibition reversed TNF-α suppression but not IL-6 suppression. Accordingly, these ω-3 PUFAs may suppress neuroinflammation through SIRT1-mediated inhibition of the microglial NF-κB stress response and ensue pro-inflammatory cytokine release, which is implicated in NAMPT-related and -unrelated pathways.

Inhibition of nicotinamide phosphoribosyltransferase and depletion of nicotinamide adenine dinucleotide contribute to arsenic trioxide suppression of oral squamous cell carcinoma.

Emerging evidence suggests that increased nicotinamide phosphoribosyltransferase (NAMPT) expression is associated with the development and prognosis of many cancers, but it remains unknown regarding its role in oral squamous cell carcinoma (OSCC). In the present study, the results from tissue microarray showed that NAMPT was overexpressed in OSCC patients and its expression level was directly correlated with differential grades of cancer. Interestingly, treatment of OSCC cells with chemotherapy agent arsenic trioxide (ATO) decreased the levels of NAMPT protein and increased cellular death in an ATO dose- and time-dependent manner. Most importantly, combination of low concentration ATO with FK866 (a NAMPT inhibitor) exerted enhanced inhibitive effect on NAMPT protein and mRNA expressions, leading to synergistic cytotoxicity on cancer cells through increasing cell apoptosis and depleting intracellular nicotinamide adenine dinucleotide levels. These findings demonstrate the crucial role of NAMPT in the prognosis of OSCC and reveal inhibition of NAMPT as a novel mechanism of ATO in suppressing cancer cell growth. Our results suggest that ATO can significantly enhance therapeutic efficacy of NAMPT inhibitor, and combined treatment may be a novel and effective therapeutic strategy for OSCC patients.

CircRNA hsa_circ_0005105 upregulates NAMPT expression and promotes chondrocyte extracellular matrix degradation by sponging miR-26a.

Osteoarthritis (OA) is a chronic disease pathologically characterized by articular cartilage degeneration and damage. Currently, studies have found that circular RNA (circRNA) is involved in intracellular RNA regulating network and is closely related to the occurrence and development of diseases, therefore it may become a new biological marker and therapeutic target. After stimulating chondrocytes with interleukin-1 beta (IL-1ß), hsa_circ_0005105 expression was significantly upregulated, while miR-26a expression was significantly inhibited. Hsa_circ_0005105 did not influence miR-26a expression but inhibited its transcriptional activity so as to upregulate the expression of its target NAMPT. Studies further indicated that hsa_circ_0005105 can inhibit the expression of type II collagen and aggrecan, promote the expression of MMP-13 and ADAMTS-4, and the generation of PGE2, IL-6, and IL-8, but the linear sequence of hsa_circ_0005105 cannot. MiR-26a has the opposite effect, and hsa_circ_0005105 can antagonize the function of miR-26a. When NAMPT expression was downregulated, the above function of hsa_circ_0005105 was significantly weakened. Therefore, hsa_circ_0005105 can promote extracellular matrix (ECM) degradation by regulating the expression of miR-26a target NAMPT. These findings will provide new targets for treatment and prevention of OA and other orthopedic diseases.

Association of serum microRNA-21 levels with Visfatin, inflammation, and acute coronary syndromes.

MicroRNAs (miRNAs) are short non-coding RNAs that regulate gene expression. It seems that microRNA-21 (miR-21) and Visfatin, a novel adipocytokine, play roles in inflammation and atherosclerosis. The aim of this study was to investigate the association of miR-21 with Visfatin, inflammation, atherosclerosis and acute coronary syndrome (ACS). Based on coronary angiography and electrocardiogram (ECG), 53 patients with ACS and 52 patients with stable CAD were enrolled in this study. We assayed serum miR-21, Visfatin, and routine chemistries using quantitative reverse transcriptase polymerase chain reaction (QRT-PCR), enzyme-linked immunosorbent assay (ELISA) and automated analyzer, respectively. We used a regression analysis to describe the relationship between the variables. Serum miR-21 level in 2-ΔCt value was significantly higher in ACS patients (10.52 ± 1.01-fold) than the stable CAD patients (4.4 ± 0.79-fold) (F = 4.59, p < 0.001). In addition, serum Visfatin was significantly higher in ACS patients (17.5 ± 0.61 ng/ml) than the stable CAD patients (12.7 ± 0.49 ng/ml) (F = 2.62, p < 0.001). Furthermore, the serum miR-21 level correlated positively with serum Visfatin level (r = 0.26, p = 0.008), hs-CRP (r = 0.29, p = 0.003), age (r = 0.21, p = 0.034) and negatively with HDL-cholesterol (r = -0.28, p = 0.004). We concluded that the increased serum miR-21 and Visfatin may be involved in the pathogenesis of ACS through promoting inflammation or may result from inflammatory responses to ACS. Furthermore, the potential role of miR-21 and Visfatin in plaque instability and inflammation warrants further investigations.

Decreased immunoreactivity of visfatin in the pancreas and liver of rats with renovascular hypertension.

Hypertension is one of the major endocrine and metabolic disorders, in which visfatin plays a significant role. The objective of this study was to evaluate the immunoreactivity of visfatin in pancreas and liver of “two kidney, one clip” (2K1C) renovascular hypertension model in rats. The studies were carried out on the pancreas and liver of rats. After a 6-week period of the renal artery clipping procedure, 2K1C rats developed a stable hypertension. Paraffin sections were stained with hematoxylin and eosin (for general histological examination) and processed for immunolocalization of visfatin. The intensity of immunohistochemical reaction was measured using Nikon NIS-Elements Advanced Research software. The hypertension significantly weakened the immunohistochemical reaction exhibiting visfatin in the pancreas and liver of hypertensive rats, compared to control animals. The changes induced by hypertension in the visfatin-containing cells in the pancreas and liver of the rats are discussed and needs further study.

Correlation between polymorphisms in the visfatin gene and its expression in the serum and coronary artery calcification.

We investigated the association between serum visfatin levels and single nucleotide polymorphisms (SNPs; rs61330082, rs2058539) in the visfatin gene and coronary artery calcification (CAC) in patients from Wenzhou, China. CAC patients (N = 206) were divided into two groups: mild CAC (MCAC) and moderate and severe CAC (MSCAC). Volunteers without CAC (N = 70) were included in the control group. The serum visfatin level was analyzed by enzyme-linked immunosorbent assay. SNPs (rs61330082, rs2058539) in the visfatin gene were analyzed by polymerase chain reaction-restriction fragment length polymorphism. Clinical data, serum visfatin levels, and genotype and allele frequencies of rs61330082 and rs2058539 were compared among the three groups. MSCAC patients expressed significantly higher serum visfatin levels (30.58 ± 6.12 ng/mL) than individuals in the MCAC (29.03 ± 1.87 ng/mL) and control (24.45 ± 5.44 ng/mL) groups (P < 0.05). The genotype distributions and frequencies of rs61330082 differed significantly among the groups (P < 0.05), while those of rs2058539 did not. The serum visfatin level was positively correlated with the body mass index (BMI), high-density lipoprotein cholesterol (HDL-C), and insulin resistance index (IRI), and negatively correlated with the triglyceride (TG) levels (P < 0.05) of patients. Serum visfatin is associated with the development of CAC. The T allele of the rs61330082 SNP in the visfatin gene had a cardioprotective effect on patients with CAC; the SNP at rs2058539 was not significantly associated with CAC. The BMI, HDL-C, IRI, and TG levels influenced the development of CAC.

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.

Nicotinamide phosphoribosyltransferase/pre-B-cell colony enhancing factor/visfatin plasma levels and clinical outcome in patients with dilated cardiomyopathy.

BACKGROUND: Nicotinamide phosphoribosyltransferase (Nampt) is an enzyme involved in nicotinamide adenine dinucleotide biosynthesis. Nampt functions as gatekeeper of energy status and survival in cardiac myocytes in animal models of ischemia-reperfusion and might regulate inflammatory processes. Therefore, we performed for the 1st time a clinical study to determine the effects of Nampt on cardiac function in patients with nonischemic dilated (DCM) and inflammatory (DCMi) cardiomyopathy. METHODS AND RESULTS: A total of 113 patients were enrolled in the study and classified into control (n = 25), DCM (n = 38), and DCMi (n = 50) groups. Cardiac functional and inflammatory parameters as well as plasma Nampt and cardiac mRNA and protein Nampt expression were determined at baseline and follow-up after 6 months. Patients with DCM (1.04 ± 0.8 ng/mL; P < .001) and DCMi (1.07 ± 0.7 ng/mL; P < .001) showed significantly increased Nampt plasma concentrations at baseline compared with the control group (0.57 ± 0.5 ng/mL). Patients with higher Nampt concentrations in both heart failure groups showed significant better improvement of cardiac functional parameters (correlation between Nampt plasma levels and the change of left ventricular ejection fraction after 6 months: DCM: r = 0.698, P < .001; DCMi: r = 0.503, P < .001). Moreover, cardiac inflammation did not influence Nampt expression, and Nampt concentrations did not modulate cardiac inflammation in DCMi. A multivariate linear regression model revealed high plasma Nampt expression to contribute to better improvement of cardiac function in patients of both heart failure groups. Moreover, heart failure patients with high plasma Nampt levels showed suppressed cardiac TNF-α and IL-6 mRNA expression after 6 months' follow-up as well as lower B-type natriuretic peptide levels compared with heart failure patients with low Nampt plasma concentrations. CONCLUSIONS: High Nampt expression in patients with nonischemic DCM and DCMi is associated with a favorable outcome and improvement in functional status.

Visfatin/pre-B-cell colony-enhancing factor (PBEF), a proinflammatory and cell motility-changing factor in rheumatoid arthritis.

Adipokines such as adiponectin and visfatin/pre-B-cell colony-enhancing factor (PBEF) have been recently shown to contribute to synovial inflammation in rheumatoid arthritis (RA). In this study, we evaluated the pathophysiological implication of visfatin/PBEF in the molecular patterns of RA synovial tissue, focusing on RA synovial fibroblasts (RASFs), key players in RA synovium. Expression of visfatin/PBEF in synovial fluid and tissue of RA patients was detected by immunoassays and immunohistochemistry. RASFs were stimulated with different concentrations of visfatin/PBEF over varying time intervals, and changes in gene expression were evaluated at the RNA and protein levels using Affymetrix array, real-time PCR, and immunoassays. The signaling pathways involved were identified. The influence of visfatin/PBEF on fibroblast motility and migration was analyzed. In RA synovium, visfatin/PBEF was predominantly expressed in the lining layer, lymphoid aggregates, and interstitial vessels. In RASFs, visfatin/PBEF induced high amounts of chemokines such as IL-8 and MCP-1, proinflammatory cytokines such as IL-6, and matrix metalloproteinases such as MMP-3. Phosphorylation of p38 MAPK was observed after visfatin/PBEF stimulation, and inhibition of p38 MAPK showed strong reduction of visfatin-induced effects. Directed as well as general fibroblast motility was increased by visfatin/PBEF-induced factors. The results of this study indicate that visfatin/PBEF is involved in synovial fibroblast activation by triggering fibroblast motility and promoting cytokine synthesis at central sites in RA synovium.

Nampt expression increases during osteogenic differentiation of multi- and omnipotent progenitors.

Despite emerging data showing that metabolic changes occur with stem cell differentiation, the cross-talk between factors governing energy metabolism and epigenetic modification is not understood. Nicotinamide adenine dinucleotide (NAD) participates in both energy metabolism and protein modification processes. Changes of the intracellular NAD concentration have been shown to correlate with differentiation of adult and embryonic stem cells. In the present study, we investigated the expression pattern of Nampt, the rate-limiting enzyme in NAD salvaging pathway, during osteogenic differentiation of the multipotent mouse fibroblast C3H10T1/2 and the omnipotent preosteoblast MC3T3-E1 cells. We found that Nampt was increasingly expressed during differentiation in both cell models. The increase of Nampt was associated with higher NAD concentration and Sirt1 activity. Knockdown of Nampt or addition of its specific inhibitor FK866 leads to lower intracellular NAD concentration and decline in osteogenesis. These findings indicate that osteogenic differentiation correlates with intracellular NAD metabolism in which Nampt plays a regulatory role.

Visfatin, a potential biomarker and prognostic factor for endometrial cancer.

OBJECTIVE: Visfatin, a newly discovered adipocytokine, is thought to play a role in the pathogenesis of metabolic-syndrome-related cancers. The aim of this study was to assess the clinical significance of serum levels and tissue expression of visfatin in relation to endometrial cancer (EC). METHODS: A total of 234 EC patients were included in this study. Serum visfatin, metabolic and anthropometric parameters were measured in EC patients and controls. Serum visfatin levels were detected using ELISA. Tissue expression of visfatin was analyzed using immunohistochemistry in tissue microarrays. The correlation between clinicopathological variables and visfatin in EC tissues and the prognostic value of visfatin for overall survival was evaluated. RESULTS: Serum levels of visfatin were significantly higher in EC patients than in controls (P<0.05). In univariate and multivariate logistic regression models, a positive association between EC and serum visfatin, BMI, waist-to-hip ratio, diabetes, and hypertension was evident (P<0.05). Visfatin expression was significantly higher in EC tissue than in normal endometrial tissue (P=0.001). Moreover, serum visfatin levels were significantly positively correlated with tissue expression of visfatin in EC patients (P<0.05). High visfatin expression in EC tissues was significantly associated with advanced FIGO stage (P=0.016) and myometrial invasion ≥1/2 (P=0.023). The overall survival rate of EC patients was significantly higher in the group with negative visfatin expression than with positive visfatin expression (P=0.035). CONCLUSIONS: Visfatin is a potential serum biomarker and prognostic factor for EC that may indicate high risk for EC and EC progression. It may also be a novel potential therapeutic target for EC.

A key role for Pre-B cell colony-enhancing factor in experimental hepatitis.

UNLABELLED: Pre-B cell colony-enhancing factor (PBEF), also known as nicotinamide phosphoribosyltransferase or visfatin, plays an important role in metabolic, inflammatory, and malignant diseases. Recent evidence suggests that blocking its enzymatic activity using a specific small-molecule inhibitor (FK866) might be beneficial in acute experimental inflammation. We investigated the role of PBEF in human liver disease and experimental hepatitis. PBEF serum levels and hepatic expression were determined in patients with chronic liver diseases. These studies were followed by in vivo experiments using concanavalin A (ConA) and D-galactosamine/lipopolysaccharide (LPS) models of experimental hepatitis. PBEF was either overexpressed by hydrodynamic perfusion or inhibited by FK866. In vivo findings were corroborated studying inflammatory responses of lentivirally PBEF-silenced or control FL83B mouse hepatocytes. Here, we demonstrate that PBEF serum levels were increased in patients with chronic liver diseases irrespective of disease stage and etiology. In particular, we observed enhanced PBEF expression in hepatocytes. Liver-targeted overexpression of PBEF rendered mice more susceptible to ConA- and D-galactosamine/LPS-induced hepatitis compared with control animals. In contrast, inhibition of PBEF using FK866 protected mice from ConA-induced liver damage and apoptosis. Administration of FK866 resulted in depletion of liver nicotinamide adenine dinucleotide+ levels and reduced proinflammatory cytokine expression. Additionally, FK866 protected mice in the D-galactosamine/LPS model of acute hepatitis. In vitro, PBEF-silenced mouse hepatocytes showed decreased responses after stimulation with LPS, lipoteichoic acid, and tumor necrosis factor α. In primary murine Kupffer cells, FK866 suppressed LPS-induced interleukin (IL)-6 production, whereas incubation with recombinant PBEF resulted in increased IL-6 release. CONCLUSION: Our data suggest that PBEF is of key importance in experimental hepatitis. Its specific inhibition might be considered a novel treatment option for inflammatory liver diseases.

Expression of visfatin mRNA in peripheral blood mononuclear cells is not correlated with visfatin mRNA in omental adipose tissue in women with polycystic ovary syndrome.

BACKGROUND: Visfatin, which is secreted predominantly from visceral adipose tissue, has an insulin-mimetic action and may play a role in the regulation of insulin sensitivity in humans. Peripheral blood mononuclear cells (PBMCs) from venous blood samples are the most accessible tissue for the analysis of gene expression. The aims of the study were to compare the expression of visfatin in PBMCs with that in omental adipose tissue in women with polycystic ovary syndrome (PCOS). METHODS: Visfatin mRNA was measured in omental adipose tissue and PBMCs from 10 women with PCOS and 10 healthy controls, matched for BMI and age, using the real-time polymerase chain reaction (PCR). RESULTS: The expression of visfatin mRNA in both omental adipose tissue and PBMCs from the women with PCOS was significantly higher (P = 0.01 and P = 0.05, respectively) than that in the controls. This finding indicated that mononuclear cells are a potential source of visfatin in women with PCOS. However, only the expression of visfatin mRNA in adipose tissue, not that in PBMCs, showed a significant positive correlation with insulin levels 2h after glucose loading (P = 0.044, r(2) = 0.45), and with homeostasis model assessment-insulin resistance (HOMA(IR); P = 0.035, r(2) = 0.47). In addition, the expression of visfatin mRNA in PBMCs did not correlate with the expression of visfatin mRNA in omental adipose tissue. CONCLUSIONS: PCOS is associated with increased visfatin mRNA concentrations in PBMCs and in omental adipose tissue. However, only visfatin mRNA concentration in omental adipose tissue is closely correlated with BMI and insulin resistance.

Analysis of an Indian diabetes prevention programme on association of adipokines and a hepatokine with incident diabetes.

To study the association and possible predictive role of visfatin, resistin, fetuin-A and chemerin with incident type 2 diabetes (T2DM) among Asian Indians with prediabetes. Their association with insulin resistance, ß-cell function, glycaemia and anthropometry were also studied. This is a nested case-control study of a large 2-year prospective prevention trial in persons at high risk of developing T2DM. Baseline HbA1c values between 6.0% (42 mmol/mol) and 6.2% (44 mmol/mol) were chosen for this analysis (n = 144). At follow-up, persons with incident T2DM (HbA1c ≥ 6.5%, 48 mmol/mol) were grouped as cases (n = 72) and those reverted to normoglycaemia, (HbA1c < 5.7% (39 mmol/mol) as controls (n = 72). Insulin resistance showed the strongest association with incident T2DM ((Odds Ratio (OR): 23.22 [95%CI 6.36-84.77]; p < 0.0001). Baseline visfatin (OR: 6.56 [95%CI 2.21-19.5]; p < 0.001) and fetuin-A (OR: 1.01 [95%CI (1.01-1.04)]; p < 0.0001) independently contributed to the conversion of prediabetes to T2DM. The contribution was significantly higher when their elevated levels coexisted (OR: 12.63 [95%CI 3.57-44.63]; p < 0.0001). The area under the curve was 0.77 ± SE 0.4 (95%CI 0.69-0.85) and 0.80 ± SE 0.04 (95%CI 0.73-0.88) for visfatin (median 17.7 ng/ml, sensitivity and specificity: 75%, p < 0.0001) and fetuin-A (mean 236.2 µg/ml, sensitivity: 71%, specificity: 75%, p < 0.0001) respectively. Higher baseline visfatin and fetuin-A concentrations are strongly associated with incident T2DM and are predictive of future diabetes.