pre-B Cell Colony-enhancing Factor Research Articles

Association of Visfatin gene polymorphism with obesity related metabolic disorders among Pakistani population: a case control study

In recent years, the global prevalence of obesity and its associated metabolic disorders has reached alarming levels, presenting a significant challenge to public health worldwide. Visfatin, also known as pre-B cell colony-enhancing factor (PBEF) or nicotinamide phosphoribosyltransferase (NAMPT), is an adipokine that has been implicated in various physiological processes, including glucose homeostasis, lipid metabolism, and inflammation. The main objective of this proposed study is to find out the association between visfatin genetic variants and metabolic syndrome. The sample size of the study consisted of 300 blood samples (150 control and 150 cases). This study found that the genotypic frequency of visfatin SNPs, including rs2302559 (OD: 18.222; 95% CI 10.228–32.466; p-value < 0.001) and rs1215113036 (OD: 129.40; 95% CI 44.576–375.693; p-value < 0.001) were significantly associated with metabolic syndrome. Moreover, the frequency of the mutant alleles of both visfatin SNPs was found to be higher in patients with metabolic syndrome as compared to controls. Results of the current study indicate that people with any genetic variation of Visfatin, such as rs2302559 and rs1215113036, are more likely to develop metabolic syndrome. Visfatin genetic variants are linked to an increased risk of metabolic syndrome, implying it’s role in disease pathophysiology.

Updated Functional Roles of NAMPT in Carcinogenesis and Therapeutic Niches.

Simple SummaryThe advantages and applications of using the non-invasive way to detect serum biomarkers for assessing cancer diagnosis and prognosis have been explored. Nicotinamide phosphoribosyltransferase (NAMPT), also designated as pre-B-cell colony-enhancing factor (PBEF) or visfatin, is a secreted adipokine known to modulate tumor malignancies. Its significance in predicting cancer patient’s survival outcome further renders the implementation of NAMPT in clinical practice. In this review, recent discoveries of NAMPT in cancer studies were focused and integrated. We aim to provide updates for researchers who are proposing relevant objectives.Nicotinamide phosphoribosyltransferase (NAMPT) is notable for its regulatory roles in tumor development and progression. Emerging evidence regarding NAMPT somatic mutations in cancer patients, NAMPT expressional signatures in normal tissues and cancers, and the prognostic significance of NAMPT in many cancer types has attracted attention, and NAMPT is considered a potential biomarker of cancer. Recent discoveries have demonstrated the indirect association and direct biological functions of NAMPT in modulating cancer metastasis, proliferation, angiogenesis, cancer stemness, and chemoresistance to anticancer drugs. These findings warrant further investigation of the underlying mechanisms to provide knowledge for developing novel cancer therapeutics. In this review article, we explore recent research developments involving the oncogenic activities of NAMPT by summarizing current knowledge regarding NAMPT somatic mutations, clinical trials, transcriptome data, and clinical information and discoveries related to the NAMPT-induced signaling pathway in modulating hallmarks of cancer. Furthermore, the comprehensive representation of NAMPT RNA expression in a pancancer panel as well as in specific normal cell types at single-cell level are demonstrated. The results suggest potential sites and cell types that could facilitate NAMPT-related tumorigenesis. With this review, we aim to shed light on the regulatory roles of NAMPT in tumor development and progression, and provide information to guide future research directions in this field.

Description of Visfatin Adipokine and its Roles on Inflammation and Coronary Heart Disease

Coronary heart disease (CHD) is a generic designation for a group of related syndromes resulting from myocardial ischemia – an imbalance between cardiac blood supply (perfusion) and myocardial oxygen demand. Visfatin (VF) is a recently discovered adipokine with different functions, Visfatin is mainly found in visceral adipose tissue and mimics insulin in lowering plasma glucose levels and, Visfatin emerges as a player in the development and progression of atherosclerotic lesions by directly promoting smooth muscle cell proliferation, Aberrant angiogenesis is now considered a feature of the atherogenic process in both coronary and carotid diseases. This adipokine was previously known as a pre-B-cell colony-enhancing factor (PBEF) or Nicotinamide phosphoribosyltransferase (NAmPRTase or Nampt) an enzyme that in humans is encoded by the NAMPT gene and demonstrated to be an intracellular protein with a key enzyme role in nicotinamide adenine dinucleotide (NAD)

Inhibition of Pre-B Cell Colony Enhancing Factor Reduces Lung Injury in Rats Receiving Cardiopulmonary Bypass.

ObjectivePre-B cell colony enhancing factor (PBEF) is an important proinflammatory cytokine involved in acute lung injury. However, whether PBEF participates in lung injury caused by cardiopulmonary bypass (CPB) is still unknown. This study aimed to investigate the effects of silencing PBEF on lung injury and the sodium and water transport system in rats receiving CPB.MethodsMorphological changes in lung tissues were evaluated using hematoxylin and eosin (H&E) staining. PBEF was detected using immunohistochemistry. The sodium and water transport system-related proteins and cellular signaling pathways were detected by Western blotting.ResultsRats receiving CPB (model group) had more severe alveolar wall damage and higher expression of PBEF in free form than the control rats. Western blotting showed that the expression of PBEF, surfactant protein D (SP), aquaporin (AQP) 1, AQP5, and epithelial sodium channel (ENaC) was significantly higher in the lung tissue of CPB rats than control rats. By contrast, adenovirus-encoding sh-PBEF significantly reduced the expression of PBEF, SP, AQP1, AQP5, and ENaC in the lung tissues of rats treated with CPB. The phosphorylation levels of extracellular signal-regulated protein kinases 1 and 2 (ERK1/2), protein kinase B (AKT), and p38 mitogen-activated protein kinase (MAPK) were significantly increased in the lung tissue of rats that received CPB, and were downregulated by adenovirus-encoding sh-PBEF.ConclusionAdenovirus-encoding sh-PBEF could reduce lung injury and repair the sodium–water transport system in rats receiving CPB, likely through reducing MAPK, ERK1/2, and Akt signaling pathways.

The Role of Pre-B-cell Colony- Enhancing Factor in Egyptian Children with Hemophagocytic Lymphohistiocytosis

Abstract Background Hemophagocytic lymphohistiocytosis (HLH) caused by a highly active but ineffective immune response, including impaired or absent function of natural killer cells and cytotoxic T cells, and the release of pro-inflammatory cytokines. Pre-B-cell colony-enhancing factor (PBEF) is an inflammatory cytokine involved in several inflammatory diseases and it has been identified to react with several cytokines involved in HLH. Objective we aimed to evaluate the role of PBEF as a diagnostic and prognostic marker in Egyptian children with HLH. Subjects and Methods This is a cross-sectional study, recruited fifteen children with HLH from the Pediatric Hematology/Oncology unit. Patients underwent thorough clinical assessment lying stress on disease manifestation, classification, treatment and prognosis. Plasma concentration of PBEF was determined using an enzyme-linked immunosorbent assay. Results PBEF was significantly increase in patients group than control group. Four patients were classified as primary HLH, seven patients were as secondary HLH and four patients had unknown classification due to waiting for genotyping. Seven patients of the study group were died and eight patients still alive. PBEF level showed a significant positive correlation with serum ferritin and triglycerides level and negative correlation with fibrinogen level. Conclusion The elevated PBEF level and its correlation with the widely available biochemical markers for diagnosis of pediatric HLH indicating that it may be involved in its inflammatory process.

Relationship between NAMPT/PBEF/visfatin and prognosis of patients with malignant tumors: a systematic review and meta-analysis.

Nicotinamide phosphoribosyltransferase (NAMPT), also known as pre-B-cell colony-enhancing factor (PBEF) or visfatin, has been reported to be a crucial factor involved in tumor metabolism, angiogenesis and cell apoptosis. However, its definite roles in patients with malignant cancer remain unclear. Three online databases PubMed, Embase and Web of Science were looked through comprehensively for eligible articles, published before November, 2018. Pooled hazard ratios (HRs) with 95% confidence intervals (CIs) of overall survival (OS) or disease-free survival time or recurrence-free survival (DFS/RFS) were calculated to determine the associations between NAMPT expression and cancer prognosis. A total of ten eligible studies were finally enrolled for this analysis. Our results indicated that elevated NAMPT expression was associated with poor OS in breast cancer by both univariate and multivariate analysis (pooled HR =3.23, 95% CI: 1.93-5.41, I2=21.1%, P=0.283; pooled HR =3.34, 95% CI: 2.13-5.22, I2=0.0%, P=0.791; respectively) and in gastric cancer by univariate analysis (pooled HR =2.47, 95% CI: 1.07-5.73, I2=91.1%, P=0.001). Moreover, high expression of NAMPT was also related to poor DFS/RFS in breast cancer by univariate and multivariate analysis (pooled HR =3.85, 95% CI: 2.59-5.71, I2=0.0%, P=0.700; pooled HR =3.43, 95% CI: 2.36-4.99, I2=0.0%, P=0.737; separately). Similar results could be found in urothelial carcinoma (pooled HR =3.14, 95% CI: 1.73-5.71, I2=47.8%, P=0.166; pooled HR =3.06, 95% CI: 1.57-5.98, I2=0.0%, P=0.860). Besides, the translational level of NAMPT was also validated by UALCAN and the Human Protein Atlas database [immunohistochemistry (IHC)]. Our results shed light on that NAMPT might be an oncogenic factor in breast cancer, gastric cancer and urothelial carcinoma.

Akt1-mediated CPR cooling protection targets regulators of metabolism, inflammation and contractile function in mouse cardiac arrest.

Therapeutic hypothermia initiated during cardiopulmonary resuscitation (CPR) in pre-clinical studies appears to be highly protective against sudden cardiac arrest injury. Given the challenges to implementing CPR cooling clinically, insights into its critical mechanisms of protection could guide development of new CPR drugs that mimic hypothermia effects without the need for physical cooling. Here, we used Akt1-deficient mice that lose CPR hypothermia protection to identify hypothermia targets. Adult female C57BL/6 mice (Akt1+/+ and Akt1+/-) underwent 8 min of KCl-induced asystolic arrest and were randomized to receive hypothermia (30 ± 0.5°C) or normothermia. Hypothermia was initiated during CPR and extended for 1 h after resuscitation. Neurologically scored survival was measured at 72 h. Other outcomes included mean arterial pressure and target measures in heart and brain related to contractile function, glucose utilization and inflammation. Compared to northothermia, hypothermia improved both 2h mean arterial pressure and 72h neurologically intact survival in Akt1+/+ mice but not in Akt1+/- mice. In Akt1+/+ mice, hypothermia increased Akt and GSK3β phosphorylation, pyruvate dehydrogenase activation, and NAD+ and ATP production while decreasing IκBα degradation and NF-κB activity in both heart and brain at 30 min after CPR. It also increased phospholamban phosphorylation in heart tissue. Further, hypothermia reduced metabolic and inflammatory blood markers lactate and Pre-B cell Colony Enhancing Factor. Despite hypothermia treatment, all these effects were reversed in Akt1+/- mice. Taken together, drugs that target Akt1 and its effectors may have the potential to mimic hypothermia-like protection to improve sudden cardiac arrest survival when administered during CPR.

Visfatin and Its Role in the Regulation of the Reproductive System

Visfatin, also called the pre-B cell colony enhancing factor, is produced mainly by the visceral fat and in the target cells activates the 3-phosphoinositide pathway, the cascade of mitogen-activated protein kinases and a number of the other effector systems. In the recent years, evidence has been obtained that visfatin regulates the functional activity of the hypothalamic-pituitary-gonad axis. Visfatin controls the synthesis and secretion of both gonadoliberin by the hypothalamic neurons and gonadotropins by the pituitary gonadotrophs, and also affects the steroidogenesis and folliculogenesis, acting on the reproductive tissues. One of the mechanisms of the steroidogenic action of visfatin is the potentiation of the stimulating effect of insulin-like growth factor-1 on the synthesis of sex steroid hormones. Since the production of visfatin and the activity of its signaling system vary significantly in the conditions of diabetes mellitus and metabolic syndrome, this directly affects its regulation of the reproductive functions in these metabolic disorders. Visfatin plays an important role in maintaining the functional activity of the ovaries and the normal quality of oocytes during aging. The review is devoted to the problem of the structural and functional organization of visfatin and its signaling system, the role of visfatin in the regulation of the female and male reproductive systems, and the mechanisms of visfatin action on the hypothalamic-pituitary-gonad axis.

The role of visfatin in cancer proliferation, angiogenesis, metastasis, drug resistance and clinical prognosis.

Visfatin, also known as nicotinamide phosphoribosyltransferase or pre-B-cell colony-enhancing factor (PBEF), is an adipocytokine secreted by adipocytes, macrophages and inflamed endothelial tissue. Related reports have indicated a positive correlation between the visfatin level and obesity and cancer risk. In addition to its original function, visfatin is multifunctional and plays critical roles in the promotion of several processes relevant to cancer progression including cancer cell proliferation, angiogenesis, metastasis and drug resistance. The relative expression of visfatin and the potential visfatin receptor on a pan-cancer scale was determined based on the transcriptome analysis data in The Cancer Genome Atlas. We further show the clinical association of its signaling axis with the survival of cancer patients, which reveals its prognostic power for specific cancer types. This review illustrates visfatin’s biological functions related to cancer progression and demonstrates its clinical significance in predicting outcomes of cancer patients.

Visfatin Serum Levels Predict Mortality in Critically Ill Patients.

The adipokine visfatin, also termed pre-B-cell colony-enhancing factor (PBEF), is mainly derived from adipose tissue but has been implicated in the regulation of innate immune responses. We hypothesized that visfatin could be a potential circulating biomarker in critical illness and sepsis. We therefore measured serum levels of visfatin in a cohort of 229 critically ill medical patients upon admission to the intensive care unit (ICU). In comparison to 53 healthy controls, visfatin levels were significantly elevated in medical ICU patients, especially in patients with sepsis. Visfatin serum concentrations were strongly associated with disease severity and organ failure but did not differ between patients with or without obesity or type 2 diabetes. Visfatin levels correlated with biomarkers of renal failure, liver dysfunction, and other adipokines (e.g., resistin, leptin, and adiponectin) in critically ill patients. High visfatin levels at ICU admission indicated an increased mortality, both at the ICU and during long-term follow-up of approximately two years. Our data therefore demonstrate that circulating visfatin is a valuable biomarker for risk and prognosis assessment in critically ill patients. Furthermore, visfatin seems to be involved in the pathogenesis of excessive systemic inflammation, supporting further research on visfatin as a therapeutic target.

Clinical Significance of Serum Visfatin in Renal Transplant Recipients

Chronic antibody-mediated rejection is the most common cause of late graft loss in renal transplant recipients. Visfatin is a pre-B cell colony-enhancing factor secreted by activated lymphocytes. We hypothesize that visfatin may play a role in the augmentation of B cell colonies and facilitate antibody-mediated rejection. Renal transplant recipients were randomly selected for the study. Fasting blood samples were obtained for the assay of visfatin. The participants were prospectively followed up for 3 years. A total of 146 patients were recruited for the study and were divided into 3 groups according to tertile of serum visfatin level. At the end of follow-up, 6 patients had graft loss, including 1 graft loss in tertile 1, 3 in tertile 2, and 2 in tertile 3 (P = .60). Fourteen patients experienced at least 1 episode of acute rejection, while 21 patients were diagnosed as having chronic rejection. The distribution of acute rejection was 10.2% in tertile 1, 10.2% in tertile 2, and 8.3% in tertile 3 (P = .94); chronic rejection occurred in 10.2%, 16.3%, and 16.7%, respectively (P = .59). We conclude that serum visfatin level was not correlated with either graft failure or patient mortality in a 3-year observation period.

The role of pre-B-cell colony-enhancing factor in Egyptian children with hemophagocytic lymphohistiocytosis

Background Hemophagocytic lymphohistiocytosis (HLH) is caused by a highly active but ineffective immune response, including impaired or absent function of natural killer cells and cytotoxic T cells, and the release of proinflammatory cytokines. Pre-B-cell colony-enhancing factor (PBEF) is an inflammatory cytokine involved in several inflammatory diseases, and it has been identified to react with several cytokines involved in HLH. Objective We aimed to study the clinicoepidemiological characteristics of Egyptian children with HLH and to evaluate the role of PBEF as a diagnostic and prognostic marker in Egyptian children with HLH. Patients and methods This is a cross-sectional study that recruited fifteen children with HLH from the Pediatric Hematology/Oncology unit. There were 11 male and four female, and their median age (interquartile range) was 13 months. Patients underwent thorough clinical assessment, laying stress on disease manifestation, classification, treatment, and prognosis. Plasma concentration of PBEF was determined using an enzyme-linked immunosorbent assay. Results Four patients were classified as having primary HLH, seven patients were as having secondary HLH and four patients had unknown classification owing to waiting for genotyping. The risk factors before presentation were as follow: four had viral infection, three patients had malignancy, and two patients had immune deficiency before presentation. Seven patients of the study group died and eight patients were still alive. PBEF was significantly increased in the patient group than the control group. PBEF level showed a significant positive correlation with serum ferritin and triglycerides level and a negative correlation with fibrinogen level. Conclusion The elevated PBEF level and its correlation with the widely available biochemical markers for diagnosis of pediatric HLH indicates that it may be involved in its inflammatory process.

Polydatin Attenuates Atherosclerosis in ApoE Mice through PBEF Mediated Reduction of Cholesterol Deposition.

Cholesterol metabolism becomes imbalanced during the formation of macrophage-derived foam cells. Pre-B-cell colony-enhancing factor (PBEF) has recently been found to affect lipid deposition and inflammation in atherosclerosis. Here, we aimed to study the effects and molecular mechanism of Polydatin on atherosclerosis in ApoE-knockout (ApoE ) mice. Thirty ApoE mice were fed a high-fat diet (HFD) for 12 weeks, and then treated with Polydatin for another 12 weeks. Whole aortas and cryosections were stained with oil red O. Blood lipid, PBEF and cytokine levels were measured by ELISA. The mRNAs of cholesterol metabolism-related genes were determined by qRT-PCR and protein levels by Western blotting. Cell cholesterol content and viability were determined in macrophages and RAW 264.7 cells. PBEF siRNA was used to study the effect of Polydatin on cholesterol metabolism in macrophages incubated with ox-LDL. Polydatin lowered blood lipids and decreased atherosclerotic lesions in ApoE mice. The expression of cytokines and the mRNA of cholesterol metabolism-related genes were markedly regulated by Polydatin. Meanwhile, PBEF mRNA and protein were both greatly down-regulated by Polydatin. In vitro, Polydatin protected RAW 264.7 cells treated by ox-LDL and inhibited cholesterol uptake by macrophages. The PBEF siRNA result indicates that Polydatin can modulate cholesterol metabolism in macrophages, partly through down-regulation of PBEF. In conclusion, Polydatin relieves atherosclerosis injury in ApoE mice, mainly through down-regulation of PBEF and inhibition of PBEF-inducing cholesterol deposits in macrophages.

NAD metabolism fuels human and mouse intestinal inflammation

ObjectiveNicotinamide phosphoribosyltransferase (NAMPT, also referred to as pre-B cell colony-enhancing factor or visfatin) is critically required for the maintenance of cellular nicotinamide adenine dinucleotide (NAD) supply catalysing the rate-limiting step...

Emodin alleviates severe acute pancreatitis-associated acute lung injury by decreasing pre-B-cell colony-enhancing factor expression and promoting polymorphonuclear neutrophil apoptosis.

The present study aimed to evaluate the protective effects of emodin on severe acute pancreatitis (SAP)-associated acute lung injury (ALI), and investigated the possible mechanism involved. SAP was induced in Sprague-Dawley rats by retrograde infusion of 5% sodium taurocholate (1 ml/kg), after which, rats were divided into various groups and were administered emodin, FK866 [a competitive inhibitor of pre-B-cell colony-enhancing factor (PBEF)] or dexamethasone (DEX). DEX was used as a positive control. Subsequently, PBEF expression was detected in polymorphonuclear neutrophils (PMNs) isolated from rat peripheral blood by reverse transcription-quantitative polymerase chain reaction and western blotting. In addition, histological alterations, apoptosis in lung/pancreatic tissues, apoptosis of peripheral blood PMNs and alterations in the expression of apoptosis-associated proteins were examined by hematoxylin and eosin staining, terminal deoxynucleotidyl-transferase-mediated dUTP nick end labeling assay, Annexin V/propidium iodide (PI) assay and western blotting, respectively. Serum amylase activity and wet/dry (W/D) weight ratios were also measured. An in vitro study was also conducted, in which PMNs were obtained from normal Sprague-Dawley rats and were incubated with emodin, FK866 or DEX in the presence of lipopolysaccharide (LPS). Apoptosis of PMNs and the expression levels of apoptosis-associated proteins were examined in cultured PMNs in vitro by Annexin V/PI assay and western blotting, respectively. The results demonstrated that emodin, FK866 and DEX significantly downregulated PBEF expression in peripheral blood PMNs. In addition, emodin, FK866 and DEX reduced serum amylase activity, decreased lung and pancreas W/D weight ratios, alleviated lung and pancreatic injuries, and promoted PMN apoptosis by regulating the expression of apoptosis-associated proteins: Fas, Fas ligand, B-cell lymphoma (Bcl)-2-associated X protein, cleaved caspase-3 and Bcl-extra-large. In addition, the in vitro study demonstrated that emodin, FK866 and DEX significantly reversed the LPS-induced decrease of apoptosis in PMNs by regulating the expression of apoptosis-associated proteins. In conclusion, the present study demonstrated that emodin may protect against SAP-associated ALI by decreasing PBEF expression, and promoting PMN apoptosis via the mitochondrial and death receptor apoptotic pathways.

Inhibition of pre-B cell colony-enhancing factor attenuates inflammation induced by hyperoxia in EA.hy926 cells.

The aim of this study was to investigate the role of pre-B cell colony-enhancing factor(PBEF) in the pathogenesis of bronchopulmonary dysplasia(BPD) using an established cell model of BPD. For this purpose, EA.hy926 cell cultures were divided into 4groups as follows: the air group as the blank control, the hyperoxia group, the hyperoxia plus PBEF siRNA group and the hyperoxia plus scramble siRNA group. Cell viability and the generation of reactive oxygen species(ROS) were determined using respective kits. Moreover, the protein and mRNA expression levels of PBEF, interleukin-8(IL-8) and tumor necrosis factor-α(TNF-α) were also detected by corresponding methods. Compared with the hyperoxia group, the ROS levels in the hyperoxia plus PBEFsiRNA group were significantly reduced(P<0.01). The silencing of PBEF increased cell viability compared with the hyperoxia group. The protein and mRNA expression levels of PBEF, IL-8 and TNF-α were all decreased in the hyperoxia plus PBEFsiRNA group compared with the hyperoxia group(P<0.01). Our study thus demonstrates that the inhibition of PBEF attenuates oxidative stress and inflammation induced by hyperoxia in EA.hy926 cells, suggesting that PBEF may be a potential diagnostic and therapeutic target, which may be used for the development of novel treatment strategies for BPD.

Regulation and Function of Extracellular Nicotinamide Phosphoribosyltransferase/Visfatin.

Nicotinamide phosphoribosyltransferase (NAMPT) is an adipokine-enzyme, which was described as to play bioactivities both in the intracellular and in the extracellular environment. However, while the functions of intracellular NAMPT (iNAMPT) are well known, much less is known on extracellular NAMPT (eNAMPT), also called visfatin or pre-B cell colony-enhancing factor. iNAMPT catalyzes the rate-limiting step in the NAD+ biosynthesis pathway from nicotinamide. Its inhibition severely reduces intracellular NAD+ levels, achieving anti-inflammatory and anti-cancer effects. eNAMPT can be detected in the human circulation and in many extracellular environments. Studies show that eNAMPT can act as a growth factor, as an enzyme, and as a cytokine, but its true mechanism of secretion and its physiological functions are still debated. Increased levels of eNAMPT have been associated with different metabolic disorders and cancers. eNAMPT was demonstrated to modulate the pathways involved in the pathophysiology of obesity, diabetes, atherosclerosis, and cardiovascular events by regulating the oxidative stress response, apoptosis, and inflammation. In cancer, eNAMPT was shown to play a pivotal role in modulating cancer cell metabolism, in promoting epithelial-to-mesenchymal transition and in shaping the tumor microenvironment. In line with these functions, circulating eNAMPT levels are frequently increased in cancer patients. Given these pleiotropic roles of eNAMPT in human disease, this protein has attracted attention as a therapeutic target. In this narrative review, we will discuss recent evidence on eNAMPT-driven signalling, highlighting the emerging pathophysiological roles of this protein in different disorders and the potential therapeutic opportunities linked to its targeting. © 2017 American Physiological Society. Compr Physiol 7:603-621, 2017.

Self-Association and Conformational Stability of NAMPT Protein

Nicotinamide phosphoribosyltransferase (NAMPT), also known as Visfatin and Pre B-cell colony enhancing factor (PBEF), is a rate-limiting enzyme in the salvage pathway required for nicotinamide adenine dinucleotide (NAD) biosynthesis. It is a highly conserved 52-kDa protein, found in living species from bacteria to humans [1]. The protein serves as a cytokine and is involved in cellular regulation influencing cancer, ischemia, obesity, and type-II diabetes [2, 3]. Despite being involved in many regulatory pathways, there is a paucity of information concerning the function of NAMPT, due to the limitations of in vivo assays, and lack of expression systems for the protein. Here, we successfully expressed the NAMPT protein using the pET-SUMO expression vector in E.coli strain SHuffle containing a hexa-His tag for protein purification. Activity assays demonstrated functionality of the protein. Moreover, initial biophysical characterization of the protein using circular dichroism revealed secondary structural elements consistent with crystallographic data. Dynamic light scattering showed the protein exists as large oligomeric units potentially involved in the NAMPT signal amplification pathway. Hydropathy analysis indicated possible hydrophobic patches on the protein surface that explains the native oligomeric state. Most striking, we discovered that NAMPT can be solubilized in n-dodecyl-β-D-maltopyranoside detergent in monomeric form. These findings open opportunities for further structural and functional investigations. Presently we are optimizing conditions for NMR experiments on NAMPT protein. These methods [4] are complementary to X-ray crystallography, and provide valuable information on the structure and dynamics, offering an important tool for understanding biological functioning.[1] T. Wang et al. (2006) Nat. Struct. Mol. Biol 13, 661.[2] S. M. Camp et al. (2015) Sci. Rep. 5, 13135.[3] A. Fukuhara et al. (2005) Science 307, 426.[4] T. R. Molugu et al. (2016) Chem. Rev. (in press).

Pre-B-cell colony enhancing factor expression and its clinical significance in colorectal cancer

Objective To explore the expression of pre-B-cell colony enhancing factor (PBEF) in colorectal cancer, and analyze the relationship between PBEF expression and the clinical pathological characte-ristics and its diagnostic and prognostic value. Methods Sixty-eight human colorectal tumor and normal colo-rectal tissues were collected, and the expression of PBEF was detected by immunohistochemical method. The relationship between the expression level of PBEF and the characteristics of tumors and prognosis was analyzed. Results The positive expression rate of PBEF in colorectal cancer tissues was 80.9%, higher than that in normal colorectal tissues, which was 29.4% (χ2=36.41, P 0.05), sex (χ2=0.39, P>0.05), histology type (χ2=0.54, P>0.05) or penetrating serosa (χ2=1.55, P>0.05), but significantly associated with the degree of differentiation (χ2=6.29, P 0.05). Conclusion The abnormal high expression of PBEF may be closely related to the occurrence and development of colorectal cancer. Key words: Colorectal neoplasms; Immunohistochemistry; Pre-B-cell colony enhancing factor

Aberrant Expression of TIMP-2 and PBEF Genes in the Placentae of Cloned Mice Due to Epigenetic Reprogramming Error.

Cloned mice derived from somatic or ES cells show placental overgrowth (placentomegaly) at term. We had previously analyzed cloned and normal mouse placentae by using two-dimensional gel electrophoresis and mass spectrometry to identify differential protein expression patterns. Cloned placentae showed upregulation of tissue inhibitor of metalloproteinase-2 (TIMP-2), which is involved in extracellular matrix degradation and tissue remodeling, and downregulation of pre-B cell colony enhancing factor 1 (PBEF), which inhibits apoptosis and induces spontaneous labor. Here, we used Western blotting to further analyze the protein expression levels of TIMP-2 and PBEF in cloned placentae derived from cumulus cells, TSA-treated cumulus cells, intracytoplasmic sperm injection (ICSI), and natural mating (NM control). Cloned and TSA-treated cloned placentae had higher expression levels of TIMP-2 compared with NM control and ICSI-derived placentae, and there was a positive association between TIMP-2 expression and the placental weight of cloned mouse concepti. Conversely, PBEF protein expression was significantly lower in cloned and ICSI placentae compared to NM controls. To examine whether the observed differences were due to abnormal gene expression caused by faulty epigenetic reprogramming in clones, we investigated DNA methylation and histone modification in the promoter regions of the genes encoding TIMP-2 and PBEF. Sodium bisulfite sequencing did not reveal any difference in DNA methylation between cloned and NM control placentae. However, ChIP assays revealed that the level of H3-K9/K14 acetylation at the TIMP-2 locus was higher in cloned placentae than in NM controls, whereas acetylation of the PBEF promoter was lower in cloned and ICSI placenta versus NM controls. These results suggest that cloned placentae appear to suffer from failure of histone modification-based reprogramming in these (and potentially other) developmentally important genes, leading to aberrant expression of their protein products. These changes are likely to be involved in generating the abnormalities seen in cloned mouse placentae, including enlargement and/or a lack of proper placental function.