Per-Ola Carlsson

Angiotensin II and the endocrine pancreas: effects on islet blood flow and insulin secretion in rats.

Summary An intrinsic angiotensin system has been described in the pancreas, with angiotensin II specific receptors being present on both exocrine, endocrine and vascular cells. The aim of the present study was to evaluate the effects of angiotensin II on insulin secretion and blood flow regulation in the pancreas. Blood flows were determined with a microsphere technique. Infusion of angiotensin II induced a dose-dependent reduction in both whole pancreatic and islet blood flow, which was most pronounced in the former. Administration of enalaprilate, an inhibitor of angiotensin-converting enzyme, and saralasin, a non-selective angiotensin II receptor antagonist, preferentially increased islet blood flow. The effects of angiotensin II on insulin release were examined by measuring insulin concentrations in the effluents from isolated perfused pancreata. In these preparations, enalaprilate affected neither basal nor glucose-stimulated insulin release, whereas angiotensin II delayed the first phase of insulin release in response to glucose. The effect of angiotensin II was probably due to initial marked vasoconstriction. The retardation of insulin release could be avoided by adding angiotensin II to the perfusion medium 20 min before glucose administration, i. e. so that the vasoconstriction had disappeared when glucose-stimulation began. The present study suggests that the angiotensin-system is important in regulation of islet blood flow and points to a pivotal role of islet blood perfusion for an adequate insulin release. [Diabetologia (1998) 41: 127–133]

Graft vascular function after transplantation of pancreatic islets

Endogenous pancreatic islets have a dense glomerular-like angioarchitecture, which ensures an optimal delivery of oxygen and nutrients to the islet cells, provides signals from other cells in the body and disposes secreted hormones. Transplantation of isolated islets means that their vascular connection is interrupted. The islet grafts therefore depend upon endothelial cells and microvessels originating in the implantation organ for derivation of a new vascular system. A re-establishment of islet blood-flow occurs within 7–14 days after transplantation, mainly through vascular sprouting. The newly formed blood vessels acquire the morphological characteristics of those in endogenous islets. In intraportally transplanted islets to the liver, the islets become revascularized almost exclusively from tributaries to the hepatic artery. Exocrine contamination of the transplanted islets could hamper the revascularization process, whereas neither cryopreservation nor immunosuppressive drugs like cyclosporin, prednisolon and RS-61443 have any essential effects on the angiogenesis. Investigators have noticed improvements in islet graft survival and function by means of basic fibroblast growth factor (bFGF), acidic FGF and endothelial cell growth factor exposure of the grafts. The functional properties of transplanted islets are largely unknown, but evidence from experimental islet transplantation suggests that both the blood perfusion and the tissue oxygen tension of the grafted islets are chronically decreased, indicating an insufficient vascular system. In order to achieve optimal condition for survival and function of transplanted beta cells, it is important to ascertain whether impairments in vascular function are present also after clinical islet transplantations as well.

Reactive oxygen species cause diabetes-induced decrease in renal oxygen tension

Aims/hypothesisAugmented formation of reactive oxygen species (ROS) induced by hyperglycaemia has been suggested to contribute to the development of diabetic nephropathy. This study was designed to evaluate the influence of streptozotocin (STZ)-induced diabetes mellitus, as well as the effects of preventing excessive ROS formation by α-tocopherol treatment, on regional renal blood flow, oxygen tension and oxygen consumption in anaesthetized Wistar Furth rats.MethodsNon-diabetic and STZ-diabetic rats were investigated after 4 weeks with or without dietary treatment with the radical scavenger DL-α-tocopherol (vitamin E, 5%). A laser-Doppler technique was used to measure regional renal blood flow, whilst oxygen tension and consumption were measured using Clark-type microelectrodes.ResultsRenal oxygen tension, but not renal blood flow, was lower throughout the renal parenchyma of diabetic rats when compared to non-diabetic control rats. The decrease in oxygen tension was most pronounced in the renal medulla. Renal cellular oxygen consumption was markedly increased in diabetic rats, predominantly in the medullary region. Diabetes increased lipid peroxidation and protein carbonylation in the renal medulla. Treatment with α-tocopherol throughout the course of diabetes prevented diabetes-induced disturbances in oxidative stress, oxygen tension and consumption. The diabetic animals had a renal hypertrophy and a glomerular hyperfiltration, which were unaffected by α-tocopherol treatment.Conclusions/interpretationWe conclude that oxidative stress occurs in kidneys of diabetic rats predominantly in the medullary region and relates to augmented oxygen consumption and impaired oxygen tension in the tissue.

Evidence for a local angiotensin-generating system and dose-dependent inhibition of glucose-stimulated insulin release by angiotensin II in isolated pancreatic islets

Aims/hypothesisA local angiotensin-generating system has been found in the exocrine pancreas. This study aimed, primarily, to investigate the existence of a local angiotensin-generating system in the pancreatic islets and, secondly, to elucidate its role in regulating insulin secretion.MethodsReal-time RT-PCR and western blot were used to investigate if angiotensin-generating components are present in the mouse pancreatic islets, which are subject to regulation by islet transplantation. The localisation of AT1-receptors in islets was investigated by immunohistochemistry. Batch-type incubations of isolated islets were applied for studying the influence of angiotensin II on the glucose-stimulated insulin release, glucose oxidation and (pro)insulin, and total protein biosynthesis.ResultsMajor components, namely angiotensinogen, ACE, AT1- and AT2-receptors, were expressed in endogenous islets. AT1-receptors were localised to pancreatic beta cells. Exposure of the isolated islets to angiotensin II induced a dose-dependent inhibition of glucose-stimulated insulin release and inhibited (pro)insulin biosynthesis. This inhibitory action was fully preventable by pretreatment of the islets with losartan, an AT1-receptor antagonist. We also investigated if the expression of these components was changed after islet transplantation. Notably, a markedly increased expression of mRNA for the AT1-receptor was observed in islets retrieved from 4-week-old syngeneic islet transplants, a finding that was confirmed at the protein level.Conclusion/interpretationThese data indicate the existence of an islet angiotensin-generating system of potential importance in the physiological regulation of glucose-induced insulin secretion, thus diabetes mellitus. The increased expression of the AT1-receptor in islet transplants could have relevance to islet-graft function.

Female Mice are Protected against High-Fat Diet Induced Metabolic Syndrome and Increase the Regulatory T Cell Population in Adipose Tissue

Sex differences in obesity-induced complications such as type 2 diabetes have been reported. The aim of the study was to pinpoint the mechanisms resulting in different outcome of female and male mice on a high-fat diet (HFD). Mice fed control or HFD were monitored for weight, blood glucose, and insulin for 14 weeks. Circulating chemokines, islet endocrine function and blood flow, as well as adipose tissue populations of macrophages and regulatory T-lymphocytes (Treg) were thereafter assessed. Despite similar weight (43.8±1.0 and 40.2±1.5 g, respectively), male but not female mice developed hyperinsulinemia on HFD as previously described (2.5±0.7 and 0.5±0.1 pmol/l, respectively) consistent with glucose intolerance. Male mice also exhibited hypertrophic islets with intact function in terms of insulin release and blood perfusion. Low-grade, systemic inflammation was absent in obese female but present in obese male mice (IL-6 and mKC, males: 77.4±17 and 1795±563; females: 14.6±4.9 and 240±22 pg/ml), and the population of inflammatory macrophages was increased in intra-abdominal adipose tissues of high-fat-fed male but not female mice. In contrast, the anti-inflammatory Treg cell population increased in the adipose tissue of female mice in response to weight gain, while the number decreased in high-fat-fed male mice. In conclusion, female mice are protected against HFD-induced metabolic changes while maintaining an anti-inflammatory environment in the intra-abdominal adipose tissue with expanded Treg cell population, whereas HFD-fed male mice develop adipose tissue inflammation, glucose intolerance, hyperinsulinemia, and islet hypertrophy.

Clinical and Experimental Pancreatic Islet Transplantation to Striated Muscle: Establishment of a Vascular System Similar to that in Native Islets

OBJECTIVE Curing type 1 diabetes by transplanting pancreatic islets into the liver is associated with poor long-term outcome and graft failure at least partly due to inadequate graft revascularization. The aim of the current study was to evaluate striated muscle as a potential angiogenic site for islet transplantation. RESEARCH DESIGN AND METHODS The current study presents a new experimental model that is found to be applicable to clinical islet transplantation. Islets were implanted into striated muscle and intraislet vascular density and blood flow were visualized with intravital and confocal microscopy in mice and by magnetic resonance imaging in three autotransplanted pancreatectomized patients. Mice were rendered neutropenic by repeated injections of Gr-1 antibody, and diabetes was induced by alloxan treatment. RESULTS Contrary to liver-engrafted islets, islets transplanted to mouse muscle were revascularized with vessel densities and blood flow entirely comparable with those of islets within intact pancreas. Initiation of islet revascularization at the muscular site was dependent on neutrophils, and the function of islets transplanted to muscle was proven by curing diabetic mice. The experimental data were confirmed in autotransplanted patients where higher plasma volumes were measured in islets engrafted in forearm muscle compared with adjacent muscle tissue through high-resolution magnetic resonance imaging. CONCLUSIONS This study presents a novel paradigm in islet transplantation whereby recruited neutrophils are crucial for the functionally restored intraislet blood perfusion following transplantation to striated muscle under experimental and clinical situations.

Chronically decreased oxygen tension in rat pancreatic islets transplanted under the kidney capsule.

BACKGROUND A factor of potential importance in the failure of islet grafts is poor or inadequate engraftment of the islets in the implantation organ. This study measured the oxygen tension and blood perfusion in 1-, 2-, and 9-month-old islet grafts. METHODS The partial pressure of oxygen was measured in pancreatic islets transplanted beneath the renal capsule of diabetic and nondiabetic recipient rats with a modified Clark electrode (outer tip diameter 2-6 microm). The size of the graft (250 islets) was by purpose not large enough to cure the diabetic recipients. The oxygen tension in islets within the pancreas was also recorded. Blood perfusion was measured with the laser-Doppler technique. RESULTS Within native pancreatic islets, the partial pressure of oxygen was approximately 40 mm Hg (n=8). In islets transplanted to nondiabetic animals, the oxygen tension was approximately 6-7 mm Hg 1, 2, and 9 months posttransplantation. No differences could be seen between the different time points after transplantation. In the diabetic recipients, an even more pronounced decrease in graft tissue oxygen tension was recorded. The mean oxygen tension in the superficial renal cortex surrounding the implanted islets was similar in all groups (approximately 15 mm Hg). Intravenous administration of glucose (0.1 gxkg(-1)x min(-1)) did not affect the oxygen tension in any of the investigated tissues. The islet graft blood flow was similar in all groups, measuring approximately 50% of the blood flow in the kidney cortex. CONCLUSION The oxygen tension in islets implanted beneath the kidney capsule is markedly lower than in native islets up to 9 months after transplantation. Moreover, persistent hyperglycemia in the recipient causes an even further decrease in graft oxygen tension, despite similar blood perfusion. To what extent this may contribute to islet graft failure remains to be determined.

Islet capillary blood pressure increase mediated by hyperglycemia in NIDDM GK rats.

This study was performed to measure pancreatic islet capillary pressure under basal conditions and after an acute glucose stimulation of insulin release in normal rats. In addition, the islet capillary pressure was estimated in GK rats, an animal model of NIDDM. Hydrostatic pressure in single pancreatic islet capillaries was determined in vivo by direct measurement using the micropuncture technique. The pancreatic islets were visualized by injection of neutral red. This intravital staining had no effect on islet function, whole pancreatic and islet blood flow, and capillary blood pressure in the exocrine pancreas. Islet capillary blood pressure in normoglycemic Wistar F rats was estimated at 3.1 ± 0.3 mmHg (n = 15). Administration of D-glucose (1g/kg) doubled this value, whereas no effect was seen after injection of an equimolar dose of the non-metabolizable glucose-derivative 3-O-methyl glucose. In GK rats, basal islet capillary blood pressure was increased (5.7 ± 0.4 mmHg; n = 10; P < 0.001) when compared with the control Wistar F rats. Reduction of blood glucose levels in GK rats with phlorizin treatment showed this increased basal islet capillary pressure in GK rats to be glucose dependent and reversible. In the present study, we have for the first time shown that both acute and chronic hyperglycemia augment islet capillary pressure. The effects of a chronically increased islet capillary pressure on long-term islet function remain to be determined.

Polyol-pathway-dependent disturbances in renal medullary metabolism in experimental insulin-deficient diabetes mellitus in rats

Aims/hypothesisThe renal medullary region is particularly vulnerable to reduced oxygen concentration because of its low blood perfusion and high basal oxygen consumption. This study investigated renal metabolic changes in relation to the previously observed decreased oxygen tension in streptozotocin-induced diabetic rats.MethodsBlood perfusion, oxygen tension and consumption, interstitial pH, and glycolytic and purine-based metabolites were determined in the renal cortex and the medulla of non-diabetic and diabetic animals by, respectively, laser Doppler flowmetry, oxygen and pH microelectrodes, and microdialysis. The importance of increased polyol pathway activity for the observed alterations was investigated by daily treatment with the aldose reductase inhibitor AL-1576 throughout the course of diabetes.ResultsThe diabetes-induced decrease in renal oxygen tension, due to augmented oxygen consumption, did not result in manifest hypoxia in either the cortical or the medullary region, as evaluated by microdialysis measurements of purine-based metabolites. The profound alterations in medullary oxygen metabolism were, however, associated with an increased lactate : pyruvate ratio and a concomitantly decreased pH. Notably, the renal medullary changes in oxygen tension, oxygen consumption, lactate : pyruvate ratio and pH were preventable by inhibition of aldose reductase.Conclusions/interpretationSubstantial metabolic changes were observed in the renal medulla in diabetic animals. These disturbances seemed to be mediated by increased polyol pathway activity and could be prevented by inhibition of aldose reductase.

Implantation Site–Dependent Dysfunction of Transplanted Pancreatic Islets

OBJECTIVE—Clinical islet transplantations are performed through infusion of islets via the portal vein into the liver. This study aimed at characterizing the influence of the implantation microenvironment on islet graft metabolism and function. RESEARCH DESIGN AND METHODS—Islets were transplanted into their normal environment, i.e., the pancreas, or intraportally into the liver of mice. One month posttransplantation, the transplanted islets were retrieved and investigated for changes in function and gene expression. RESULTS—Insulin content, glucose-stimulated insulin release, (pro)insulin biosynthesis, and glucose oxidation rate were markedly decreased in islets retrieved from the liver, both when compared with islets transplanted into the pancreas and endogenous islets. Islets transplanted into the pancreas showed normal insulin content, (pro)insulin biosynthesis, and glucose oxidation rate but increased basal insulin secretion and impaired glucose stimulation index. Gene expression data for retrieved islets showed downregulation of pancreatic and duodenal homeobox gene-1, GLUT-2, glucokinase, mitochondrial glycerol-phosphate dehydrogenase, and pyruvate carboxylase, preferentially in intraportally transplanted islets. CONCLUSIONS—Islets transplanted into their normal microenvironment, i.e., the pancreas, display gene expression changes when compared with endogenous islets but only moderate changes in metabolic functions. In contrast, site-specific properties of the liver markedly impaired the metabolic functions of intraportally transplanted islets.