Immunocytochemical staining for islet amyloid polypeptide in pancreatic endocrine tumors

Aims/hypothesis: Islet amyloid polypeptide is originally identified as the chief constituent of amyloid in insulinomas and type 2 diabetic islets. This study aimed to identify islet amyloid polypeptide by immunocytochemical staining in pancreatic endocrine tumors including 30 cases of insulinomas and non-β-cell pancreatic endocrine tumors. Results: In normal islets, 62% of islet cells and 52% of insulin cells were granularly positive for insulin and IAPP, respectively, with more insulin positive cells than IAPP positive cells and some densely positive staining for insulin and IAPP in irregularly shaped a nuclear, degenerating islet β-cells. In pancreatic endocrine tumors, all 10 insulinomas were positive for islet amyloid polypeptide but 2 glucogonomas, 1 somatostatinoma, 6 of 7 pancreatic polypeptidomas, all 7 gastrinomas and all 3 non-functioning pancreatic endocrine tumors were negative for islet amyloid polypeptide whereas one pancreatic polypeptidoma was positive for islet amyloid polypeptide. Methods: Using commercially available rabbit anti-islet amyloid polypeptide antibody, immunocytochemical staining was performed on 30 cases of pancreatic endocrine tumors, consisting of 10 insulinomas, 2 glucagonomas, 1 somatostatinoma, 7 pancreatic polypeptidomas, 7 gastrinomas and 3 non-functioning pancreatic endocrine tumors. Pancreatic tissues containing pancreatic endocrine tumors were systematically immunostained for insulin, glucagon, somatostatin, pancreatic polypeptide, gastrin and chromogranin A, in addition to islet amyloid polypeptide. When normal pancreatic tissues adjacent to pancreatic endocrine tumors were present, insulin, glucagon, somatostatin and islet amyloid polypeptide positive cells were counted for a total of 20 islets, which were divided into large islets and medium islets for each case. Conclusions/Interpretations: All 10 insulinomas and 1 pancreatic polypeptidoma were granularly positive for islet amyloid polypeptide, suggesting all 10 insulinomas contained enough insulin granules for IAPP whereas only one non-β-cell pancreatic endocrine tumor was co-localized with islet amyloid polypeptide in their secretary granules.     

Immunocytochemical staining for islet amyloid polypeptide in pancreatic endocrine tumors

Aims/hypothesis: Islet amyloid polypeptide is originally identified as the chief constituent of amyloid in insulinomas and type 2 diabetic islets. This study aimed to identify islet amyloid polypeptide by immunocytochemical staining in pancreatic endocrine tumors including 30 cases of insulinomas and non-β-cell pancreatic endocrine tumors.

Results: In normal islets, 62% of islet cells and 52% of insulin cells were granularly positive for insulin and IAPP, respectively, with more insulin positive cells than IAPP positive cells and some densely positive staining for insulin and IAPP in irregularly shaped a nuclear, degenerating islet β-cells. In pancreatic endocrine tumors, all 10 insulinomas were positive for islet amyloid polypeptide but 2 glucogonomas, 1 somatostatinoma, 6 of 7 pancreatic polypeptidomas, all 7 gastrinomas and all 3 non-functioning pancreatic endocrine tumors were negative for islet amyloid polypeptide whereas one pancreatic polypeptidoma was positive for islet amyloid polypeptide.

Methods: Using commercially available rabbit anti-islet amyloid polypeptide antibody, immunocytochemical staining was performed on 30 cases of pancreatic endocrine tumors, consisting of 10 insulinomas, 2 glucagonomas, 1 somatostatinoma, 7 pancreatic polypeptidomas, 7 gastrinomas and 3 non-functioning pancreatic endocrine tumors. Pancreatic tissues containing pancreatic endocrine tumors were systematically immunostained for insulin, glucagon, somatostatin, pancreatic polypeptide, gastrin and chromogranin A, in addition to islet amyloid polypeptide. When normal pancreatic tissues adjacent to pancreatic endocrine tumors were present, insulin, glucagon, somatostatin and islet amyloid polypeptide positive cells were counted for a total of 20 islets, which were divided into large islets and medium islets for each case.

Conclusions/Interpretations: All 10 insulinomas and 1 pancreatic polypeptidoma were granularly positive for islet amyloid polypeptide, suggesting all 10 insulinomas contained enough insulin granules for IAPP whereas only one non-β-cell pancreatic endocrine tumor was co-localized with islet amyloid polypeptide in their secretary granules.     

Immunolocalization of insulin, glucagon, pancreatic polypeptide, and somatostatin in the pancreatic islets of the possum, Trichosurus vulpecula

Antibodies to insulin, glucagon, pancreatic polypeptide (PP), and somatostatin were used in the immunofluorescence procedure to demonstrate localization of the four hormones in cells of the pancreatic islets of the brushtailed possum, Trichosurus vulpecula. Most pancreatic islets revealed some differences in the topographical distribution and cell number of each endocrine cell type. Insulin immunoreactive cells were observed in most islets where they occurred as groups of cells peripherally and within the islet. In several islets glucagon cells were the predominant cell population and were distributed peripherally as well as centrally. Pancreatic polypeptide cells were fewer in number and usually occurred as single cells within the islet. Cells immunoreactive to antisomatostatin serum were observed in varying numbers in the peripheral and central regions of the islet. The present immunofluorescence study demonstrates differences in the topographical distribution of the four major pancreatic hormones between a marsupial species and several of the eutherian mammals.     

Expression of class II major histocompatibility complex antigens on pancreatic B cells in the NOD mouse

Summary To elucidate the role of class II major histocompatibility complex antigen expression on pancreatic B cells in the development of diabetes in the non-obese diabetic (NOD) mouse, indirect immunofluorescence was employed for I-A staining on Bouin-fixed pancreas sections of NOD mice (I-A of which was reported as d), B10.GD (I-A, d), BALB/c (I-A, d) and C3H/He (I-A, k). I-A positive islets were observed in all NOD mice examined. Positive reaction was detected in islets both with and without lymphocytic infiltrations. Double staining with anti-insulin, glucagon, somatostatin or pan creatic polypeptide antibodies revealed that I-A positive cells corresponded with insulin cells, while other types of pancreatic islet cells were virtually negative for I-A. Weaker staining was seen in islets of B10.GD and, to a lesser extent, in those of BALB/c mice. C3H/He mouse islet cells showed no I-A expression. These results demonstrated the expression of I-A antigens on pancreatic B cells in the NOD mouse.     

Paradoxical reduction in pancreatic glucagon with normalization of somatostatin and decrease in insulin in normoglycemic alloxan-diabetic dogs: a putative mechanism of glucagon irresponsiveness to hypoglycemia

In alloxan-diabetic (A-D) dogs, plasma glucagon does not increase when glycemia is decreased by insulin. Therefore, as in insulin-dependent diabetes mellitus (IDDM), increased glucose utilization is not matched by an increase in hepatic production. To explore further the abnormal effects of insulin on regulation of pancreatic glucagon, we studied content and morphology of pancreatic hormones in six normal (N) dogs, five hyperglycemic A-D (HD) dogs, and in four A-D dogs where normoglycemia was maintained by insulin (ND). Morphometric measurement of islets and of immunocytochemically localized A cells (glucagon) were performed by an image analysis system. In normal pancreas, islets of tail and body were bigger in size (tail = 4850 +/- 376 microns 2, body = 3256 +/- 198 microns 2), than the head (2009 +/- 207 microns 2). Glucagon content was 331 +/- 50 micrograms with a mean concentration of 8.5 +/- 0.9 micrograms/g in N dogs, and did not change in HD dogs (422 +/- 34 micrograms, 9.3 +/- 0.4 micrograms/g). With normoglycemia, glucagon content decreased by 5-fold (p less than 0.001). Morphometry indicated that, although A cell area per islet increased (2.7-fold), islet number decreased (70%), explaining the unchanged glucagon content in HD dogs. This decrease in islet number can also justify the dramatic glucagon decrease in ND dogs. Despite the 70% decrease in islet numbers in HD dogs, pancreatic somatostatin increased 3-fold (9.93 +/- 3.3 to 30.6 +/- 7.2 micrograms), indicating that its islet content was augmented 10-fold. Somatostatin content returned to normal with normoglycemia. Pancreatic insulin content in HD dogs was negligible (55 +/- 23 micrograms) when compared with that in N dogs (5500 micrograms) and it did not increase with normoglycemia. The distinct but markedly diminished insulin and proinsulin peaks in HD dogs nearly disappeared in ND dogs. Thus, in alloxan-diabetic HD dogs, 70% of islets are destroyed. A marked increase in glucagon in residual islets can explain the unchanged islet size despite the absence of B cells; however, the percent increase of somatostatin is larger than that of glucagon. Normoglycemia 1) normalizes somatostatin content, 2) further diminishes insulin and proinsulin synthesis presumably due to lack of hyperglycemic stimulus, and 3) paradoxically decreases pancreatic glucagon content 5-fold below its normal level. We hypothesize that with normalization of plasma insulin, glucagon content in each islet normalizes, but because of destruction of most islets, pancreatic glucagon content becomes extremely low.(ABSTRACT TRUNCATED AT 400 WORDS)     

Expression of the coiled coil domain containing protein 116 in the pancreatic islets and endocrine pancreatic tumors

Aims: Coiled coil domain containing protein 116 (CCDC116) is a product of the gene coiled coil domain containing 116 located on human chromosome 22. Its function has not yet been established. The present study focuses on the expression of this protein in human pancreatic islets and in the endocrine pancreatic tumors (EPTs). Methods and Results: Expression of the protein was evaluated by immunohistochemistry in endocrine pancreas from six patients and in various EPTs from 51 patients. In pancreatic islets, virtually all insulin, approx. 75% of the somatostatin, and approx. 60% of the pancreatic polypeptide (PP) cells were immunoreactive for the CCDC116 protein whereas glucagon, ghrelin and the exocrine cells were not. All insulinomas, gastrinomas, non-functioning sporadic tumors and the hereditary multihormonal EPTs were immunoreactive with variable relative incidence. Two of the three somatostatinomas, and one of the three ACTH-secreting tumors also expressed CCDC116. Conclusions: The CCDC116 protein is expressed in all islet cell types except the glucagon and ghrelin cells. Most of the EPTs also contained CCDC116 protein. These findings suggest that this protein may play some role for the above mentioned endocrine cells and tumors. Its function has to be investigated in future studies.     

Expression of the coiled coil domain containing protein 116 in the pancreatic islets and endocrine pancreatic tumors

Aims: Coiled coil domain containing protein 116 (CCDC116) is a product of the gene coiled coil domain containing 116 located on human chromosome 22. Its function has not yet been established. The present study focuses on the expression of this protein in human pancreatic islets and in the endocrine pancreatic tumors (EPTs). Methods and Results: Expression of the protein was evaluated by immunohistochemistry in endocrine pancreas from six patients and in various EPTs from 51 patients. In pancreatic islets, virtually all insulin, approx. 75% of the somatostatin, and approx. 60% of the pancreatic polypeptide (PP) cells were immunoreactive for the CCDC116 protein whereas glucagon, ghrelin and the exocrine cells were not. All insulinomas, gastrinomas, non-functioning sporadic tumors and the hereditary multihormonal EPTs were immunoreactive with variable relative incidence. Two of the three somatostatinomas, and one of the three ACTH-secreting tumors also expressed CCDC116. Conclusions: The CCDC116 protein is expressed in all islet cell types except the glucagon and ghrelin cells. Most of the EPTs also contained CCDC116 protein. These findings suggest that this protein may play some role for the above mentioned endocrine cells and tumors. Its function has to be investigated in future studies.     

Islet amyloid polypeptide in pancreatic islets from type 1 diabetic subjects

Aims/hypothesis:

Islet amyloid polypeptide is originally isolated as the chief constituent of amyloid deposits in type 2 diabetic islets. Islet amyloid polypeptide hyposecretion was known in type 1 diabetics and this study aimed to detect possibly reduced islet amyloid polypeptide-positive cells in type 1 diabetic islets.

Results:

Non-diabetic control islets showed about 60% of islet cells were insulin cells, and 60% of insulin cells were positive for IAPP. In type 1 diabetic islets, islets were generally smaller than control islets, consisting of weaker positive cells for insulin and islet amyloid polypeptide. Medium-sized islets still retained some insulin positive cells, whereas islet amyloid polypeptide positive cells were much less or even absent, but some insulin-negative cells were weakly islet amyloid polypeptide positive. An occasional extra-large islet, representing regenerating islets, consisting of more than 100 islet cells revealed less than 35% insulin and 20% islet amyloid polypeptide positive cells with relatively increased glucagon and somatostatin cells. Both normal and type 1 diabetic islets revealed scattered, densely insulin and islet amyloid polypeptide positive sickle-shaped cytoplasm without granular appearance, consistent with degenerating insulin cells.

Methods:

Using commercially available rabbit anti-islet amyloid polypeptide antibody, immunostaning was performed on ten cases of type 1 diabetic pancreata and eight non-diabetic controls. Both control and type 1 diabetic pancreata were systematically immunostained for insulin, glucagon, somatostatin and islet amyloid polypeptide.

Conclusion/Interpretation:

Control islets consisted of about 60% insulin cells, and about 34% of islet cells were amyloid polypeptide positive with scattered and densely positive for insulin and islet amyloid polypeptide without granular appearance, consistent with degenerating β-cells. All islets, including occasional extra-large islets from type 1 diabetics, showed less insulin cells and less islet amyloid polypeptide positive cells with twice increased glucagon and somatostatin cells of the control islets, but some insulin-negative cells were positive for islet amyloid polypeptide, suggesting the presence of islet amyloid polypeptide in degenerating and extra large regenerating islets. Thus, this immunocytochemical staining revealed generally less islet amyloid positive cells in type 1 diabetic islets, corresponding to severe hyposecretion of islet amyloid polypeptide in type 1 diabetics.Key words: immunocytochemistry, islet amyloid polypeptide, pancreatic islets, type 1 diabetes     

An immunocytochemical study of endocrine cell development in the early fetal guinea pig pancreas.

The presence of insulin, glucagon, pancreatic polypeptide, and somatostatin containing cells and their ontogenic changes were investigated immunocytochemically in the early fetal pancreas of the guinea pig (Days 25-40). In the earliest tissues examined (Day 25 and Day 30) brightly staining glucagon cells were the most predominant endocrine cell population, followed by slightly fewer and weaker staining pancreatic polypeptide cells. Insulin and somatostatin immunoreactive cells were less numerous. At Day 25 all endocrine cells were located within the pancreatic tubules where some glucagon cells also coexpressed insulin. Similar dual immunoreactivity was present at Day 30. At Day 25 some of the pancreatic polypeptide cells also showed coexpression of somatostatin which persisted until Days 35-40. At these later stages insulin and somatostatin cells were increasingly frequent. Glucagon and pancreatic polypeptide cells were also conspicuous. The four endocrine cell types were found either in the pancreatic tubules or in the developing islets where they began to acquire an adult-like topographic distribution. These studies in the fetal guinea pig show that the four islet hormonal cells cytodifferentiate from an early stage. A small proportion of endocrine cells coexpress either insulin and glucagon or pancreatic polypeptide and somatostatin.     

Islet amyloid polypeptide in pancreatic islets from type 1 diabetic subjects

Aims/hypothesis: Islet amyloid polypeptide is originally isolated as the chief constituent of amyloid deposits in type 2 diabetic islets. Islet amyloid polypeptide hyposecretion was known in type 1 diabetics and this study aimed to detect possibly reduced islet amyloid polypeptide-positive cells in type 1 diabetic islets. Results: Non-diabetic control islets showed about 60% of islet cells were insulin cells, and 60% of insulin cells were positive for IAPP. In type 1 diabetic islets, islets were generally smaller than control islets, consisting of weaker positive cells for insulin and islet amyloid polypeptide. Medium-sized islets still retained some insulin positive cells, whereas islet amyloid polypeptide positive cells were much less or even absent, but some insulin-negative cells were weakly islet amyloid polypeptide positive. An occasional extra-large islet, representing regenerating islets, consisting of more than 100 islet cells revealed less than 35% insulin and 20% islet amyloid polypeptide positive cells with relatively increased glucagon and somatostatin cells. Both normal and type 1 diabetic islets revealed scattered, densely insulin and islet amyloid polypeptide positive sickle-shaped cytoplasm without granular appearance, consistent with degenerating insulin cells. Methods: Using commercially available rabbit anti-islet amyloid polypeptide antibody, immunostaning was performed on ten cases of type 1 diabetic pancreata and eight non-diabetic controls. Both control and type 1 diabetic pancreata were systematically immunostained for insulin, glucagon, somatostatin and islet amyloid polypeptide.

Conclusion/Interpretation: Control islets consisted of about 60% insulin cells, and about 34% of islet cells were amyloid polypeptide positive with scattered and densely positive for insulin and islet amyloid polypeptide without granular appearance, consistent with degenerating β cells. All islets, including occasional extra-large islets from type 1 diabetics, showed less insulin cells and less islet amyloid polypeptide positive cells with twice increased glucagon and somatostatin cells of the control islets, but some insulin-negative cells were positive for islet amyloid polypeptide, suggesting the presence of islet amyloid polypeptide in degenerating and extra-large regenerating islets. Thus, this immunocytochemical staining revealed generally less islet amyloid positive cells in type 1 diabetic islets, corresponding to severe hyposecretion of islet amyloid polypeptide in type 1 diabetics.     

A role for kisspeptin in islet function

Aims/hypothesis We investigated the production of kisspeptin (KISS1) and the KISS1 receptor, GPR54, in pancreatic islets and determined the effects of exogenous kisspeptin on insulin secretion.Methods RT-PCR and immunohistochemistry were used to detect expression of KISS1 and GPR54 mRNAs and the production of KISS1 and GPR54 in human and mouse islets and in beta (MIN6) and alpha- (alphaTC1) cell lines. The effects of KISS1 on basal and glucose-induced insulin secretion from mouse and human islets were measured in a perifusion system.Results KISS1 and GPR54 mRNAs were both detected in human and mouse islets, and GPR54 mRNA expression was also found in the MIN6 and alphaTC1 endocrine cell lines. In sections of mouse pancreas, KISS1 and GPR54 immunoreactivities were co-localised in both beta and alpha cells within islets, but were not detected in the exocrine pancreas. Exposure of mouse and human islets to KISS1 caused a stimulation of glucose-induced (20 mmol/l) insulin secretion, but had no effect on the basal rate of secretion at a sub-stimulatory concentration of glucose (2 mmol/l). In contrast, KISS1 inhibited insulin secretion from MIN6 cells at both 2 and 20 mmol/l glucose. KISS1 had no significant effect on glucagon secretion from mouse islets.Conclusions/interpretation This is the first report to show that the GPR54/KISS1 system is expressed in the endocrine pancreas, where it influences beta cell secretory function. These observations suggest an important role for this system in the normal regulation of islet function.     

Nesfatin-1 exerts a direct, glucose-dependent insulinotropic action on mouse islet β- and MIN6 cells

Nesfatin-1 is a recently discovered multifunctional metabolic hormone abundantly expressed in the pancreatic islets. The main objective of this study is to characterize the direct effects of nesfatin-1 on insulin secretion in vitro using MIN6 cells and islets isolated from C57BL/6 mice. We also examined the expression of the nesfatin-1 precursor protein, nucleobindin 2 (NUCB2) mRNA, and nesfatin-1 immunoreactivity (ir) in the islets of normal mice and in the islets from mice with streptozotocin-induced type 1 diabetes and diet-induced obese (DIO) mice with type 2 diabetes. Nesfatin-1 stimulated glucose-induced insulin release in vitro from mouse islets and MIN6 cells in a dose-dependent manner. No such stimulation in insulin secretion was found when MIN6 cells/islets were incubated with nesfatin-1 in low glucose. In addition, a fourfold increase in nesfatin-1 release from MIN6 cells was observed following incubation in high glucose (16.7 mM) compared to low glucose (2 mM). Furthermore, we observed a significant reduction in both NUCB2 mRNA expression and nesfatin-1-ir in the pancreatic islets of mice with type 1 diabetes, while a significant increase was observed in the islets of DIO mice. Together, our findings indicate that nesfatin-1 is a novel insulinotropic peptide and that the endogenous pancreatic islet NUCB2/nesfatin is altered in diabetes and diet-induced obesity.     

Effects of islet hormones on insulin secretion from cloned B cell lines, HIT-T15 and RINm5F

The effects of the islet cell hormones glucagon, somatostatin-28 and pancreatic polypeptide on insulin secretion from cultured cloned pancreatic B cells (HIT-T15 and RINm5F) have been investigated. Glucagon stimulates the secretion of insulin from HIT-T15 cells in the absence and presence of glucose and from RINm5F cells in the absence and presence of glyceraldehyde. HIT-T15 cells were more sensitive to the stimulatory effect of glucagon than RINm5F cells. Somatostatin-28 and pancreatic polypeptide, both alone and in combination, reduced glucose- and glucagon-stimulated insulin release from HIT-T15 cells and glyceraldehyde- and glucagon-stimulated insulin release from RINm5F cells. HIT-T15 cells were more sensitive to the inhibitory actions of somatostatin-28 and pancreatic polypeptide than RINm5F cells. This study supports the hypothesis that insulin release from normal B cells may be modified by the paracrine activity of islet hormones, glucagon, somatostatin and pancreatic polypeptide and probably occurs before any fine tuning imposed by subsequently released insulin.     

Rapid depletion of somatostatin in isolated mouse pancreatic islets after treatment with cysteamine

Cysteamine (CSH; beta-mercaptoethylamine) is known to deplete pancreatic somatostatin without affecting the insulin or glucagon content. It may therefore be useful for studies of intra-islet regulation of hormone release. In the present study injection of CSH (60 mg/kg body weight) to mice decreased the somatostatin content of their isolated pancreatic islets to 50% in 1 h and 30% in 4 h as compared to islets of non-injected controls. Exposure of isolated mouse islets to CSH (100 micrograms/ml) for either 0.5 h followed by incubation in control medium for 3.5 h, or continuously for 4 h, decreased the somatostatin content to about 40% of the controls. There was no change in the islet content of insulin or glucagon. Islets pretreated with CSH (100 micrograms/ml) for 1 h in vitro showed a decreased glucose stimulation of both oxygen consumption and glucose oxidation. Measurements of insulin release after a similar preincubation of the islets indicated an increased basal release and an attenuated glucose stimulation. It is concluded that CSH rapidly decreases islet somatostatin both in vivo and in vitro. This depletion may lead to a loss of tonic inhibition by islet somatostatin on basal insulin release. It is, however, more plausible that the increased basal insulin release reflected a direct effect of CSH on the islet beta-cells.     

Tissue culture of fetal rat islets: corticosterone promotes D cell maintenance and function

Fetal pancreatic islets were cultured using a recently described technique (1). After 1 day in culture, half of the plates were continued in control medium and half were grown in identical medium supplemented with corticosterone (0.1 microgram/ml). Media were renewed daily, and culture was continued for a total of 8 days. Insulin, glucagon, and somatostatin contents in the media were determined daily. These hormones were also estimated in the tissue at the time of plating, and after 1 and 8 days in culture. Islets were fixed on day 8, and the cells containing each of these hormones were identified by immunocytochemical staining. Corticosterone supplementation of the medium resulted in a 3-fold increase in the somatostatin concentration of the medium. The insulin and glucagon contents of the supplemented medium were slightly reduced. On day 8, there was no difference in the insulin content of the cultured tissue regardless of medium. The glucagon and somatostatin contents of the tissue grown in the steroid-supplemented medium were greater (1.8- and 3.1-fold, respectively) than those of the tissue grown in control medium. D cells were rare in the islets grown in control medium volume density, 0.4%, but were more numerous in the islets maintained in supplemented medium (2.2%). Islets grown in corticosterone-supplemented medium lacked an insulin secretory response to 22 mM glucose. These findings indicate that the volume densities of the cells within the islets can be altered during an 8-day period in culture, suggesting that nutritional and other requirements of the individual subpopulations of islet cells may be different. In addition, corticosterone may prevent the maturation of the secretory responsiveness of cultured B cells to glucose.     

Direct regulation of insulin secretion by angiotensin II in human islets of Langerhans

Aims/hypothesis This study aimed to identify the expression of angiotensin II receptors in isolated human islets and beta cells and to examine the functional consequences of their activation. Materials and methods Single-cell RT-PCR was used to identify whether human islet cells express mRNA for type 1 angiotensin II receptors (AT₁), and western blotting was used to determine AT₁ protein expression by human islets and MIN6 beta cells. We measured changes in intracellular calcium by microfluorimetry using Fura 2-loaded MIN6 cells and human islet cells. Dynamic insulin secretory responses were determined by RIA following perifusion of human islets and MIN6 cells. Results Human islets expressed mRNAs for both the angiotensin precursor, angiotensinogen, and for angiotensin-converting enzyme. In addition, human and mouse beta cells expressed AT₁. These were functionally coupled to increases in intracellular calcium, which occurred at least in part through phospholipase-C-sensitive mechanisms and calcium influx through voltage-operated calcium channels. Short-term exposure of human islets and MIN6 cells to angiotensin II caused a rapid, short-lived initiation of insulin secretion at 2 mmol/l glucose and potentiation of insulin secretion induced by glucose (at 8 and 16.7 mmol/l). Conclusions/interpretation These data demonstrate that the AT₁ is expressed by beta cells and that angiotensin II effects a short-lived and direct stimulation of human and mouse beta cells to promote insulin secretion, most probably through elevations in intracellular calcium. Locally produced angiotensin II may be important in regulating a coordinated insulin secretory response from beta cells.     

Establishment of a pancreatic beta cell line that retains glucose-inducible insulin secretion: special reference to expression of glucose transporter isoforms

Two cell lines have been established from insulinomas obtained by targeted expression of the simian virus 40 T antigen gene in transgenic mice. These cell lines, designated MIN6 and MIN7, produce insulin and T antigen and have morphological characteristics of pancreatic beta cells. MIN6 cells exhibit glucose-inducible insulin secretion comparable with cultured normal mouse islet cells, whereas MIN7 cells do not. Both cell lines produce liver-type glucose transporter (GT) mRNA at high level. Brain-type GT mRNA is also present at considerable level in MIN7 cells, but is barely detectable in MIN6 cells, suggesting that exclusive expression of the liver-type GT is related to glucose-inducible insulin secretion. MIN6 cells do not express either major histocompatibility (MHC) class I or class II antigens on the cell surface. However, treatment with interferon-gamma induces high levels of MHC class I antigens, and a combination of interferon-gamma and tumor necrosis factor-alpha induces a MHC class II antigen on the cell surface. These results emphasize that the MIN6 cell line retains physiological characteristics of normal beta cells. The MIN6 cell line will be especially useful to analyze the molecular mechanisms by which beta cells regulate insulin secretion in response to extracellular glucose concentrations. We discuss a possible role of GT isoforms in glucose sensing by beta cells.