Pancreatic Endocrine Hormones Research Articles

Adropin Is Expressed in Pancreatic Islet Cells and Reduces Glucagon Release in Diabetes Mellitus.

Diabetes mellitus affects 537 million adults around the world. Adropin is expressed in different cell types. Our aim was to investigate the cellular localization in the endocrine pancreas and its effect on modulating pancreatic endocrine hormone release in streptozotocin (STZ)-induced diabetic rats. Adropin expression in the pancreas was investigated in normal and diabetic rats using immunohistochemistry and immunoelectron microscopy. Serum levels of insulin, glucagon pancreatic polypeptide (PP), and somatostatin were measured using a Luminex® χMAP (Magpix®) analyzer. Pancreatic endocrine hormone levels in INS-1 832/3 rat insulinoma cells, as well as pancreatic tissue fragments of normal and diabetic rats treated with different concentrations of adropin (10-6, 10-9, and 10-12 M), were measured using ELISA. Adropin was colocalized with cells producing either insulin, glucagon, or PP. Adropin treatment reduced the number of glucagon-secreting alpha cells and suppressed glucagon release from the pancreas. The serum levels of GLP-1 and amylin were significantly increased after treatment with adropin. Our study indicates a potential role of adropin in modulating glucagon secretion in animal models of diabetes mellitus.

Analysis Of The Relationship Of Blood Sugar Control In Patients With Diabetes Mellitus With Compliance To Take Medicine In Balimbingan Hospital In 2019

Diabetes mellitus is a major public health problem because of its short-term and long-term complications. Diabetes Mellitus (DM) is a disease that involves pancreatic endocrine hormones, including insulin and glucagon. The main manifestations include disturbances of lipid, carbohydrate, and protein metabolism, which in turn stimulates hyperglycemia. This hyperglycemic condition will develop into diabetes mellitus with various forms of complications. Adherence is very important, especially in long-term treatment. In general, the level of compliance in each patient is described by the percentage of the number of drugs taken and the time of taking the drug within a certain period of time. This study aims to determine the relationship between controlled blood sugar in patients with diabetes mellitus with medication adherence at Balimbingan Hospital. The design of this study was cross sectional, with a quantitative approach with a sample of 56 patients. The data collection method in this study used a questionnaire and data on visits to patients with diabetes mellitus at Balimbingan Hospital.

Pancreatic cancer-associated diabetes mellitus is characterized by reduced β-cell secretory capacity, rather than insulin resistance

AimsThe early distinction of pancreatic cancer associated diabetes (PaCDM) in patients with elderly diabetes is critical. However, PaCDM and type 2 diabetes mellitus (T2DM) remain indistinguishable. We aim to address the differences between the pancreatic and gut endocrine hormones of patients with PaCDM and T2DM. MethodsA total of 44 participants underwent mixed meal tolerance test (MMTT). Fasting and postprandial concentrations of insulin, C-peptide, glucagon, pancreatic polypeptide (PP), glucagon-like peptide-1 (GLP-1), and gastric inhibitory peptide (GIP) were measured. Insulin sensitivity and secretion indices were calculated. One-way ANOVA with post-hoc analysis was used for statistical analysis. ResultsInsulin and C-peptide responses to MMTT were blunted in PaCDM patients compared with T2DM. Baseline concentrations and AUCs differed. PaCDM patients showed lower insulin secretion capacity but better insulin sensitivity than T2DM patients. The peak concentration and AUC of PP in T2DM group were higher than healthy controls, but in accordance with PaCDM. PaCDM patients presented lower baseline GLP-1 concentration than T2DM patients. No between-group differences were found for glucagon and GIP. ConclusionsPaCDM patients had a lower baseline and postprandial insulin and C-peptide secretion than T2DM patients. Reduced insulin secretion and improved peripheral sensitivity were found in PaCDM patients compared with T2DM.

Study of Serum Pancreatic Amylase and Lipase Enzyme in Patients with Type 2 Diabetes

Introduction: Diabetes mellitus (DM) is a metabolic disorder that is greatly exacerbated by a complete lack of insulin or insulin resistance. The pancreas is a multicellular organ, the exocrine half accounting for 84% of its volume and the endocrine half accounting for only 2%. Since these two parts of the world have a close relationship with structure and function, the disruption of one can affect the other. Hemoglobin A1C (HbA1c) represents the glycemic status of the patient over the previous three months. Evidence suggests that pancreatic endocrine hormones, especially insulin, affect pancreatic exocrine function. Insulin has a detrimental effect on exocrine acinar cells. Exocrine acinar cells attached to it contain a variety of enzymes, including amylase and lipase, which help digest certain food particles.
 Aim: Study of serum pancreatic amylase and lipase enzyme in patients with type 2 diabetes.
 Materials and Methods: This cross sectional study was done in the Department of Biochemistry, dept. of medicine and Diabetic OPD, Datta Meghe Medical College and Shalinitai Meghe Hospital and Research Centre, Nagpur. For this study 40 diagnosed type 2 diabetic patients of both sexes with age ranging 35-60 years were selected as study group.
 Results: FBS, HbA1C, serum pancreatic amylase, and lipase mean effects between control and patients showed statistically significant differences in type 2 diabetes mellitus (P <0.0001).
 Conclusion: We concluded that pancreatic amylase and lipase function are impaired in type 2 patients with diabetes, and this observation is particularly important in type 2 diabetes. It has been suggested that the analysis of pancreatic enzymes in diabetic patients may be a useful parameter in determining the progression of the disease.

2062-P: Pancreatic Inflammation Implicates Increased Cellular Plasticity in the Pancreatic Duct Glands in Type 1 Diabetes

Type 1 diabetes (T1D) is an autoimmune disease characterized by endocrine/exocrine pancreas inflammation, fibrosis and, ultimately, decreased organ mass. Accumulating evidence suggests that this chronic inflammation may play a pivotal role in the initiation and progression of endogenous pancreatic cell plasticity. Pancreatic duct glands (PDGs), a potential stem/progenitor cell niche, represent a distinct anatomical compartment of the pancreas, forming blind-ending outpouchings from the main pancreatic duct and its branches. PDGs become activated in response to inflammation in T1D, evidenced by a high frequency of epithelial cell replication and an increase in progenitor-like cells. We addressed the hypothesis that in T1D, perturbation of hormone and non-hormone expressing cells in the interlobular ducts (ILD) and PDGs is a consequence of inflammation. Pancreatic sections from 21 T1D (disease duration 3 - 84 y) and 22 age-matched nondiabetic (ND) donors were evaluated for replication and expression of known pancreatic endocrine hormones in ducts and PDGs. We confirmed increased duct and PDG replication in T1D (T1D vs. ND— ILD: 2.8 ± 0.9 vs. 0.7 ± 0.1%, p<0.05; PDG: 3.4 ± 1.0 vs. 1.1 ± 0.4%, p<0.05) and reduced frequency of insulin expressing cells (T1D vs. ND— ILD: 0.0 ± 0.0 vs. 0.4 ± 0.1%, p<0.001; PDGs: 0.01 ± 0.01 vs. 0.8 ± 0.3%, p<0.05). The frequency of PDG cell replication was highest in T1D donors with acute pancreatitis. No differences were found in the frequency of other endocrine cell types regardless of the presence of pancreatic inflammation. These data suggest that increased pancreatic ductal cell replication in the PDG compartment of subjects with T1D is associated with sustained autoimmunity-related inflammation. Efforts are underway to determine specificity of pancreatic ductal cell changes to islet or acinar inflammation and the identity of replicating ILD and PDG cells, to better understand the mechanisms underlying pancreatic ductal cell plasticity. Disclosure S.S. Kulkarni: None. A.E. Butler: None. M.A. Atkinson: None.

Establishment of a rapid and footprint-free protocol for differentiation of human embryonic stem cells into pancreatic endocrine cells with synthetic mRNAs encoding transcription factors

BackgroundTransplantation of pancreatic β cells generated in vitro from pluripotent stem cells (hPSCs) such as embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) has been proposed as an alternative therapy for diabetes. Though many differentiation protocols have been developed for this purpose, lentivirus-mediated forced expression of transcription factors (TF)—PDX1 and NKX6.1—has been at the forefront for its relatively fast and straightforward approach. However, considering that such cells will be used for therapeutic purposes in the future, it is desirable to develop a procedure that does not leave any footprint on the genome, as any changes of DNAs could potentially be a source of unintended, concerning effects such as tumorigenicity. In this study, we attempted to establish a novel protocol for rapid and footprint-free hESC differentiation into a pancreatic endocrine lineage by using synthetic mRNAs (synRNAs) encoding PDX1 and NKX6.1. We also tested whether siPOU5F1, which reduces the expression of pluripotency gene POU5F1 (also known as OCT4), can enhance differentiation as reported previously for mesoderm and endoderm lineages.MethodssynRNA-PDX1 and synRNA-NKX6.1 were synthesized in vitro and were transfected five times to hESCs with a lipofection reagent in a modified differentiation culture condition. siPOU5F1 was included only in the first transfection. Subsequently, cells were seeded onto a low attachment plate and aggregated by an orbital shaker. At day 13, the degree of differentiation was assessed by quantitative RT-PCR (qRT-PCR) and immunohistochemistry for endocrine hormones such as insulin, glucagon, and somatostatin.ResultsBoth PDX1 and NKX6.1 expression were detected in cells co-transfected with synRNA-PDX1 and synRNA-NKX6.1 at day 3. Expression levels of insulin in the transfected cells at day 13 were 450 times and 14 times higher by qRT-PCR compared to the levels at day 0 and in cells cultured without synRNA transfection, respectively. Immunohistochemically, pancreatic endocrine hormones were not detected in cells cultured without synRNA transfection but were highly expressed in cells transfected with synRNA-PDX1, synRNA-NKX6.1, and siPOU5F1 at as early as day 13.ConclusionsIn this study, we report a novel protocol for rapid and footprint-free differentiation of hESCs to endocrine cells.

No Compensatory Increase in Non-ß Hormone Expressing Cells in the Pancreatic Duct Glands in Type 1 Diabetes

Longstanding type 1 diabetes (T1D) entails a near total loss of β cells secondary to autoimmunity, fibrosis and decreased total organ mass, but accumulating evidence suggests there may be an endogenous attempt to regenerate β cells. Pancreatic duct glands (PDGs) represent a distinct anatomical compartment of the pancreas, forming blind ending outpouchings from the main pancreatic duct and its branches. The PDG is purported to be a potential stem/progenitor cell niche. PDGs respond to inflammation in T1D by becoming activated, evidenced by a high frequency of epithelial cell replication and an increase in progenitor-like cells. Hence, we addressed the hypothesis that in T1D, perturbation of hormone-expressing endocrine cells in the interlobular ducts (ILD) and PDGs exist. Pancreatic sections from 10 T1D (disease duration 8-58 year) and 11 similarly aged control (ND) donors were evaluated for replication and expression of known pancreatic endocrine hormones in ducts and PDG. We confirmed increased duct and PDG replication in T1D (T1D vs. ND— ILD: 4.5 ± 1.8 vs. 0.8 ± 0.2%, p<0.05; PDG: 3.4 ± 1.3 vs. 0.4 ± 0.2%, p<0.05) and reduced frequency of insulin expressing cells (T1D vs. ND— ILD: 0.0 ± 0.0 vs. 0.4 ± 0.1%, p<0.001; PDGs: 0.± 0.07 vs. 2.4 ± 0.8%, p<0.05). No differences were found in the frequency of other known endocrine cell types (T1D vs. ND— ILD: Glucagon: 1.5 ± 0.7 vs. 2.1 ± 0.6%, Somatostatin: 1.2 ± 0.4 vs. 1.6 ± 0.3%, Pancreatic Polypeptide: 0.4 ± 0.2 vs. 1.3 ± 0.3%; PDGs: Glucagon: 0.9 ± 0.9 vs. 1.6 ± 1.0%, Somatostatin: 1.96 ± 0.8 vs. 3.7 ± 1.9%, Pancreatic Polypeptide: 1.7 ± 1.1 vs. 2.1 ± 0.8%; p=ns, all). These data suggest that increased cell replication and the deficiency of insulin expressing cells in the PDG compartment of subjects with T1D are not associated with a corresponding increase in non-insulin hormone-expressing endocrine cells. Furthermore, the consistent observation of increased cell replication in the ducts and PDGs in T1D may implicate regenerative mechanisms towards an exocrine cell fate. Disclosure S.S. Kulkarni: None. M.A. Atkinson: Other Relationship; Self; Patent Issued. A. Butler: None.

Targeted Mutation of NGN3 Gene Disrupts Pancreatic Endocrine Cell Development in Pigs

The domestic pig is an attractive model for biomedical research because of similarities in anatomy and physiology to humans. However, key gaps remain in our understanding of the role of developmental genes in pig, limiting its full potential. In this publication, the role of NEUROGENIN 3 (NGN3), a transcription factor involved in endocrine pancreas development has been investigated by CRISPR/Cas9 gene ablation. Precomplexed Cas9 ribonucleoproteins targeting NGN3 were injected into in vivo derived porcine embryos, and transferred into surrogate females. On day 60 of pregnancy, nine fetuses were collected for genotypic and phenotypic analysis. One of the piglets was identified as an in-frame biallelic knockout (Δ2/Δ2), which showed a loss of putative NGN3-downstream target genes: NEUROD1 and PAX4, as well as insulin, glucagon, somatostatin and pancreatic polypeptide-Y. Fibroblasts from this fetus were used in somatic cell nuclear transfer to generate clonal animals to qualify the effect of mutation on embryonic lethality. Three live piglets were born, received colostrum and suckled normally, but experienced extreme weight loss over a 24 to 36-hour period requiring humane euthanasia. Expression of pancreatic endocrine hormones: insulin, glucagon, and somatostatin were lost. The data support a critical role of NGN3 in porcine endocrine pancreas development.

Role of PI3K p110β in the differentiation of human embryonic stem cells into islet-like cells

To investigate the effects of the PI3K inhibitors on the differentiation of insulin-producing cells derived from human embryonic stem cells. Here, we report that human embryonic stem cells induced by phosphatidylinositol-3-kinase (PI3K) p110β inhibitors could produce more mature islet-like cells. Findings were validated by immunofluorescence analysis, quantitative real-time PCR, insulin secretion in vitro and cell transplantation for the diabetic SCID mice. Immunofluorescence analysis revealed that unihormonal insulin-positive cells were predominant in cultures with rare polyhormonal cells. Real-time PCR data showed that islet-like cells expressed key markers of pancreatic endocrine hormones and mature pancreatic β cells including MAFA. Furthermore, this study showed that the expression of most pancreatic endocrine hormones was similar between groups treated with the LY294002 (nonselective PI3K inhibitor) and TGX-221 (PI3K isoform selective inhibitors of class 1β) derivatives. However, the level of insulin mRNA in TGX-221-treated cells was significantly higher than that in LY294002-treated cells. In addition, islet-like cells displayed glucose-stimulated insulin secretion in vitro. After transplantation, islet-like cells improved glycaemic control and ameliorated the survival outcome in diabetic mice. This study demonstrated an important role for PI3K p110β in regulating the differentiation and maturation of islet-like cells derived from human embryonic stem cells.

Islet-like organoids derived from human pluripotent stem cells efficiently function in the glucose responsiveness in vitro and in vivo

Insulin secretion is elaborately modulated in pancreatic ß cells within islets of three-dimensional (3D) structures. Using human pluripotent stem cells (hPSCs) to develop islet-like structures with insulin-producing ß cells for the treatment of diabetes is challenging. Here, we report that pancreatic islet-like clusters derived from hESCs are functionally capable of glucose-responsive insulin secretion as well as therapeutic effects. Pancreatic hormone-expressing endocrine cells (ECs) were differentiated from hESCs using a step-wise protocol. The hESC-derived ECs expressed pancreatic endocrine hormones, such as insulin, somatostatin, and pancreatic polypeptide. Notably, dissociated ECs autonomously aggregated to form islet-like, 3D structures of consistent sizes (100–150 μm in diameter). These EC clusters (ECCs) enhanced insulin secretion in response to glucose stimulus and potassium channel inhibition in vitro. Furthermore, ß cell-deficient mice transplanted with ECCs survived for more than 40 d while retaining a normal blood glucose level to some extent. The expression of pancreatic endocrine hormones was observed in tissues transplanted with ECCs. In addition, ECCs could be generated from human induced pluripotent stem cells. These results suggest that hPSC-derived, islet-like clusters may be alternative therapeutic cell sources for treating diabetes.

Cell type-specific deletion in mice reveals roles for PAS kinase in insulin and glucagon production.

Aims/hypothesisPer-Arnt-Sim kinase (PASK) is a nutrient-regulated domain-containing protein kinase previously implicated in the control of insulin gene expression and glucagon secretion. Here, we explore the roles of PASK in the control of islet hormone release, by generating mice with selective deletion of the Pask gene in pancreatic beta or alpha cells.MethodsFloxed alleles of Pask were produced by homologous recombination and animals bred with mice bearing beta (Ins1Cre; PaskBKO) or alpha (PpgCre [also known as Gcg]; PaskAKO) cell-selective Cre recombinase alleles. Glucose homeostasis and hormone secretion in vivo and in vitro, gene expression and islet cell mass were measured using standard techniques.ResultsIns1Cre-based recombination led to efficient beta cell-targeted deletion of Pask. Beta cell mass was reduced by 36.5% (p < 0.05) compared with controls in PaskBKO mice, as well as in global Pask-null mice (38%, p < 0.05). PaskBKO mice displayed normal body weight and fasting glycaemia, but slightly impaired glucose tolerance, and beta cell proliferation, after maintenance on a high-fat diet. Whilst glucose tolerance was unaffected in PaskAKO mice, glucose infusion rates were increased, and glucagon secretion tended to be lower, during hypoglycaemic clamps. Although alpha cell mass was increased (21.9%, p < 0.05), glucagon release at low glucose was impaired (p < 0.05) in PaskAKO islets.Conclusions/interpretationThe findings demonstrate cell-autonomous roles for PASK in the control of pancreatic endocrine hormone secretion. Differences between the glycaemic phenotype of global vs cell type-specific null mice suggest important roles for tissue interactions in the control of glycaemia by PASK.Electronic supplementary materialThe online version of this article (doi:10.1007/s00125-016-4025-1) contains peer-reviewed but unedited supplementary material, which is available to authorised users.

In vitro differentiation of human amniotic epithelial cells into insulin-producing 3D spheroids.

Regenerative medicine and stem cell therapy may represent the solution for the treatment of non-curable human diseases such as type 1 diabetes. In this context of growing demand for functional and safe stem cells, human amniotic epithelial cells (hAECs) from term placenta have attracted increasing interest for their wide availability, stem cell properties, and differentiation plasticity, which make them a promising tool for stem cell-based therapeutic applications. We initially assayed the stemness characteristics of hAECs in serum-free conditions. Subsequently we developed a culture procedure on extracellular matrix for the formation of three-dimensional (3D) spheroids. Finally, we tested the immunomodulation and differentiation potential of hAEC spheroids: the presence of pancreatic endocrine hormones was revealed with transmission electron microscopy and immunofluorescence analyses; the release of C-peptide in hyperglycemic conditions was assayed with ELISA. The serum-free culture conditions we applied proved to maintain the basic stemness characteristics of hAECs. We also demonstrated that 3D spheroids formed by hAECs in extracellular matrix can be induced to differentiate into insulin-producing cells. Finally, we proved that control and induced cells equally inhibit the proliferation of activated mononuclear cells. The results of this study highlight the properties of amnion derived epithelial cells as promising and abundant source for cell-based therapies. In particular we are the first group to show the in vitro pancreatic induction of hAECs cultured on extracellular matrix in a 3D fashion. We accordingly propose the outcomes of this study as a novel contribution to the development of future cell replacement therapies involving placenta-derived cells.

Relationship of visfatin level to pancreatic endocrine hormone level, HOMA-IR index, and HOMA β-cell index in overweight women who performed hydraulic resistance exercise.

[Purpose] This study aimed to examine the correlation of visfatin level to pancreatic endocrine hormone level, homeostasis model assessment of insulin resistance (HOMA-IR) index, and HOMA β-cell index in hydraulic resistance exercise. Furthermore, it investigated the relationship between visfatin level and other variables affected by exercise in overweight women. [Subjects and Methods] The exercise group trained for 12 weeks, 70 minutes/day, 5 days/week. Visfatin level, pancreatic endocrine hormone level, HOMA-IR index, and HOMA β-cell index were measured before and after the intervention. Based on the blood insulin and glucose concentrations, HOMA-IR index, the indicator of insulin resistance, and HOMA β-cell index, the indicator of insulin secretion level, were assessed. [Results] Interaction effects on visfatin level, insulin level, HOMA-IR index, and HOMA β-cell index were observed. Interaction effects on glucagon and glucose levels were not observed between the intervention groups. The correlations of visfatin level to insulin, glucagon, and glucose levels, and HOMA-IR and HOMA β-cell indexes were not significant for any of the subjects. [Conclusion] Therefore, the 12-week resistance exercise affected body composition, visfatin level, insulin level, HOMA-IR index, and HOMA β-cell index. Finally, visfatin was not related to insulin, glucagon, and glucose levels, and HOMA-IR and HOMA β-cell indexes.

In vitro Differentiation of Human Adipose Tissue-Derived Stem Cells into Islet-Like Clusters Promoted by Islet Neogenesis-Associated Protein Pentadecapeptide

Human adipose tissue-derived stem cells (hASCs) are considered an ideal tool for the supply of insulin-producing cells to treat diabetes mellitus, with high differentiation efficiency. Islet neogenesis-associated protein (INGAP) is an initiator of islet neogenesis, and the peptide sequence comprising amino acids 104-118, named INGAP pentadecapeptide (INGAP-PP), has been shown to increase β-cell mass in animals and human pathological states. Here, we report a novel 4-step method to promote hASCs to differentiate into islet-like clusters (ILCs) more efficiently by adding INGAP-PP. The hASCs were isolated, purified and differentiated using a 4-step protocol including trichostatin A, INGAP-PP/scrambled peptide (Scrambled-P), dexamethasone, nicotinamide, glucagon-like peptide-1, transforming growth factor β1 and exendin-4. Results showed that ILCs in the INGAP-PP group were more similar to the fresh islets with regard to both size and morphology and expressed significantly higher levels of both insulin and C-peptide than those in the Scrambled-P group. Moreover, the ILCs from the INGAP-PP group secreted higher levels of insulin and C-peptide than those from the Scrambled-P group in response to both a low (5.6 m<smlcap>M</smlcap>) and high (25 m<smlcap>M</smlcap>) glucose challenge and secreted 6 times more hormones under the high-glucose challenge. Real-time PCR and immunocytochemistry showed that ILCs of the INGAP-PP group expressed human pancreatic endocrine hormones and transcription factors. Transplantation of ILCs into diabetic rats partially reversed diabetes and prolonged their life span. In conclusion, the INGAP-PP protocol can efficiently induce hASCs to differentiate into ILCs in vitro, and thus hASCs could be a promising source of cells for transplantation to treat diabetes mellitus.

Dynamic expression of microRNAs during the differentiation of human embryonic stem cells into insulin-producing cells

Human embryonic stem (hES) cells with the capacity of self-renewal and multilineage differentiation are promising sources for generation of pancreatic islet cells for cell replacement therapy in diabetes. Here we induced hES cells into insulin-producing cells (IPCs) in a stepwise process which recapitulated islet organogenesis by directing cells through the stages resembling definitive endoderm, gut-tube endoderm, pancreatic precursor and cells that expressed pancreatic endocrine hormones. The dynamic expression of microRNAs (miRNAs) during the differentiation was analyzed and was compared with that in the development of human pancreatic islets. We found that the dynamic expression patterns of miR-375 and miR-7 were similar to those seen in the development of human fetal pancreas, whereas the dynamic expression of miR-146a and miR-34a showed specific patterns during the differentiation. Furthermore, the expression of Hnf1β and Pax6, the predicted target genes of miR-375 and miR-7, was reciprocal to that of miR-375 and miR-7. Over-expression of miR-375 down-regulated the expression of gut-endoderm/pancreatic progenitor specific markers Hnf1β and Sox9. Therefore, the miRNAs may directly or indirectly regulate the expression of pancreatic islet organogenesis-specific transcription factors to control the differentiation and maturation of pancreatic islet cells.

Neuroendocrine Pancreatic Tumors: Guidelines for Management and Update

Pancreatic neuroendocrine tumors (PanNETs) are a diverse group of rare neoplasms. Commonly referred to as islet cell tumors, PanNETs are classified as functional or nonfunctional depending on their production of specific pancreatic endocrine hormones (e.g. insulin, gastrin, glucagon, and others) and association with the resultant clinical syndromes. While most PanNETs are sporadic, syndromic patients, in particular those with multiple endocrine neoplasia type 1 (MEN-1) and von Hippel Lindau (VHL), are at increased risk of developing these tumors. Recent investigations of patients with sporadic and syndromic PanNETs have elucidated critical pathways in tumor development, such as mammalian target of rapamycin (mTOR) signaling and its downstream growth factors such as vascular endothelial growth factor (VEGF). Prognosis ranges from favorable for localized, low-grade neoplasms to poor for advanced, high-grade tumors. Regardless of the stage at presentation, surgery is the first-line therapy for patients with disease amenable to surgical resection. We favor formal pancreatectomy with conventional lymph node sampling for the vast majority of patients, either through open or laparoscopic techniques. Those with insulinomas, however, may be candidates for enucleation. Cytoreductive surgery is also recommended for patients with locoregional recurrences or hepatic metastases. Regional adjuvants such as radiofrequency ablation (RFA), transarterial chemoembolization (TACE), and others are often employed in an attempt to palliate symptoms and prolong survival. Unfortunately, cytotoxic chemotherapy has been largely ineffective in treating patients with PanNETs. The somatostatin analogue octreotide, however, has been effective in palliating symptoms and slowing the progression of disease. Other promising systemic agents, including sunitinib and everolimus, have targeted critical PanNET signaling pathways. In summary, surgery remains the principal therapeutic strategy for patients with PanNETs, but continued research may identify more robust systemic therapies for those with advanced disease.

Nkx2.2 and Arx genetically interact to regulate pancreatic endocrine cell development and endocrine hormone expression

Nkx2.2 and Arx are essential pancreatic transcription factors. Nkx2.2 is necessary for the appropriate specification of the islet alpha, beta, PP and epsilon cell lineages, whereas Arx is required to form the correct ratio of alpha, beta, delta and PP cells. To begin to understand the cooperative functions of Nkx2.2 and Arx in the development of endocrine cell lineages, we generated progenitor cell-specific deletions of Arx on the Nkx2.2 null background. The analysis of these mutants demonstrates that expansion of the ghrelin cell population in the Nkx2.2 null pancreas is not dependent on Arx; however, Arx is necessary for the upregulation of ghrelin mRNA levels in Nkx2.2 mutant epsilon cells. Alternatively, in the absence of Arx, delta cell numbers are increased and Nkx2.2 becomes essential for the repression of somatostatin gene expression. Interestingly, the dysregulation of ghrelin and somatostatin expression in the Nkx2.2/Arx compound mutant (Nkx2.2null;ArxΔpanc) results in the appearance of ghrelin+/somatostatin+ co-expressing cells. These compound mutants also revealed a genetic interaction between Nkx2.2 and Arx in the regulation of the PP cell lineage; the PP cell population is reduced when Nkx2.2 is deleted but is restored back to wildtype numbers in the Nkx2.2null;ArxΔpanc mutant. Moreover, conditional deletion of Arx in specific pancreatic cell populations established that the functions of Arx are necessary in the Neurog3+ endocrine progenitors. Together, these experiments identify novel genetic interactions between Nkx2.2 and Arx within the endocrine progenitor cells that ensure the correct specification and regulation of endocrine hormone-producing cells.

MIN6 Is Not a Pure Beta Cell Line but a Mixed Cell Line with Other Pancreatic Endocrine Hormones

MIN6 cells retains glucose-stimulated insulin secretion (GSIS) as isolated islets. We comprehensively evaluated the gene expression and production of other islet hormones in MIN6 cells. Islet hormones were demonstrated by immunohistochemical staining and measured by ELISA. The gene expression profiles of MIN6 cells were compared with those in the mouse islets obtained by the laser capture micro-dissection (LCM). MIN6 cells excreted insulin, glucagon, somatostatin and ghrelin. They expressed mRNAs of insulin I and II, proglucagon, somatostatin, pancreatic polypeptide (PP) and ghrelin which were shown in the mouse pancreatic islet core and periphery obtained by LCM. A variety of genes closely related to the islet hormone producing cells were expressed in MIN6. Confocal laser scanning microscopy revealed that MIN6 cells included not only insulin positive cells but also insulin and glucagon or somatostin double positive cells. Glucagon, somatostatin and ghrelin were detectable in the culture medium. The present study clearly demonstrated that MIN6 produce pancreatic endocrine cells. It would be possible to use this cell line as a model to research the development, cell differentiation and function of pancreatic islets.

MicroRNA miR-7 is preferentially expressed in endocrine cells of the developing and adult human pancreas

MicroRNAs (miRNA) are small non-coding RNAs that inhibit gene expression through binding to complementary messenger RNA sequences. miRNAs have been predicted to target genes important for pancreas development, proper endocrine cell function and metabolism. We previously described that miRNA-7 (miR-7) was the most abundant and differentially expressed islet miRNA, with 200-fold higher expression in mature human islets than in acinar tissue. Here we have analyzed the temporal and spatial expression of miR-7 in human fetal pancreas from 8 to 22 weeks of gestational age (wga). Human fetal (8–22 wga) and adult pancreases were processed for immunohistochemistry, in situ hybridization, and quantitative RT-PCR of miRNA and mRNA. miR-7 was expressed in the human developing pancreas from around 9 wga and reached its maximum expression levels between 14 and 18 wga, coinciding with the exponential increase of the pancreatic endocrine hormones. Throughout development miR-7 expression was preferentially localized to endocrine cells and its expression persisted in the adult pancreas. The present study provides a detailed analysis of the spatiotemporal expression of miR-7 in developing human pancreas. The specific localization of miR-7 expression to fetal and adult endocrine cells indicates a potential role for miR-7 in endocrine cell differentiation and/or function. Future functional studies of a potential role for miR-7 function in islet cell differentiation and physiology are likely to identify novel targets for the treatment of diabetes and will lead to the development of improved protocols for generating insulin-producing cells for cell replacement therapy.

In vitro transformation of adult rat hepatic progenitor cells into pancreatic endocrine hormone-producing cells

Pancreatic and duodenal homeobox factor 1 (Pdx-1) plays an important role in initiating differentiation toward pancreatic endocrine cells. The transdifferentiation or transformation of hepatocytes into pancreatic endocrine cells could be feasible, due to their similar cellular origins. Our goal in this study was to see if small hepatocytes (SHs) could give rise to pancreatic endocrine cells via exogenous Pdx-1 gene expression. SHs were cultured for 10 days before adenovirus (Adt)-mediated Pdx-1 gene transfection. We performed western blot analysis for pancreatic transcription factors in the nuclei and reverse-transcription polymerase chain reaction (RT-PCR) for the gene expression of pancreatic endocrine hormones. Confocal laser microscanning analysis was used to observe the transformation of SHs. Pancreatic transcription factors such as Pdx-1, Ngn3, NeuroD, Nkx2.2, and Pax6 were induced after Adt-Pdx-1 gene transfection. The mRNA expression of pancreatic endocrine hormones (insulin, glucagon, and somatostatin) was induced after the gene transfection. Pdx-1 was expressed in the nucleus, where the cells were positive for one or more of the hormones and cytokeratin (CK) 8. Some cells were positive for multiple hormones. The insulin level increased while the glucagon level decreased after the glucose loading test, depending on the glucose concentration. SHs are transformed into functional pancreatic endocrine cells after Pdx-1 gene transfection.