Changes In Islet Function Research Articles

Islet hypoplasia of adult offspring rats caused by intrauterine chronic hypoxia is compensated by up-regulation of INS and PDX-1

ABSTRACT Background: Intrauterine chronic hypoxia (ICH) can lead to pancreatic dysmetabolism in offspring. This study aimed to determine the changes in islet function of offspring through a rat ICH model and detect the factors affecting islet function. Methods: Twenty couples of healthy Sprague – Dawley adult rats were randomly mated, and the pregnant rats were randomly allocated to ICH and normal control (NC) groups. Pregnant rats in the ICH group were placed in a hypoxic chamber with 13% oxygen concentration for hypoxia treatment twice a day for 4 h until delivery at 21 days. NC group is inlet with normal air from beginning to end. After delivery, blood was taken from the heart of pregnant rats for blood gas analysis. The weight of the offspring rats was measured at 12 h after birth and 16 weeks after birth. At 16 weeks, the immunohistochemical results of β-cell total, islet area, insulin (INS), and glucose transporter 2 (GLUT2) proteins were obtained from the islets. The mRNA data of INS and pancreatic and duodenal homeobox 1 (PDX-1) genes were obtained from pancreas. Results: We found the β-cell total, islet area, and the positive cell area of INS and GLUT2 of offspring rats in ICH group were lower than those of NC group, while the levels of INS and PDX-1 genes were higher in ICH group than in NC group. Conclusions: ICH can lead to islet hypoplasia in adult male offspring rats. However, this is within the compensatory range.

297-LB: Dorzagliatin, a Glucokinase Activator, Repairs Defective Islet Function in High-Fat Diet-Induced Diabetic Mice

The DREAM study showed long-term benefits for glucose control in newly diagnosed type 2 diabetes (T2D) patients even after withdrawal of dorzagliatin treatment, a newly approved glucokinase activator for T2D treatment, who were considered to be in the remission for diabetes. The purpose of the current study is to examine the mechanisms of dorzagliatin-induced diabetes remission by evaluating the changes in islet function over time after a period of administering dorzagliatin in high-fat diet (HFD) induced obesity and diabetes mouse models. Diabetes mice were generated after 7 months of HFD, then received dorzagliatin or vehicle controls via oral gavage once a day for 19 days (8 days of 30 mg/Kg and 11 days of 15 mg/Kg). After drug withdrawal, one group of mice remained in HFD and another group changed to a normal diet. Islets were then isolated at different time points after drug withdrawal, and the serum and liver were collected for evaluation. The glucose ramp-stimulated insulin secretion (GSIS) was evaluated in islet perifusion. Gene expression was measured by qPCR in islets and by RNA-seq in the liver. Compared to normal mouse islets, HFD-induced diabetic islets in vehicle groups have impaired GSIS with about 40% reduction in maximum insulin secretion. In contrast, GSIS was fully recovered to the levels of normal islets in 10, 18 and 38 days after dorzagliatin withdrawal, but returned to defective GSIS levels after 45 days of drug discontinuation. In addition, islet GSIS can be fully recovered after 80 days of diet change. In conclusion, the defective GSIS in HFD-induced diabetes mouse islets is reversible and can be fully restored by dorzagliatin even after a long period of drug withdrawal while still remaining in HFD. The mechanisms are likely due to increased glucokinase gene expression caused by a short treatment of dorzagliatin. Diet change alone can recover islet function but requires weight reduction and a longer duration. Disclosure D. Han: Employee; Nanjing AscendRare Pharmaceutical Technology. S. Meng: Employee; Nanjing AscendRare Pharmaceutical Technology. X. Zhou: Employee; Nanjing AscendRare Pharmaceutical Technology. R. Li: Employee; Nanjing AscendRare Pharmaceutical Technology. C. Li: Employee; Hua Medicine, Nanjing AscendRare Pharmaceutical Technology.

1645-P: Diabetes-Associated Variation in TCF7L2 Is Associated with a Longitudinal Decline in Glucose Tolerance Accompanied by Impaired Suppression of Postprandial Glucagon Secretion

Diabetes-associated alleles in TCF7L2 increase the rate of conversion from prediabetes to type 2 diabetes (T2DM). Multiple studies have shown an association of the T allele at rs7903146 with impaired β-cell function and more recently with defective suppression of α-cell secretion by glucose. However, the temporal relationship of these abnormalities is uncertain. To study the longitudinal changes in islet function, we recruited 140 individuals half of whom were homozygous for the diabetes-associated allele (TT) at rs7903146. The remainder were homozygous for the protective allele (CC). Subjects were studied on 2 occasions, 3 years apart using an oral 75g glucose challenge. This was labelled with 13C-glucose, in concert with intravenously infused tracers to enable measurement of endogenous glucose production, meal appearance and glucose disposal. Indices for insulin secretion and action were estimated using the oral minimal model. Fasting, peak and integrated (443 ± 27 vs. 428 ± 50 mmol/l per 6hr, p = 0.54) post-prandial glucose concentrations did not differ over time in people with the CC genotype. In contrast, post-prandial integrated glucose concentrations increased (441 ± 29 vs. 544 ± 37 mmol/l per 6hr, p < 0.01) in people with the TT genotype. This was accompanied by an increase in integrated postprandial glucagon concentrations (526 ± 39 vs. 618 ± 61 pmol/l per 2hr, p = 0.04) that was not observed in the CC genotype (537 ± 50 vs. 525 ± 65 pmol/l per 2hr, p = 0.07). There were no baseline differences in β-cell function between genotype groups. Disposition Index in both TT (950 ± 130 vs. 868 ± 92 10-4 dl/kg/min per μU/ml, baseline vs. follow-up study respectively, p=0.42) and CC (896 ± 177 vs. 1062 ± 289 10-4 dl/kg/min per μU/ml, p=0.65) groups did not change. This data would suggest that α-cell dysfunction associated with the TT genotype (rs7903146) precedes β-cell dysfunction in people prior to the development of T2DM. Disclosure M.Zeini: None. K.Muthusamy: None. A.M.Egan: None. M.C.Laurenti: None. C.Cobelli: None. C.Dalla man: Research Support; Sanofi-Aventis Deutschland GmbH, Becton, Dickinson and Company. A.Vella: Advisory Panel; Rezolute, Inc., Consultant; Crinetics Pharmaceuticals, Inc., Hanmi Pharm. Co., Ltd., Zealand Pharma A/S, Other Relationship; Novo Nordisk. Funding National Institutes of Health (DK116723)

Remodeling of the Acetylproteome by SIRT3 Manipulation Fails to Affect Insulin Secretion or β Cell Metabolism in the Absence of Overnutrition

SUMMARYSIRT3 is a nicotinamide adenine dinucleotide (NAD+)- dependent mitochondrial protein deacetylase purported to influence metabolism through post-translational modification of metabolic enzymes. Fuel-stimulated insulin secretion, which involves mitochondrial metabolism, could be susceptible to SIRT3-mediated effects. We used CRISPR/Cas9 technology to manipulate SIRT3 expression in β cells, resulting in widespread SIRT3-dependent changes in acetylation of key metabolic enzymes but no appreciable changes in glucose- or pyruvate-stimulated insulin secretion or metabolomic profile during glucose stimulation. Moreover, these broad changes in the SIRT3-targeted acetylproteome did not affect responses to nutritional or ER stress. We also studied mice with global SIRT3 knockout fed either standard chow (STD) or high-fat and high-sucrose (HFHS) diets. Only when chronically fed HFHS diet do SIRT3 KO animals exhibit a modest reduction in insulin secretion. We conclude that broad changes in mitochondrial protein acetylation in response to manipulation of SIRT3 are not sufficient to cause changes in islet function or metabolism.

Early deficits in insulin secretion, beta cell mass and islet blood perfusion precede onset of autoimmune type 1 diabetes in BioBreeding rats

Aims/hypothesisGenetic studies show coupling of genes affecting beta cell function to type 1 diabetes, but hitherto no studies on whether beta cell dysfunction could precede insulitis and clinical onset of type 1 diabetes are available.MethodsWe used 40-day-old BioBreeding (BB) DRLyp/Lyp rats (a model of spontaneous autoimmune type 1 diabetes) and diabetes-resistant DRLyp/+ and DR+/+ littermates (controls) to investigate beta cell function in vivo, and insulin and glucagon secretion in vitro. Beta cell mass was assessed by optical projection tomography (OPT) and morphometry. Additionally, measurements of intra-islet blood flow were performed using microsphere injections. We also assessed immune cell infiltration, cytokine expression in islets (by immunohistochemistry and qPCR), as well as islet Glut2 expression and ATP/ADP ratio to determine effects on glucose uptake and metabolism in beta cells.ResultsDRLyp/Lyp rats were normoglycaemic and without traces of immune cell infiltrates. However, IVGTTs revealed a significant decrease in the acute insulin response to glucose compared with control rats (1685.3 ± 121.3 vs 633.3 ± 148.7; p < 0.0001). In agreement, insulin secretion was severely perturbed in isolated islets, and both first- and second-phase insulin release were lowered compared with control rats, while glucagon secretion was similar in both groups. Interestingly, after 5–7 days of culture of islets from DRLyp/Lyp rats in normal media, glucose-stimulated insulin secretion (GSIS) was improved; although, a significant decrease in GSIS was still evident compared with islets from control rats at this time (7393.9 ± 1593.7 vs 4416.8 ± 1230.5 pg islet−1 h−1; p < 0.0001). Compared with controls, OPT of whole pancreas from DRLyp/Lyp rats revealed significant reductions in medium (4.1 × 109 ± 9.5 × 107 vs 3.8 × 109 ± 5.8 × 107 μm3; p = 0.044) and small sized islets (1.6 × 109 ± 5.1 × 107 vs 1.4 × 109 ± 4.5 × 107 μm3; p = 0.035). Finally, we found lower intra-islet blood perfusion in vivo (113.1 ± 16.8 vs 76.9 ± 11.8 μl min−1 [g pancreas]−1; p = 0.023) and alterations in the beta cell ATP/ADP ratio in DRLyp/Lyp rats vs control rats.Conclusions/interpretationThe present study identifies a deterioration of beta cell function and mass, and intra-islet blood flow that precedes insulitis and diabetes development in animals prone to autoimmune type 1 diabetes. These underlying changes in islet function may be previously unrecognised factors of importance in type 1 diabetes development.

Multifunctional in vivo imaging of pancreatic islets during diabetes development

Pancreatic islet dysfunction leading to insufficient glucose-stimulated insulin secretion triggers the clinical onset of diabetes. How islet dysfunction develops is not well understood at the cellular level, partly owing to the lack of approaches to study single islets longitudinally in vivo Here, we present a noninvasive, high-resolution system to quantitatively image real-time glucose metabolism from single islets in vivo, currently not available with any other method. In addition, this multifunctional system simultaneously reports islet function, proliferation, vasculature and macrophage infiltration in vivo from the same set of images. Applying our method to a longitudinal high-fat diet study revealed changes in islet function as well as alternations in islet microenvironment. More importantly, this label-free system enabled us to image real-time glucose metabolism directly from single human islets in vivo for the first time, opening the door to noninvasive longitudinal in vivo studies of healthy and diabetic human islets.

Multifunctional in vivo imaging of pancreatic islets during diabetes development.

Pancreatic islet dysfunction leading to insufficient glucose-stimulated insulin secretion triggers the clinical onset of diabetes. How islet dysfunction develops is not well understood at the cellular level, partly owing to the lack of approaches to study single islets longitudinally in vivo Here, we present a noninvasive, high-resolution system to quantitatively image real-time glucose metabolism from single islets in vivo, currently not available with any other method. In addition, this multifunctional system simultaneously reports islet function, proliferation, vasculature and macrophage infiltration in vivo from the same set of images. Applying our method to a longitudinal high-fat diet study revealed changes in islet function as well as alternations in islet microenvironment. More importantly, this label-free system enabled us to image real-time glucose metabolism directly from single human islets in vivo for the first time, opening the door to noninvasive longitudinal in vivo studies of healthy and diabetic human islets.

Short-term glucosamine infusion increases islet blood flow in anesthetized rats

Impaired glucose tolerance and type 2 diabetes in rodents are associated with increased islet blood flow. If this is important for modulation of the endocrine function is at present unknown. We evaluated if glucosamine infusion, which induces peripheral insulin resistance and glucose intolerance, could be used to acutely increase islet blood flow. We infused anaesthetized Sprague-Dawley rats for 2 h with glucosamine (6 mg/kg body weight), in some cases followed by glucose administration. The former induced a 2-fold increase in serum insulin concentrations while plasma glucose remained unchanged. In vitro an augmented insulin response to hyperglycemia and decreased insulin content in batch type islet incubations with glucosamine for 24 h were seen. After 2 h glucosamine exposure in vitro, insulin release was decreased. In vivo glucosamine infusion increased islet blood flow, without affecting other regional blood flow values. Glucose increased islet blood flow to the same extent in control and glucosamine-infused rats. When exposed to 10 mmol/L glucosamine arterioles of isolated perfused islets showed a 10% dilation of their vascular smooth muscle. Thus, application of this model leads to acute hyperinsulinemia in vivo but a decreased insulin release in vitro, which suggests that effects not located to β cells are responsible for the effects seen in vivo. An increased islet blood flow in previously healthy animals was also seen after glucose administration, which can be used to further dissect the importance of blood flow changes in islet function.

Nicotinamide mononucleotide protects against pro-inflammatory cytokine-mediated impairment of mouse islet function

Nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme for NAD(+) biosynthesis, exists as intracellular NAMPT (iNAMPT) and extracellular NAMPT (eNAMPT). eNAMPT, secreted from adipose tissue, promotes insulin secretion. Administration of nicotinamide mononucleotide (NMN), a product of the eNAMPT reaction, corrects impaired islet function in Nampt ( +/- ) mice. One of its potential targets is the NAD(+)-dependent deacetylase sirtuin 1. We hypothesised that altered NAMPT activity might contribute to the suppression of islet function associated with inflammation, and aimed to determine whether NMN could improve cytokine-mediated islet dysfunction. Acute effects of NMN on cytokine-mediated islet dysfunction were examined in islets incubated with TNFα and IL1β, and in mice fed a fructose-rich diet (FRD) for 16 weeks. Changes in iNAMPT, eNAMPT and inflammation levels were determined in FRD-fed mice. FRD-fed mice displayed markedly lower levels of circulating eNAMPT, with impaired insulin secretion and raised islet expression of Il1b. NMN administration lowered Il1b expression and restored suppressed insulin secretion in FRD-fed mice. NMN also restored insulin secretion in islets cultured with pro-inflammatory cytokines. The changes in islet function corresponded with changes in key markers of islet function and differentiation. The anti-inflammatory effects of NMN were partially blocked by inhibition of sirtuin 1. Chronic fructose feeding causes severe islet dysfunction in mice. Onset of beta cell failure in FRD-fed mice may occur via lowered secretion of eNAMPT, leading to increased islet inflammation and impaired beta cell function. Administration of exogenous NMN to FRD-fed mice corrects inflammation-induced islet dysfunction. Modulation of this pathway may be an attractive target for amelioration of islet dysfunction associated with inflammation.

Evidence of functional impairment of syngeneically transplanted mouse pancreatic islets retrieved from the liver.

A drawback in pancreatic islet transplantation is the large number of islets needed to obtain insulin independence in patients with diabetes. This most likely reflects extensive posttransplantation islet cell death and functional impairment of the remaining endocrine cells. We aimed to develop an experimental method to retrieve transplanted islets from the mouse liver, which would enable comparisons of transplanted and endogenous islets and provide valuable information on functional changes induced by intraportal transplantation. Transplanted islets were obtained by retrograde perfusion of the liver with collagenase. The identity of retrieved tissue as transplanted islets was confirmed by intravital staining, immunohistochemistry, and electron microscopy. The retrieved islets, irrespective of whether they had resided in diabetic or nondiabetic recipients, had a markedly lower insulin content and glucose-stimulated insulin release when compared with isolated endogenous islets. The glucose oxidation rate was also markedly lower in the retrieved islets, suggesting mitochondrial dysfunction. These disturbances in insulin content, insulin release, and glucose oxidation rate were not reversed by a few days of culture after retrieval. The results implicate changes in islet function after intraportal transplantation. Such dysfunction may contribute to the high number of islets needed for successful transplantation in diabetic individuals.

Accurate measurement of endogenous insulin secretion does not require separate assessment of C-peptide kinetics.

The implication of beta-cell failure as an early defect in type 2 diabetes exacerbates the need for accurate but facile assessment of islet cell secretory rate, particularly in large group studies in which individual assessment of C-peptide kinetics is impractical. This study was designed to examine whether it is possible to obtain accurate secretory rates from the extended combined model, which provides insulin and C-peptide kinetics from plasma measurements of the two peptides. Equimolar intraportal infusions of insulin and C-peptide that are designed to simulate insulin secretion rates during both oral and intravenous glucose tolerance tests were used to generate plasma insulin and C-peptide data in conscious dogs that were examined under clamped glucose conditions. The plasma peptide kinetics were analyzed using the extended combined model to generate estimates of prehepatic insulin secretion that were then compared with the known intraportal infusion rates. The extended combined model was able to reproduce the known intraportal infusion profiles. The model-predicted rates were similar to those calculated with methods that require separate assessment of C-peptide kinetics. Simulation results supported lesser clearance of insulin during rapid changes of portal insulin (as measured by an intravenous glucose tolerance test) versus slow changes in portal insulin (as measured by an oral glucose tolerance test). The extended combined model accurately calculates prehepatic insulin appearance. It may be possible to apply this approach to large studies of beta-cell function designed to identify changes in islet function in subjects at risk for diabetes. Such an approach could strengthen epidemiological and genetic studies of the pathogenesis of diabetes.

Osmotic tolerance limits of canine pancreatic islets.

Future improvements in the recovery and function of pancreatic islets following cryopreservation will require a more precise quantification of the stresses that occur at each stage of the cryopreservation protocol. Changes in solution osmolality during the addition and dilution of cryoprotectants and during freezing and thawing induce changes in islet volume that may exceed tolerable limits. The aim of this study was to determine the range of solution osmolalities that results in significant changes in islet function. Islets were isolated from canine pancreases by collagenase digestion and Euro-Ficoll purification. Following 12-h culture at 37 degrees C, islets were counted and dispensed into multiwell plate inserts. Islet function was assessed in each well immediately before and 24 h following a 10-min osmotic challenge with hypo- or hyperosmotic solutions of PBS (0, 75, 150, 300, 600, 1200, or 2300 mOsm/kg) at 22 degrees C. Canine islets reached their osmotic equilibrium within 10 min. Duplicate wells were used for each osmolality treatment for each of six donors (n = 12). No significant differences in basal or glucose-stimulated insulin secretion were found between wells prior to the osmotic challenge (3.35 +/- 0.45 and 20.98 +/- 3.36 microIU/IE/h, respectively). Following the osmotic challenge and 24-h in vitro tissue culture, a significant increase in basal secretion was observed for islets exposed to 0 and 75 mOsm/kg solutions and a significant decrease for islets exposed to 2300 mOsm/kg solution. Islets exposed to 0 and 2300 mOsm/kg solutions showed significant decreases in the stimulated insulin secretion when compared to controls. Solution osmolalities of 150-1200 mOsm/kg appear to be tolerated by canine islets with no significant deviations in insulin secretion. The corresponding tolerable volume range was 152.6 +/- 6.8% to 60 +/- 5.1% of the isotonic islet volume. The minimum critical volume was used in a theoretical analysis of the islet volumes that would result from equilibrium freezing in dimethyl sulfoxide (DMSO). The calculations show that 1.5 mol/l DMSO is sufficient to prevent damage to islets due to excessive shrinkage. Further refinement of cryoprotectant addition and dilution protocols, and cooling and warming protocols for canine islets, are now possible.

Effect of tyrosine kinase inhibitors on islets of Langerhans: evidence for tyrosine kinases in the regulation of insulin secretion

In order to determine if tyrosine kinase activation is involved in the changes in islet function, the effect of tyrosine kinase inhibitors on insulin secretion and islet cell proliferation was examined in cultured islets of Langerhans. When islets were exposed to 100 microM genistein or 2 microM herbimycin A, large 5- to 10-fold increases in insulin secretion were observed. The effect on insulin secretion was detected within 1 hr and was maintained for at least 4 days. The glucose sensitivity of islets exposed to genistein was dramatically increased as demonstrated by a shift of the glucose-dose response curve to lower glucose concentrations. In contrast, islet cell proliferation was dramatically reduced in the presence of these tyrosine kinase inhibitors in the absence or presence of PRL. These very large changes observed in islets suggest that tyrosine kinases may have important roles in the regulation of beta-cell function.

Effect of tyrosine kinase inhibitors on islets of Langerhans: evidence for tyrosine kinases in the regulation of insulin secretion.

In order to determine if tyrosine kinase activation is involved in the changes in islet function, the effect of tyrosine kinase inhibitors on insulin secretion and islet cell proliferation was examined in cultured islets of Langerhans. When islets were exposed to 100 microM genistein or 2 microM herbimycin A, large 5- to 10-fold increases in insulin secretion were observed. The effect on insulin secretion was detected within 1 hr and was maintained for at least 4 days. The glucose sensitivity of islets exposed to genistein was dramatically increased as demonstrated by a shift of the glucose-dose response curve to lower glucose concentrations. In contrast, islet cell proliferation was dramatically reduced in the presence of these tyrosine kinase inhibitors in the absence or presence of PRL. These very large changes observed in islets suggest that tyrosine kinases may have important roles in the regulation of beta-cell function.

Adaptation of islets of Langerhans to pregnancy: increased islet cell proliferation and insulin secretion correlates with the onset of placental lactogen secretion.

To elucidate the temporal profile of adaptive changes of the islets of Langerhans to the increased insulin demands of pregnancy, we have studied islet cell proliferation and insulin secretion during gestation in the rat. 5-Bromo-2'-deoxyuridine incorporation into dividing islet cells was significantly (P less than 0.05) increased over age-matched controls by day 10, rose continuously to a peak at day 14, and then returned to control levels by day 18. By day 20, cell division was significantly inhibited (P less than 0.05). The pattern of changes in insulin secretory profiles observed with perfused pancreata of pregnant animals was similar to that obtained for islet cell proliferation. Both the threshold of glucose-stimulated insulin secretion and the amount of above threshold insulin secretion began to diverge from controls by day 10. By day 12, the glucose-stimulation threshold was significantly decreased from 5.7 mM glucose to 3.3 mM (P less than 0.05), remained at this low level through day 15, and returned toward normal by day 20. Concomitant with the increased sensitivity of B cells to glucose, the above threshold insulin secretion was significantly increased by day 12 (P less than 0.05), peaked at day 15, and returned to control levels by day 20. This insulin secretory data demonstrates that the increased sensitivity of B cells to glucose is an important component of the adaptation of islets during pregnancy to the increased demand for insulin at physiological concentrations of plasma glucose. To correlate the above changes in islet cell proliferation and insulin secretion with levels of placental lactogen (PL), serum lactogenic hormone activity was measured by Nb2 lymphoma cell replication assays. This analysis revealed the expected biphasic pattern: a midpregnancy peak at day 12, followed by a nadir at day 14, and then continuously elevated levels until term. The bioassay data agreed with the known secretory profiles of rat (r) PL-I (midpregnancy) and rPL-II (late pregnancy). Our results provide the first systematic evaluation of changes in islet function during pregnancy in the rat. In addition, they provide evidence that rPL-I may be the critical hormonal signal which triggers the primary adaptive changes in islet function characteristic of pregnancy. The return to normal values of insulin secretion and inhibition of cell division observed at day 20 in the presence of high concentrations of rPL-II suggests that other inhibitory influences become dominant in the later stages of rat pregnancy.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of a diabetogenic strain of encephalomyocarditis (EMC) virus on protein synthesis in mouse islets of Langerhans

The effects of a diabetogenic strain of encephalomyocarditis (EMC) virus on total protein and insulin biosynthesis in mouse islets of Langerhans have been studied in tissue culture. In dispersed mouse islets, the rates of protein biosynthesis were assessed by measuring the incorporation of [3H]leucine into proteins. In infected dispersed islets incubated in 20 mM-glucose, both insulin and total protein biosynthesis were decreased at 6 h; only insulin biosynthesis was significantly decreased at 3 h. In whole islets, EMC virus brought about a decrease in glucose-stimulated protein and insulin biosynthesis as early as 2 h after infection without concomitant effects on insulin release. This inhibition of protein biosynthesis was still apparent at 20 h post-infection, at which time insulin release was found to be markedly elevated, and the islet insulin content was moderately decreased. At 44 h post-infection, glucose-induced insulin biosynthesis was preferentially inhibited. Infected islets at this later time point also displayed elevated levels of insulin release, and a marked loss of islet insulin content. When insulin mRNA and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA levels were assessed by dot-blot hybridization using appropriate cDNA probes, levels of insulin mRNA were shown to decrease steadily during the first 20 h of infection, in contrast with the levels of GAPDH mRNA. At 44 h post-infection, both types of mRNA were markedly decreased. It is suggested that there is an initial early 'shut-off' of protein synthesis without other detectable changes in islet function. This is followed by a phase where both insulin mRNA levels and insulin synthesis are dramatically decreased.

THE EFFECT OF DUCT OBLITERATION ON THE HISTOLOGY AND ENDOCRINE FUNCTION OF THE CANINE PANCREAS

Although duct obliteration is a safe and effective method for ablation of exocrine secretion in segmental pancreas transplantation, it remains to be clarified whether its effects are restricted to the exocrine tissue. In 20 dogs (beagles 9-15 kg) the right lobe of the pancreas was removed and the ductal system of the left lobe was injected with the duct-obliterants neoprene (6 dogs), polyisoprene (6 dogs), or prolamine (8 dogs). In this study, i.v. glucose tolerance tests (the results of which are expressed in K values) and relaparotomies for taking biopsies were performed at 1, 3, and 12 months after duct obliteration. Biopsies were studied histologically and immunohistochemically in a qualitative and semiquantitative fashion. Three prolamine-injected dogs developed diabetes. All other dogs maintained normal fasting blood glucose levels but showed reduced K values at 1 month after duct obliteration. Further deterioration of glucose tolerance was not observed up to 12 months. Differences in K values depending on the type of obliterant were insignificant at all intervals. The exocrine tissue was completely replaced by fibrosis at 3 months after duct obliteration, and the architecture of the islets was disrupted. Morphometrical analysis of relative numbers of different endocrine cell types showed transient changes at 1 month after duct obliteration, but did not differ from unmodified controls at 12 months. We conclude that the effects of duct obliteration are not restricted to the exocrine pancreatic tissue, but that the endocrine pancreas is interfered with as well. Changes in islet function and histology are brought about during the first month after duct obliteration and stabilize thereafter.