GIP_HUMAN [22-51] Peptide Encoded by the Glucose-Dependent Insulinotropic Polypeptide (GIP) Gene Suppresses Insulin Expression and Secretion in INS-1E Cells and Rat Pancreatic Islets.

GIP_HUMAN [22-51] is a recently discovered peptide that shares the same precursor molecule with glucose-dependent insulinotropic polypeptide (GIP). In vivo, chronic infusion of GIP_HUMAN [22-51] in ApoE-/- mice enhanced the development of aortic atherosclerotic lesions and upregulated inflammatory and proatherogenic proteins. In the present study, we evaluate the effects of GIP_HUMAN [22-51] on insulin mRNA expression and secretion in insulin-producing INS-1E cells and isolated rat pancreatic islets. Furthermore, we characterize the influence of GIP_HUMAN [22-51] on cell proliferation and death and on Nf-kB nuclear translocation. Rat insulin-producing INS-1E cells and pancreatic islets, isolated from male Wistar rats, were used in this study. Gene expression was evaluated using real-time PCR. Cell proliferation was studied using a BrdU incorporation assay. Cell death was quantified by evaluating histone-complexed DNA fragments. Insulin secretion was determined using an ELISA test. Nf-kB nuclear translocation was detected using immunofluorescence. GIP_HUMAN [22-51] suppressed insulin (Ins1 and Ins2) in INS-1E cells and pancreatic islets. Moreover, GIP_HUMAN [22-51] promoted the translocation of NF-κB from cytoplasm to the nucleus. In the presence of a pharmacological inhibitor of NF-κB, GIP_HUMAN [22-51] was unable to suppress Ins2 mRNA expression. Moreover, GIP_HUMAN [22-51] downregulated insulin secretion at low (2.8 mmol/L) but not high (16.7 mmol/L) glucose concentration. By contrast, GIP_HUMAN [22-51] failed to affect cell proliferation and apoptosis. We conclude that GIP_HUMAN [22-51] suppresses insulin expression and secretion in pancreatic ß cells without affecting ß cell proliferation or apoptosis. Notably, the effects of GIP_HUMAN [22-51] on insulin secretion are glucose-dependent.

The biological role of neuronostatin, a hormone encoded by the somatostatin gene / Biologiczna rola neuronostatyny, hormonu kodowanego przez gen somatostatyny.

Neuronostatin is a peptide hormone encoded by the somatostatin gene. Neuronostatin was discovered in 2008 using bioinformatics methods. Studies in rodents have shown that it exerts a widespread effects in the central nervous system, as well as in peripheral tissues. It was demonstrated that neuronostatin reduces food and water intake, delays gastrointestinal transit, and may have important role in adipogenesis. It also increases glucagon secretion from pancreatic islet alpha cells. In addition, it has been proven that neuronostatin can affect cardiac contractility and blood pressure, and may be involved in processes related to memory, pain sensation and anxiety. In addition neuronostatin can induce a depression-like effect. In this article we review the literature addressing the biological role of neuronostatin in the organism.

Expression and localization of the neuropeptide phoenixin-14 and its receptor GRP173 in the canine reproductive organs and periovarian adipose tissue.

Phoenixin-14 (PNX-14) is a regulatory neuropeptide encoded by the SMIM20 gene, which has been implicated in the reproductive cycle by modulating the hypothalamic-pituitary-gonadal (HPG) axis. Recently, we showed that PNX-14 is downregulated in bitches with cystic endometrial hyperplasia and pyometra. The objective of this study was to determine the expression of Smim20, PNX-14, and its putative receptor GRP173 in the canine ovary (both healthy and those with ovarian cysts), periovarian adipose tissue (PAT) and in the endometrium during the oestrous cycle. The expression was analysed by RT-qPCR and Western blot. In tissue sections, peptides were localised by immunofluorescent assays, and blood plasma concentrations of PNX-14 were detected by EIA. The results demonstrated increased levels of PNX in bitches in the anestrus groups compared to diestrus animals. The expression of GPR173 increased in PAT during the diestrus phase and endometrial tissue in late diestrus bitches. In the ovary, strong signals of PNX-14 and GPR173 were detected in the luteal and follicular cells. Furthermore, bitches with cystic ovaries were characterised by elevated circulating PNX levels and a significantly higher expression of PNX and GPR173 in gonadal tissues, when compared with healthy animals. Moreover, a positive correlation between PNX and progesterone in the blood of healthy bitches was noted, which changed to a negative correlation in females affected by cystic ovaries. These studies expand the knowledge regarding the expression and localization of the PNX/GRP173 system in canine reproductive organs during physiological and pathological conditions.

Levels of spexin and its receptors GALR2 and GALR3 in the hypothalamus and ovary of letrozole-induced polycystic ovary syndrome in rats.

Spexin (SPX) is a newly identified neuropeptide, a natural ligand for the galanin receptors (GALR) 2/3, which is involved in maintaining physiological functions including female reproduction. One of the most common endocrine disorder in reproductive system is polycystic ovary syndrome (PCOS), however the role of SPX in PCOS is still unknown. The objective of this study was to determine the expression of mRNA and peptide levels of SPX and its receptors GALR2/3 in the hypothalamus and ovary (by real time PCR and Western blot) as well as plasma levels of SPX (ELISA) in letrozole - induced PCOS rats. We observed that SPX plasma level does not change in PCOS rats. In the hypothalamus transcript level of Spx and Galr3 were significantly higher in PCOS rats compared to the control, while mRNA of Galr2 and protein expression of GALR2/3 were lower. Moreover, expression of Spx and Galr2/3 mRNA as well as GALR2/3 peptide production were lower in the ovary of PCOS rats. In summary, while our results did not show differences in plasma SPX levels, we observed tissue-dependent significant differences in the SPX/GALR2/3 levels between PCOS and control rats, what indicates possible new mechanisms of PCOS neuroendocrinology.

Canine cystic endometrial hyperplasia and pyometra may downregulate neuropeptide phoenixin and GPR173 receptor expression.

The most common uterine diseases affecting bitches are cystic endometrial hyperplasia (CEH) and pyometra. The neuropeptide phoenixin (PNX) and its receptor (GPR173) are potential key factors involved in the proliferative and inflammatory regulation of the reproductive system in females. This study aimed to evaluate the expression of PNX and GPR173 by qPCR, western blot and immunofluorescence assays in the endometrium of bitches suffering from CEH or pyometra compared to clinically healthy females. Additionally, PNX and progesterone (P4) plasma concentrations were analysed. The results showed a significantly lower expression levels of PNX and GPR173 (mRNA and protein production) in bitches with the CEH or pyometra groups compared to healthy animals. Immunofluorescence staining examination also confirmed a lower concentration of PNX and GPR173 signals in bitches with pathological uteri. Moreover, a lower concentration of PNX blood levels in bitches suffering from pyometra was observed. The PNX concentration was negatively correlated with P4 but only in healthy bitches. These results illustrate that the development of canine uterine disorders may cause a lower expression of PNX and its receptor GPR173.

Phoenixin as a New Target in the Development of Strategies for Endometriosis Diagnosis and Treatment.

Small integral membrane protein 20/phoenixin (SMIM20/PNX) and its receptor GPR173 (G Protein-Coupled Receptor 173) play a role in the regulation of the hypothalamic-pituitary-gonadal axis (HPG). The aim of the study was to determine PNX, FSH, LH, and 17ß-estradiol association in women with endometriosis, and the expression of SMIM20/PNX signaling via GPR173. Serum PNX, FSH, LH, and 17ß-estradiol concentrations were measured by enzyme and electrochemiluminescence immunoassay. SMIM20/PNX and GPR173 expression in the eutopic and ectopic endometrium was assessed by qPCR and immunohistochemistry. Reduced PNX level, increased LH/FSH ratio and elevated 17ß-estradiol concentration were found in patients with endometriosis. No differences in SMIM20 expression were observed between the studied endometria. GPR173 expression was lower in ectopic than in eutopic endometria. SMIM20 expression was mainly restricted to stroma. GPR173 was detected in some eutopic and ectopic stromal cells and in eutopic glandular epithelial cells. Discriminant analysis indicates the diagnostic relevance of PNX and LH/FSH ratio in patients with endometriosis. In women with endometriosis, reduced PNX levels and GPR173 expression may be responsible for HPG axis dysregulation. These new insights may contribute to a better understanding of the pathophysiology of endometriosis and provide the basis for a new strategy for diagnosis and treatment of endometriosis.

Two weeks of moderate intensity locomotor training increased corticosterone concentrations but did not alter the number of adropin-immunoreactive cells in the hippocampus of diabetic type 2 and control rats.

Adropin (ADR) plays a role in metabolism regulation and its alterations in obesity and diabetes have been found. Treatment with ADR was beneficial in metabolic diseases, and physical exercise increased ADR concentrations in obese patients. However, data on the distribution of ADR in the brain are sparse. The role of metabolic status and physical exercise on its expression in the brain is undiscovered. We hypothesized that diabetes type 2 (DM2) and/or exercise will alter number of ADR-immunoractive (-ir) cells in the rat brain. Animals were divided into groups: diabetes type 2 (receiving high-fat diet and injections of streptozotocin) and control (fed laboratory chow diet; C). Rats were further divided into: running group (2 weeks of forced exercise on a treadmill) and non-running group. Body mass, metabolic and hormonal profiles were assessed. Immunohistochemistry was run to study ADR-ir cells in the brain. We found that: 1) in DM2 animals, running decreased insulin and increased glucose concentrations; 2) in C rats, running decreased insulin concentrations and had no effect on glucose concentration in blood; 3) running increased corticosterone (CORT) concentrations in DM2 and C rats; 4) ADR-ir cells were detected in the hippocampus and ADR-ir fibers in the arcuate nucleus of the hypothalamus, which is a novel location; 5) metabolic status and running, however, did not change number of these cells. We concluded that 2 weeks of forced moderate intensity locomotor training induced stress response present as increased concentration of CORT and did not influence number of ADR-ir cells in the brain.

Expression of Transforming Growth Factor Beta Isoforms in Canine Endometrium with Cystic Endometrial Hyperplasia-Pyometra Complex.

Cystic endometrial hyperplasia (CEH) and pyometra are the most frequently diagnosed uterine diseases affecting bitches of different ages. Transforming growth factor beta (TGF-ß) has been classified in females as a potential regulator of many endometrial changes during the estrous cycle or may be involved in pathological disorders. The aim of this study was to determine the expression of TGF-ß1, -ß2 and -ß3 in the endometrium of bitches suffering from CEH or a CEH-pyometra complex compared to clinically healthy females (control group; CG). A significantly increased level of TGF-ß1 mRNA expression was observed in the endometrium with CEH-pyometra compared to CEH and CG. Protein production of TGF-ß1 was identified only in the endometrium of bitches with CEH-pyometra. An increase in TGF-ß3 mRNA expression was observed in all the studied groups compared to CG. The expression of TGF-ß2 mRNA was significantly higher in CEH and lower in CEH-pyometra uteri. The results indicate the presence of TGF-ß cytokines in canine endometrial tissues affected by proliferative and degenerative changes. However, among all TGF-ß isoforms, TGF-ß1 could potentially be a key factor involved in the regulation of the endometrium in bitches with CEH-pyometra complex.

Adropin Slightly Modulates Lipolysis, Lipogenesis and Expression of Adipokines but Not Glucose Uptake in Rodent Adipocytes.

Adropin is a peptide hormone which modulates energy homeostasis and metabolism. In animals with diet-induced obesity, adropin attenuates adiposity and improves lipid and glucose homeostasis. Adropin promotes the proliferation of rodent white preadipocytes and suppresses their differentiation into adipocytes. By contrast, the effects of adropin on mature white adipocytes are unknown. Therefore, we aimed to evaluate the effects of adropin on lipolysis, lipogenesis and glucose uptake in white rodent adipocytes. We assessed the effects of adropin on the mRNA expression of adiponectin, resistin and visfatin. White preadipocytes were isolated from male Wistar rats. Differentiated 3T3-L1 cells were used as a surrogate model of white adipocytes. Lipolysis was measured by the evaluation of glycerol and free fatty acid secretion using colorimetric kits. The effects of adropin on lipogenesis and glucose uptake were measured using radioactive-labelled glucose. The expression of adipokine mRNA was studied using real-time PCR. Our results show that adropin slightly promotes lipolysis in rat adipocytes and 3T3-L1 cells. Adropin suppresses lipogenesis in rat adipocytes without influencing glucose uptake. In addition, adropin stimulates adiponectin mRNA expression and suppresses the expression of resistin and visfatin. These results indicate that adropin may be involved in controlling lipid metabolism and adipokine expression in white rodent adipocytes.

The effects of neuronostatin on proliferation and differentiation of rat primary preadipocytes and 3T3-L1 cells.

Neuronostatin is a peptide hormone encoded by the somatostatin gene. Biological effects of neuronostatin are mediated through activation of GPR107. There is evidence indicating that neuronostatin modulates energy homeostasis by suppressing food intake and insulin secretion, while stimulating glucagon secretion. While it was found that neuronostatin receptor is expressed in white adipose tissue, the role of neuronostatin in controlling adipose tissue formation is unknown. The aim of this study is to investigate the effects of neuronostatin on proliferation and differentiation of rat primary preadipocytes and 3T3-L1 cells. We found that neuronostatin receptor GPR107 is expressed in rat preadipocytes and 3T3-L1 cells. Neuronostatin promotes proliferation of preadipocytes via AKT activation. Downregulation of GPR107 mRNA expression and protein production results in an attenuation of neuronostatin-induced stimulation of preadipocyte proliferation. Moreover, neuronostatin reduces intracellular lipid content and the expression of adipogenesis-modulating genes C/ebpα, C/ebpß, Pparγ, and Fabp4. In summary, these results show that neuronostatin, AKT-dependently, stimulates the proliferation of preadipocytes via GPR107. In contrast, neuronostatin inhibits the differentiation of preadipocytes into mature adipocytes.

The Role of Peptide Hormones Discovered in the 21st Century in the Regulation of Adipose Tissue Functions.

Peptide hormones play a prominent role in controlling energy homeostasis and metabolism. They have been implicated in controlling appetite, the function of the gastrointestinal and cardiovascular systems, energy expenditure, and reproduction. Furthermore, there is growing evidence indicating that peptide hormones and their receptors contribute to energy homeostasis regulation by interacting with white and brown adipose tissue. In this article, we review and discuss the literature addressing the role of selected peptide hormones discovered in the 21st century (adropin, apelin, elabela, irisin, kisspeptin, MOTS-c, phoenixin, spexin, and neuropeptides B and W) in controlling white and brown adipogenesis. Furthermore, we elaborate how these hormones control adipose tissue functions in vitro and in vivo.

Phoenixin: More than Reproductive Peptide.

Phoenixin (PNX) neuropeptide is a cleaved product of the Smim20 protein. Its most common isoforms are the 14- and 20-amino acid peptides. The biological functions of PNX are mediated via the activation of the GPR173 receptor. PNX plays an important role in the central nervous system (CNS) and in the female reproductive system where it potentiates LH secretion and controls the estrus cycle. Moreover, it stimulates oocyte maturation and increases the number of ovulated oocytes. Nevertheless, PNX not only regulates the reproduction system but also exerts anxiolytic, anti-inflammatory, and cell-protective effects. Furthermore, it is involved in behavior, food intake, sensory perception, memory, and energy metabolism. Outside the CNS, PNX exerts its effects on the heart, ovaries, adipose tissue, and pancreatic islets. This review presents all the currently available studies demonstrating the pleiotropic effects of PNX.

Adropin stimulates proliferation but suppresses differentiation in rat primary brown preadipocytes.

Adropin is a peptide hormone encoded by Energy Homeostasis Associated (Enho) gene. Adropin modulates glucose and lipid metabolism, and adiposity. Recently, we found that adropin suppresses differentiation of rodent white preadipocytes into mature fat cells. By contrast, the role of adropin in controlling brown adipogenesis is largely unknown. Therefore, in the present study we evaluated the effects of adropin on proliferation and differentiation of adipocyte precursor cells in rats. Brown adipocyte precursor cells were isolated from male Wistar rats. Cell replication was measured by BrdU incorporation. Gene expression was studied using real time PCR. Protein phosphorylation and production was assessed by Western blot. Lipid accumulation was evaluated by Oil Red O staining. Colorimetric kits were used to evaluate glycerol and free fatty acids release. We report here that adropin stimulates proliferation of brown preadipocytes. Moreover, in brown preadipocytes, adropin suppresses mRNA expression of adipogenic genes (C/ebpα, C/ebpß, Pgc1α, Pparγ and Prdm16) during differentiation process. In addition, adropin suppresses UCP1 protein production in brown adipocytes. Finally, adropin reduces intracellular lipid content in brown adipocytes. These results indicate that adropin stimulates proliferation of brown preadipocytes and suppresses their differentiation into mature adipocytes.

Levels of the neuropeptide phoenixin-14 and its receptor GRP173 in the hypothalamus, ovary and periovarian adipose tissue in rat model of polycystic ovary syndrome.

Phoenixin (PNX) is a newly discovered peptide produced by proteolytic cleavage of a small integral membrane protein 20 (Smim20), which acts as an important regulator of energy homeostasis and reproduction. Since dysfunction of reproduction is characteristic in polycystic ovarian syndrome (PCOS), the role of PNX in pathogenesis of PCOS needs further investigation. The objective of this study was to determine expression of Smim20, PNX-14 and its receptor GRP173 in the hypothalamus, ovary and periovarian adipose tissue (PAT) of letrozole induced PCOS rats. Phosphorylation of extracellular signal-regulated kinase (ERK1/2), protein kinases A (PKA) and B (Akt) were also estimated. We observed that PCOS rats had high weight gain and a number of ovarian cyst, high levels of testosterone, luteinizing hormone and PNX-14, while low estradiol. Smim20 mRNA expression was higher in the ovary and PAT, while PNX-14 peptide production was higher only in the ovary of PCOS rat. Moreover, in PCOS rats Gpr173 level was lower in PAT but at the protein level increased only in the ovary. Depending on the tissues, kinases phosphorylation were significantly differ in PCOS rats. Our results showed higher levels of PNX-14 in PCOS rats and indicated some novel findings regarding the mechanisms of PCOS pathophysiology.

Adropin as A Fat-Burning Hormone with Multiple Functions-Review of a Decade of Research.

Adropin is a unique hormone encoded by the energy homeostasis-associated (Enho) gene. Adropin is produced in the liver and brain, and also in peripheral tissues such as in the heart and gastrointestinal tract. Furthermore, adropin is present in the circulatory system. A decade after its discovery, there is evidence that adropin may contribute to body weight regulation, glucose and lipid homeostasis, and cardiovascular system functions. In this review, we summarize and discuss the physiological, metabolic, and pathophysiological factors regulating Enho as well as adropin. Furthermore, we review the literature addressing the role of adropin in adiposity and type 2 diabetes. Finally, we elaborate on the role of adropin in the context of the cardiovascular system, liver diseases, and cancer.

Effects of adropin on proliferation and differentiation of 3T3-L1 cells and rat primary preadipocytes.

Adropin is a protein encoded by Energy Homeostasis Associated (Enho) gene which is expressed mainly in the liver and brain. There is evidence that biological effects of adropin are mediated via GPR19 activation. Animal studies showed that adropin modulates adiposity as well as lipid and glucose homeostasis. Adropin deficient animals have a phenotype closely resembling that of human metabolic syndrome with are obesity dyslipidemia and impaired glucose production. Animals treated with exogenous adropin lose weight, in addition to having reduced expression of lipogenic genes in the liver and fat tissue. While it was shown that adropin may contribute to energy homeostasis and body weight regulation, the role of this protein in controlling fat tissue formation is largely unknown. Thus, in the present study we investigated the effects of adropin on adipogenesis using 3T3-L1 cells and rat primary preadipocytes. We found a low Enho mRNA expression in 3T3-L1 cells and rat primary preadipocytes. Adropin stimulated proliferation of 3T3-L1 cells and rat primary preadipocytes. Stimulation of 3T3-L1 cell proliferation was mediated via ERK1/2 and AKT. Adropin reduced lipid accumulation as well as expression of proadipogenic genes in 3T3-L1 cells and rat preadipocytes, suggesting that this protein attenuates differentiation of preadipocytes into mature fat cells. In summary, these results show that adropin modulates proliferation and differentiation of preadipocytes.

Phoenixin-14 stimulates proliferation and insulin secretion in insulin producing INS-1E cells.

Phoenixin (PNX) is a recently discovered neuropeptide which modulates appetite, pain sensation and neurons of the reproductive system in the central nervous system. PNX is also detectable in the circulation and in peripheral tissues. Recent data suggested that PNX blood levels positively correlate with body weight as well as nutritional status suggesting a potential role of this peptide in controlling energy homeostasis. PNX is detectable in endocrine pancreas, however it is unknown whether PNX regulates insulin biosynthesis or secretion. Using insulin producing INS-1E cells and isolated rat pancreatic islets we evaluated therefore, whether PNX controls insulin expression, secretion and cell proliferation. We identified PNX in pancreatic alpha as well as in beta cells. Secretion of PNX from pancreatic islets was stimulated by high glucose. PNX stimulated insulin mRNA expression in INS-1E cells. Furthermore, PNX enhanced glucose-stimulated insulin secretion in INS-1E cells and pancreatic islets in a time-dependent manner. Stimulation of insulin secretion by PNX was dependent upon cAMP/Epac signalling, while potentiation of cell growth and insulin mRNA expression was mediated via ERK1/2- and AKT-pathway. These results indicate that PNX may play a role in controlling glycemia by interacting with pancreatic beta cells.

Spexin: A novel regulator of adipogenesis and fat tissue metabolism.

Spexin (SPX, NPQ) is a novel peptide involved in the regulation of energy metabolism. SPX inhibits food intake and reduces body weight. In obese humans, SPX is the most down-regulated gene in fat. Therefore, SPX might be involved in the regulation of lipid metabolism. Here, we study the effects of SPX on lipolysis, lipogenesis, glucose uptake, adipogenesis, cell proliferation and survival in isolated human adipocytes or murine 3T3-L1 cells. SPX and its receptors, GALR2 and GALR3, are present at mRNA and protein levels in murine 3T3-L1 cells and human adipocytes. SPX inhibits adipogenesis and down-regulates mRNA expression of proadipogenic genes such as Pparγ, C/ebpα, C/ebpß and Fabp4. SPX stimulates lipolysis by increasing the phosphorylation of hormone sensitive lipase (HSL). Simultaneously, SPX inhibits lipogenesis and glucose uptake in human adipocytes and murine 3T3-L1 cells. SPX has no effect on murine 3T3-L1 cell proliferation and viability. Moreover, our research showed that the SPX effect on adipocytes metabolism is mediated via GALR2 and GALR3 receptors. SPX is a novel regulator of lipid metabolism in murine 3T3-L1 and human adipocytes.

Phoenixin-14 stimulates differentiation of 3T3-L1 preadipocytes via cAMP/Epac-dependent mechanism.

Phoenixin-14 (PNX) is a newly discovered peptide produced by proteolytic cleavage of the small integral membrane protein 20 (Smim20). Previous studies showed that PNX is involved in controlling reproduction, pain, anxiety and memory. Furthermore, in humans, PNX positively correlates with BMI suggesting a potential role of PNX in controlling fat accumulation in obesity. Since the influence of PNX on adipose tissue formation has not been so far demonstrated, we investigated the effects of PNX on proliferation and differentiation of preadipocytes using 3T3-L1 and rat primary preadipocytes. We detected Smim20 and Gpr173 mRNA in 3T3-L1 preadipocytes as well as in rat primary preadipocytes. Furthermore, we found that PNX peptide is produced and secreted from 3T3-L1 and rat primary adipocytes. PNX increased 3T3-L1 preadipocytes proliferation and viability. PNX stimulated the expression of adipogenic genes (Pparγ, C/ebpß and Fabp4) in 3T3-L1 adipocytes. 3T3-L1 preadipocytes differentiated in the presence of PNX had increased lipid content. Stimulation of cell proliferation and differentiation by PNX was also confirmed in rat preadipocytes. PNX failed to induce AKT phosphorylation, however, PNX increased cAMP levels in 3T3-L1 cells. Suppression of Epac signalling attenuated PNX-induced Pparγ expression without affecting cell proliferation. Our data show that PNX stimulates differentiation of 3T3-L1 and rat primary preadipocytes into mature adipocytes via cAMP/Epac-dependent pathway. In conclusion our data shows that phoenixin promotes white adipogenesis, thereby may be involved in controlling body mass regulation.

TRPV6 modulates proliferation of human pancreatic neuroendocrine BON-1 tumour cells.

Highly Ca(2+) permeable receptor potential channel vanilloid type 6 (TRPV6) modulates a variety of biological functions including calcium-dependent cell growth and apoptosis. So far, the role of TRPV6 in controlling growth of pancreatic neuroendocrine tumour (NET) cells is unknown. In the present study, we characterize the expression of TRPV6 in pancreatic BON-1 and QGP-1 NET cells. Furthermore, we evaluate the impact of TRPV6 on intracellular calcium, the activity of nuclear factor of activated T-cells (NFAT) and proliferation of BON-1 cells. TRPV6 expression was assessed by real-time PCR and Western blot. TRPV6 mRNA expression and protein production were down-regulated by siRNA. Changes in intracellular calcium levels were detected by fluorescence calcium imaging (fura-2/AM). NFAT activity was studied by NFAT reporter assay; cell proliferation by bromodeoxyuridine (BrdU), MTT and propidium iodine staining. TRPV6 mRNA and protein are present in BON-1 and QGP-1 NET-cells. Down-regulation of TRPV6 attenuates BON-1 cell proliferation. TRPV6 down-regulation is associated with decreased Ca(2+) response pattern and reduced NFAT activity. In conclusion, TRPV6 is expressed in pancreatic NETs and modulates cell proliferation via Ca(2+)-dependent mechanism, which is accompanied by NFAT activation.