Pancreatic cancer is known to be one of the most life-threatening cancers characterized by aggressive local invasion and distant metastasis. The high basal level of autophagy in pancreatic cancer may be responsible for the low chemotherapeutic drug response rate and poor disease prognosis. However, the clinical application of autophagy inhibitors was unsatisfactory due to their toxicity and minimal single-agent anticancer efficacy. Hence, oncologists begin to consider the tumor microenvironment when exploring new drug targets. In the present study, the anti-tumorigenic mechanisms of two major phytochemicals derived from Chinese medicinal herbs had been investigated against pancreatic cancer development. Calycosin is a bioactive isoflavonoid of the medicinal plant Astragalus membranaceus. Our results have shown that calycosin inhibited the growth of various pancreatic cancer cells both in vitro and in vivo by inducing cell cycle arrest and apoptosis. Alternatively, calycosin also facilitated MIA PaCa-2 pancreatic cancer cell migration in vitro and increased the expression of epithelial-mesenchymal transition (EMT) biomarkers in vivo. Further mechanistic study suggests that induction of the Raf/MEK/ERK pathway and facilitated polarization of M2 tumor-associated macrophage in the tumor microenvironment both contribute to the pro-metastatic potential of calycosin in pancreatic cancer. These events appear to be associated with calycosin-evoked activation of TGF-β signaling, which may explain the paradoxical drug actions due to the dual roles of TGF-β as both tumor suppressor and tumor promoter in pancreatic cancer development under different conditions. Isoliquiritigenin (ISL) is a chalcone obtained from the medicinal plant Glycyrrhiza glabra, which can be a precursor for chemical conversion to form calycosin. Results have shown that ISL decreased the growth and EMT of pancreatic cancer cells in vitro, probably due to modulation of autophagy. ISL-induced inhibition of autophagy subsequently promoted reactive oxygen species (ROS) production, leading to induction of apoptosis in pancreatic cancer cells. Such phenomenon also contributed to the synergistic growth-inhibitory effect in combined treatment with the orthodox chemotherapeutic drug 5-fluorouracil. In addition, ISL-induced tumor growth inhibition in vivo was further demonstrated in a tumor xenograft mice model of pancreatic cancer. ISL promoted apoptosis and inhibited autophagy in the tumor tissues. Study on immune cells indicates that ISL could reduce the number of myeloid-derived suppressor cells (MDSCs) both in tumor tissue and in peripheral blood, while CD4+ and CD8+ T cells were increased correspondingly. In vitro test has revealed that ISL inhibited the polarization of M2 macrophage along with its inhibition of autophagy in M2 macrophage. These immunomodulating effects of ISL had reversed the pro-invasive role of M2 macrophage in pancreatic cancer.In conclusion, calycosin acts as a "double-edged sword" on the growth and metastasis of pancreatic cancer, which may be related to the dual roles of TGF-β and its influence on the tumor microenvironment. Alternatively, ISL consistently inhibited the growth and metastatic drive of pancreatic cancer through regulation of autophagy and reprogramming of the immune system. The differential modes of action of these compounds have provided new insights in the development of effective pancreatic cancer treatment adjuvants.

Indiana University-Purdue University Indianapolis (IUPUI) / Pancreatic ductal adenocarcinoma (PDAC), which comprises 85% of pancreatic cancers, is the 4th leading cause of cancer death in the United States with a 5-year survival rate of 8%. While human PDACs (hPDACs) are hypovascular, they also overexpress a number of angiogenic growth factors and receptors. Additionally, the use of anti-angiogenic agents in murine models of PDAC leads to reduced tumor volume, tumor spread, and microvessel density (MVD), and improved survival. Nonetheless, clinical trials using anti-angiogenic therapy have been overwhelmingly unsuccessful in hPDAC. On the other hand, pancreatic neuroendocrine tumors (PNETs) account for only 2% of pancreatic tumors, yet they are very vascular and classically angiogenic, respond to anti-angiogenic therapy, and confer a better prognosis than PDAC even in the metastatic setting. In an effort to compare and contrast the angiogenic transcriptomes of these two tumor types, we analyzed RNA-Sequencing (RNA-Seq) data from The Cancer Genome Atlas (TCGA) and found that a pro-angiogenic gene signature is present in 35% of PDACs and that it is mostly distinct from the angiogenic signature present in PNETs. The pro-angiogenic PDAC subgroup also exhibits a transcriptome that reflects active TGF-β signaling, less frequent SMAD4 inactivation than PDACs without the signature, and up-regulation of several pro-inflammatory genes, including members of JAK signaling pathways. Consequently, targeting the TGF-β receptor type-1 kinase with SB505124 and JAK1/2 with ruxolitinib blocks proliferative crosstalk between human pancreatic cancer cells (PCCs) and human endothelial cells (ECs). Additionally, treatment of the KRC (oncogenic Kras, homozygous deletion of Rb1) and KPC (oncogenic Kras, mutated Trp53) genetically engineered PDAC mouse models with ruxolitinib suppresses murine PDAC (mPDAC) progression only in the KRC model, which shows superior enrichment and differential expression of the human pro-angiogenic gene signature as compared to KPC tumors. These findings suggest that targeting both TGF-β and JAK signaling in the 35% of PDAC patients whose cancers exhibit an pro-angiogenic gene signature should be explored in a clinical trial.

Orientador: Débora Cristina Damasceno / Coorientador: Yuri Karen Sinzato / Banca: Mara Sandra Hoshida / Banca: Luciane Dias Alargão Dias Melicio / Resumo: Introdução:Muitos estudos mostram a regeneração pancreática em um ambiente diabético e não exploram este processo no organismo saudável. Para compreender melhor os mecanismos envolvidos na regeneração das ilhotas pancreáticas sem o envolvimento do diabete. Objetivo:Avaliar a estrutura morfológicadas ilhotas pancreáticas desde o período neonatal até a idade adulta de ratas. Metodologia:Recém-nascidos do sexo feminino foram distribuídos aleatoriamente em diferentes grupos de estudo para coleta do pâncreas e soro em diferentes períodos de vida (5, 15, 90 dias de vida e no 18,5º dia de prenhez). Para a análise histopatológica, foram considerados os aspectos morfológicos observados na coloração de hematoxilina e eosina (HE). A análise morfométrica foi realizada em sistema computadorizado de imagem (software KS-300, versão 3.0, Zeiss). Para análise imunoistoquímica, foram utilizados os anticorpos anti insulina, anti PDX-1, anti GLP-1, anti Ki-67 e anti caspase-3. A avaliação imunoistoquímica foi realizada por contagem de células positivas através da grade de Weibel modificada. Resultados:Com cinco dias de vida, foi observado menor área comparado a dos outros períodos. Na fase adulta e durante a prenhez, as ratas apresentaram diminuição do número de células positivas para insulina e aumento de células quiescentes comparado às dos dias 5 e 15 de vida pós-natal. Um pico de proliferação celular foi observado no 15°dia de vida e, em todos os momentos estudados, foi observado maior frequência de morte celular das células não-beta (β) das ilhotas pancreáticas (células α, δe PP) em relação às células βpancreáticas. Durante a prenhez, houve diminuição do número de células positivas para GLP-1 em relação aos dias 5 e 15 de vida. Conclusão:As ilhotas pancreáticas apresentaram mudançasem sua morfologia, sobretudo durante os primeiros 15 dias de vida, sendo estas mudanças suficientes para ... / Abstract: Not available / Mestre

The human pancreatic cancer cell line T3M4, is known to produce transforming growth factor-alpha (TGF-alpha); as well as overexpress the receptor for this ligand, epidermal growth factor (EGF) receptor. TGF-alpha messenger RNA (mRNA) levels were assayed using northern blot, after addition of epidermal growth factor or TGF-alpha. The level of TGF-alpha mRNA was found to increase 2-fold at 2 hours and then return to near basal levels at 10 hours, after treatment with either ligand. Both ligands were also equipotent in a 2 hour dose response assay, with half maximal stimulation seen at 1 nM and maximal stimulation reached at 4 nM. Furthermore, there appeared to be a 2-fold increase in TGF-alpha transcription as determined by nuclear runoff experiments. Induction of TGF-alpha mRNA coupled with the overexpression of the EGF receptor, may result in a potent autocrine cycle; which may be found in other cancers.

Background: Metastases to the pancreas are rare, accounting for less then 2 % of all pancreatic malignancies. However, both the benefit of extended tumor resection and the ideal oncological approach have not been established for such cases; therefore, we evaluated patients with metastasis to the pancreas who underwent pancreatic resection. Methods: Between 1994 and 2012, 676 patients underwent pancreatic surgery in our institution. We retrospectively reviewed patients’ medical records according to survival, and surgical and non-surgical complications. Student’s t-test and the log-rank test were used for statistical analysis. Results: Eighteen patients (2.7 %) received resection for pancreatic metastases (12 multivisceral resections and 6 standard resections). The pancreatic metastases originated from renal cell carcinoma (n = 10), malignant melanoma (n = 2), neuroendocrine tumor of the ileum (n = 1), sarcoma (n = 1), colon cancer (n = 1), gallbladder cancer (n = 1), gastrointestinal stromal tumor (n = 1), and non-small cell lung cancer (n = 1). The median time between primary malignancy resection to metastasectomy was 83 months (range, 0–228 months). Minor surgical complications (Grade I-IIIa) occurred in six patients (33.3 %) whereas major surgical complications (Grade IIIb-V) occurred in three patients (16.6 %). No patients died during hospitalization. The median follow-up was 76 months (range, 10–165 months). One-year, 3-year and 5-year survival for standard resection versus multivisceral resection was 83, 50, and 56 % versus 83, 66, and 50, respectively. Twelve patients died after a median of 26 months (range, 5–55 months). Conclusions: A surgical approach with curative intent is justified in select patients suffering from metastases to the pancreas and offers good long-term survival. The resection of pancreatic metastases of different tumor types was associated with favorable morbidity and mortality when compared with resection of the primary pancreatic malignancies. Our findings also demonstrated that multivisceral resection was feasible, with acceptable long term outcomes, even though morbidity rates tended to be higher after multivisceral resection than after standard resection.

Dissertation (PhD)--Stellenbosch University, 2004. / ENGLISH ABSTRACT: Introduction Despite advances in insulin therapy, the side effects associated with diabetes mellitus still remain. Pancreas transplantation has benefited diabetics with end-stage renal failure by reversing the diabetic state and preventing or reversing the progression of diabetes associated diseases. Currently the side effects associated with lifelong immunosuppression preclude pancreas transplantation as a viable treatment option for both type I and II diabetics. In the laboratory, transplanted rat foetal pancreata have been shown to be able to reverse the clinical signs of streptozotocin-induced diabetes in an isogeneic model. Reversal of diabetes by allogeneic foetal rat pancreas transplantation, although possible has proved to be more difficult due to fierce rejection of the grafts and the diabetogenic effects of conventional immunosuppressants. Aims One of the goals, focus and intentions of this laboratory study in rodents, is to contribute new information to the scientific literature. The potential to “reverse” the diabetic state by allogeneic foetal pancreatic transplantation, was the main stimulus for this study. Methods Foetal pancreata of 16-18 days gestation were transplanted into a surgically prepared renal subcapsular space. Immunosuppressive protocols used to prevent rejection of the allogeneic foetal rat pancreata included donor specific transfusion (DST), cyclosporine [a calcineurin inhibitor (CsA)], mycophenolate mofetil [a purine syntase inhibitor (MMF)], and a mouse anti-rat CD4 monoclonal antibody (W3/25). Immunosuppressants were used as monotherapies and in combination. Results Isogeneic foetal rat pancreas transplantation resulted in the growth and development of mature insulin producing islets of Langerhans at the site of engraftment. Allogeneic foetal pancreatic transplantation without immunosuppression resulted in complete rejection of the grafts at 14 days post-transplantation. Histological assessment of allografts at 14 and 30 days post-transplantation showed that CsA was able to prevent acute rejection in our rat models although graft scores and survival were improved if CsA was combined with MMF. Intraperitoneal anti-CD4 monoclonal injections were well tolerated, and if given daily effectively prolonged graft survival up to 30 days. Combining DST with anti-CD4 and CsA induction therapy provided long-term graft survival without daily immunosuppression. This combination, together with allogeneic foetal rat pancreas transplantation, was effective in reversing the clinical signs of experimentally induced diabetes. To my knowledge these are the first published results in which reversal of streptozotocin induced diabetes was achieved by fully MHC mismatched foetal rat pancreatic transplantation. Conclusion Foetal rat pancreatic transplantation is a potential source of endocrine replacement, which, with effective immunosuppression allows for the development of functional islets able to reverse the clinical signs of experimentally induced diabetes in an allogeneic rat model. An unique immunosuppressive protocol, with potential clinical relevance in the human, combines anti-CD4 mAb, CsA and DST induction therapy, which alleviates the burden of daily immunosuppression and associated side effects. / AFRIKAANSE OPSOMMING: Inleiding Ten spyte van die vordering met modeme insulienterapie bly die newe-effekte, waarmee diabetes mellitus geassosieer is, steeds ‘n probleem vir diabete. Diabetiese pasiente met eindstadium nierversaking trek geweldig voordeel uit nier-pankreasoorplantings wat die diabetes omkeer en die progressie van diabetesverwantesiektes voorkom of selfs omkeer. Die newe-effekte van lewenslange immuunonderdrukking skakel pankreasoorplanting uit as ‘n lewensvatbare behandelingsopsie vir tipe I of II diabete. In ‘n streptozotosien-gei'nduseerde diabetiese rotmodel kan isogenei'ese fetale pankreasoorplanting die kliniese tekens van diabetes omkeer. Die omkering van streptozotosien-gei'nduseerde diabetes deur allogeneiese fetale pankreasoorplanting behoort moontlik te wees indien verwerping en die diabetogeniese newe-effekte van konvensionele immuunonderdrukkers oorkom word. Doelstellings Een van die mikpunte, fokusse en oogmerke van hierdie laboratorium studie in knaagdiere, is om ‘n betekenisvolle bydrae tot nuwe kennis in die wetenskaplike literatuur, te maak. Die potensiaal om die diabetiese toestand deur allogeneiese fetale pankeasoorplanting om te keer, was die hoof stimulus vir die studie. Metodes Fetale rotpankreata van 16-18 dae gestasie was in ‘n chirurgies voorbereide spasie onder die nierkapsel oorgeplant. Immuunonderdrukkende protokolle, vir die voorkomming van verwerping van die allogeneiese fetale pankreasoorplantings, het donorspesifiekeoortappings (DST), siklosporien [‘n kalsineurien inhibitor (CsA)], mikofenolaat mofetiel [‘n purien sintase inhibitor (MMF)] en ‘n anti-rot CD4 monoklonale antiliggaam (W3/25) ingesluit. Die immuunonderdrukkers is as mono- of as kombinasieterapie gebruik. Resultate IsogeneTese fetale rotpankreasoorplanting het tot die ontwikkeling van volwasse insulienproduseerende eilande van Langerhans gelei, wat die kliniese tekens van streptozotosien-gei'nduseerde diabetes kon omkeer. Allogenei'ese fetale rotpankreasoorplanting sonder immuunonderdrukking het tot algehele verwerping van die oorplanting binnel4 dae na oorplanting gelei. Histologiese beoordeling van die oorplantings 14 en 30 dae na oorplanting het getoon dat CsA akute verwerping van fetale pankreasoorplantings in die rotmodelle voorkom. Indien CsA met MMF gekombineer word, word die oorplantings-telling en oorlewing verbeter. Intraperitoneale anti-CD4 monoklonale inspuitings was goed verdra, en indien daagliks toegedien, het dit die oorlewing van die pankreasoorplantings effektief tot 30 dae verleng. Die kombinasie van DST, anti-CD4 en CsA induksieterapie het tot langtermyn oorlewing van die pankreasoorplantings gelei sonder verdere daaglikse immuunonderdrukking. Die induksieterapie in kombinasie met allogenei'ese fetale pankreasoorplanting was effektief in die omkering van die kliniese tekens van streptozotosien-gei'nduseerde diabetes in die rot. Hierdie is, sover ek weet, die eerste keer dat omkering van streptozotosien-gei'nduseerde diabetes suksesvol met ‘n volledige MHC onverenigbare allogenei'ese fetale pankreasoorplanting behaal is. Gevolgtrekkings Fetale rotpankreasoorplanting is ‘n potensiele bron vir endokrien vervangingsterapie, wat met effektiewe immuunonderdrukking tot die ontwikkeling van funksionele eilande van Langerhans lei, wat die vermoe het om die kliniese tekens van experimenteel-ge'induseerde diabetes in ‘n allogeneiese rotmodel om te keer. ‘n Unieke immuunonderdrukkingsprotokol, met kliniese relevansie, kombineer DST met anti-CD4 mAb en CsA induksieterapie wat die las van daaglikse immuunonderdrukking en die geassosieerde newe-effekte van konvensionele immuunonderdrukking verlig.

Thesis (PhD)--Stellenbosch University, 2005. / ENGLISH ABSTRACT: A high fat diet (HFD) reduces beta-cell mass, impairs glucose signalling and is involved in the development of Type 2 diabetes. Malnutrition during gestation is hypothesized to irreversibly damage beta-cell development. The transcription factors Pdx-1 and Pax 4 are involved in islet cell development. Pdx-1 is reported to regulate expression of GLUT-2, glucokinase (GK) and the insulin gene. Aims The aim of this study is to investigate, in the neonatal and weanling rat, the effect of exposure to a HFD in utero and/or lactation on weight, glucose and insulin concentrations, islet cell development, pancreatic transcription factors and glucose sensing genes. Methods Neonatal and weanling rats were exposed to a maternal HFD for defined periods of gestation and/or lactation. After termination, pups were weighed and glucose and insulin concentrations determined. mRNA expression of Pdx-1, Pax 4, GLUT-2 and GK was quantified by LightCycler PCR. Pancreatic sections were immunostained for insulin and glucagon (islet cell development), and for Pdx-1, GLUT-2 and GK (beta-cell function) followed by image analysis. Results: Exposure to an in utero HFD throughout gestation resulted in hyperglycaemic pups with reduced beta-cell volume and number, Pdx-1 and GK immunoreactivity. In contrast the alpha-cell volume, number and size were augmented in neonates exposed to a HFD throughout gestation. Most weanlings were hyperglycaemic and hypoinsulinaemic. In some weanlings, reduced beta-cell number and beta- and alpha-cell size was observed. Pdx-1 mRNA was overexpressed in weanlings exposed to a maternal HFD for the final week of gestation or throughout both gestation and lactation, but reduced in those only exposed throughout lactation. Pax 4 mRNA was reduced in weanlings exposed to a maternal HFD for the first or final week of gestation, throughout gestation or throughout lactation. In most of the weanlings, GLUT-2 mRNA expression was reduced whereas immunoreactivity for GLUT-2 was increased. Both GK mRNA expression and immunoreactivity were reduced in most of the weanlings. Conclusions • Exposure to an in utero HFD throughout gestation induced hyperglycaemia in neonates. The reduced Pdx-1 expression appears to play a role in the compromised beta-cell development, and concomitant with the reduced GK levels, contributes to the hyperglycaemia in these neonates and may make them susceptible to beta-cell failure. • In most weanlings exposed to a HFD in utero and/or during lactation the hyperglycaemia and hypoinsulinaemia suggest compromised beta-cell function. The GK mRNA expression and immunoreactivity were reduced thereby impairing glycolysis which would result in reduced insulin secretion contributing to the hyperglycaemia. Furthermore, beta-cell development is adversely affected by the HFD in some weanlings. This would contribute to reduced beta-cell function and may eventually result in beta-cell failure. GLUT-2 immunoreactivity was increased in some, suggesting a compensatory adaptative mechanism to restore glucose homeostasis. • A maternal HFD has adverse effects both in neonates and weanlings on beta-cell development, transcription factor and glucose sensing gene expression and induced hyperglycaemia and hypoinsulinaemia in some of the offspring. Ways to ameliorate the HFD-induced attenuation of key beta-cell genes to ensure normal beta-cell function are important for future research in Type 2 diabetes. / AFRIKAANSE OPSOMMING: ‘n Hoe vet diet (HVD) verminder beta-sel masse, glukose signale en speel ‘n rol in Tipe 2 diabetes. Die hipothese is dat wanvoeding gedurende swangerskap lei tot onomkeerbare betasel beskadiging. Die transkripsiefaktore Pdx-1 en Pax 4 speel rolle in eilandselontwikkeling. Daar is bewyse dat Pdx-1 die uitdrukking van die GLUT-2, glucokinase (GK) en insulin gene reguleer. Doelstelling: Die doel van hierdie studie is om, in die pasgebore en gespeende rot, die effek van ’n HVD in utero en/of laktasie op gewig, glukose en insulin konsentrasies, eilandselontwikkeling, pankreatiese transksripsiefaktore en op glukosewaarnemingsgene te ondersoek. Metodes: Pasgebore en gespeende rotte is vir bepaalde periodes van gestasie en/of laktasie blootgestel aan ’n HVD van die moeder. Na terminase, is kleinjies geweeg en die glukose- en insulienkinsentrasies bepaal. mRNA uitdrukking van Pdx-1, Pax 4, GLUT-2 en GK is geoes met LightCycler PCR. Snitte van die pankreas is gekleur met insulien en glukagon (eilandsontwikkeling) en vir Pdx-1, GLUT-2 en GK (betaselfunksie) gevolg deur beeldanalise. Resultate: Bloodstelling aan ’n in utero HVD regdeur gestasie het hiperglisemie versoorsaak in pasgebore rotte met verlaagde betasel volume en aantal, Pdx-1 en GK immunoreaktiwiteit. In teenstelling daarmee was die alfasel se volume, aantal en grootte verhoog in pasgebore rotte wat regdeur gestasie aan 'n HVD blootgestel was. Meeste van die gespeende rotte was hiperglisemies en hipoinsulinemies. In sommige gespeende rotte, was daar ’n verlaging van betasel hoeveelheid en grootte en in alfasel groote. Oormatige uitdrukking van Pdx-1 mRNA het plaasgevind in speenlinge wat aan ’n HVD van die moeder vir die laaste week van gestasie of regdeur gestasie en laktasie blootgestel was, maar dit was verlaag in die speenlinge wat net tydens laktasie blootgestel was. Pax 4 mRNA was verlaag in speenlinge wat aan ’n HVD van die moeder blootgestel was vir die eerste of laaste week van gestasie, regdeur gestasie of regdeur laktasie. In meeste van die speenlinge is onder-uitdrukking van GLUT-2 mRNA, maar ’n verhoging van GLUT-2 immunoreaktiwiteit gevind. Beide GK mRNA uitdrukking en immunoreaktiwiteit was laer in meeste van die speenlinge. Gevolgtrekkings: • Blootstelling aan ’n in utero HVD regdeur gestasie lei tot hiperglisemie in pasgebore rotte. Die verlaagde Pdx-1 immunoreaktiwiteit speel klaarblyklik ’n rol by die geaffekteerde betaselontwikkeling. Dit, saam met die verlaagde immunoreaktiwiteit vir GK, kan bydra tot die hiperglisemie in hierdie pasgebore rotte. In die meeste van die speenlinge wat aan ’n HVD blootgestel was, dui die hiperglisemie en hipoinsulinemie op geaffekteerde betaselfunksie. Die GK mRNA uitdrukking en immunoreaktiwiteit is verlaag, wat weer glikolise benadeel, en dit sal lei tot verminderde insulienafskeiding wat bydra tot die hiperglisemie. Betaselontwikkeling word voorts negatief beinvloed deur die HVD, wat blyk uit die verlaagde aantal en grootte van betaselle. Dit sal bydra tot verminderde betaselfunksie. Dit kan uiteindelik tot betaselversaking lei. GLUT-2 immunoreaktiwiteit was verhoog in hierdie speenlinge, wat dui op ’n kompenserende aanpassingsmeganisme om glukose homeostase te herstel. ’n HVD van die moeder het ’n negatiewe uitwerking op betaselontwikkeling, transkripsiefaktor en glukosewaarneming geenuitdrukking in beide die pasgebore en gespeende rotte, en geinduseerde hiperglisemie en hipoinsulinemie in sommige kleintjies. Dis belangrik vir toekomstige Tipe 2 diabetes navorsing dat daar na gekyk moet word om die HVD-geinduseerde verlaging van sleutel betaselgene te verbeter vir optimale betaselfunksie.

published_or_final_version / Surgery / Doctoral / Doctor of Philosophy

Placental insufficiency-induced intrauterine growth restriction (IUGR) increases risk of mortality and morbidity in newborn infants and domestic animals. IUGR fetuses are typically exposed to prolonged hypoxemia, hypoglycemia, and hypercatecholaminemia, which results in perinatal pancreatic β-cell dysfunction. Recent evidence indicates that chronic exposure to norepinephrine in utero suppresses insulin secretion through α2-adrenergic receptors (ARs), but if the adrenergic actions are blocked compensatory hyper insulin secretion response is observed in the IUGR sheep fetus. In the current studies, we demonstrate that chronic NE exposure alone can produce the compensatory enhancement of β-cell responsiveness following termination of a chronic NE infusion. In the fetus NE was continuously infused at 1-4 μg/min for seven days starting at 131 days of gestational age (term = 145 days). During treatment, NE infused fetuses had higher (P < 0.05) plasma NE concentrations and lower (P < 0.01) insulin concentrations than vehicle infused control fetuses. Glucose stimulated insulin secretion (GSIS), which measures β-cell function, prior to NE treatment was not different between treatments. However, insulin concentrations during hyperglycemic steady state period of GSIS studies and area under the curve of glucose-potentiated arginine-induced insulin secretion were higher (P < 0.01) than control values and this augmentation was confirmed at 3 hours, 24 hours, and five days in NE-infused fetuses after discontinuing the infusion. Pancreatic islets isolated within 10 hours post NE infusion had lower (P < 0.05) mRNA expression of α1D (58%), α2A (43%), α2C (42%), α1 (67%) adrenergic receptors (ARs), and uncoupling protein 2 (40%) compared to islets from controls. Isolated islets from NE-infused fetuses 5 days after NE treatment had lower (P < 0.05) inhibitory responsiveness from NE and a greater (P < 0.05) maximal insulin release with glucose simulation in static incubations compared to controls. These findings show that following chronic NE exposure insulin secretion responsiveness was augmented and was coupled with desensitized adrenergic signaling. Moreover, this compensatory β-cell enhancement persists for days indicating chronic NE exposure permanently alters β-cell responsiveness.

Type 1 diabetes (T1D) results from a loss of functional insulin-producing pancreatic beta cells. The etiology of T1D is poorly understood, but the detection of infiltrating inflammatory cells in the pancreas and circulating autoantibodies has led to the common notion that an autoimmune process plays a central role in the pathogenesis of the disease. The aim of this doctoral thesis was to assess various aspects of the immunopathology of type 1 diabetes. To this purpose, studies have been conducted on pancreatic material from the Network for Pancreatic Organ Donors with Diabetes (nPOD) collection, the Nordic Network for Islet Transplantation, and the Diabetes Virus Detection (DiViD) study. Paper I is a study on pancreatic tissue from organ donors with varying duration of T1D as well as non-diabetic donors and subjects with other types of diabetes, in which persistent expression of glucose transporters was shown on the beta cell membrane despite several years of T1D. Glucose transporter 1 was also confirmed as the predominant glucose transporter on human pancreatic islets. In paper II, we report on signs of inflammation in the exocrine but not in the endocrine pancreas in non-diabetic organ donors with diabetes-related autoantibodies, suggesting that diabetes-associated autoantibodies can occur in response to unspecific pancreatic lesions. Paper III aimed to characterize the T cell-infiltration of pancreatic islets in material from recent-onset T1D patients. Insulitis was shown in all subjects, but with distinct differences in expression analysis of T- and B cell activation to cell-mediated allorejected kidney transplant. Also Paper IV was conducted on material from recent-onset cases and showed increased islet glucagon content, in combination with a reduced number of islets but sustained mean islet size. Together, these results provide expansion of our knowledge of the immunopathology in T1D, and will hopefully assist in bringing us towards a deeper understanding of T1D aetiology and eventually an effective cure.