RESUMO
Type 1 diabetes mellitus (T1DM) is predominantly managed using insulin replacement therapy, however, pancreatic microcirculatory disturbances play a critical role in T1DM pathogenesis, necessitating alternative therapies. This study aimed to investigate the protective effects of glycine supplementation on pancreatic microcirculation in T1DM. Streptozotocin-induced T1DM and glycine-supplemented mice (n = 6 per group) were used alongside control mice. Pancreatic microcirculatory profiles were determined using a laser Doppler blood perfusion monitoring system and wavelet transform spectral analysis. The T1DM group exhibited disorganized pancreatic microcirculatory oscillation. Glycine supplementation significantly restored regular biorhythmic contraction and relaxation, improving blood distribution patterns. Further-more, glycine reversed the lower amplitudes of endothelial oscillators in T1DM mice. Ultrastructural deterioration of islet microvascular endothelial cells (IMECs) and islet microvascular pericytes, including membrane and organelle damage, collagenous fiber proliferation, and reduced edema, was substantially reversed by glycine supplementation. Additionally, glycine supplementation inhibited the production of IL-6, TNF-α, IFN-γ, pro-MMP-9, and VEGF-A in T1DM, with no significant changes in energetic metabolism observed in glycine-supplemented IMECs. A statistically significant decrease in MDA levels accompanied by an increase in SOD levels was also observed with glycine supplementation. Notably, negative correlations emerged between inflammatory cytokines and microhemodynamic profiles. These findings suggest that glycine supplementation may offer a promising therapeutic approach for protecting against pancreatic microcirculatory dysfunction in T1DM.
Assuntos
Diabetes Mellitus Tipo 1 , Ilhotas Pancreáticas , Camundongos , Animais , Microcirculação , Células Endoteliais , Ilhotas Pancreáticas/irrigação sanguínea , Ilhotas Pancreáticas/metabolismo , Suplementos NutricionaisRESUMO
Accumulating evidence demonstrates that microcirculation plays a role in the pathogenesis of hypertension. In the current study, we demonstrated that pancreatic islet microvascular vasomotion of spontaneously hypertensive rats (SHRs) lost the ability to regulate blood flow perfusion and exhibited a lower microvascular blood perfusion pattern which was negative correlated with blood glucose level. SHRs administrated with insulin revealed an improvement of pancreatic islet microvascular vasomotion and blood perfusion pattern. In vitro, the expressions of endothelial nitric oxide synthase (eNOS) and phospho-eNOSser1177 (p-eNOSser1177) were significantly decreased in high glucose exposed islet endothelial cells (iECs), accompanied with a higher ratio of eNOS monomer to eNOS dimer and a significantly increased malondialdehyde and nitrite levels. Meanwhile, barrier function, tube formation and migration capacities of high glucose exposed iECs were significantly inhibited. In contrast, iECs dysfunction induced by glucose toxicity and oxidative stress was attenuated or improved by supplement with insulin, L-arginine and ß-mercaptoethanol. In summary, our findings suggest that functional status of pancreatic islet microvascular vasomotion is impaired in SHRs and provide evidence that treatment with insulin, L-arginine and ß-mercaptoethanol improves endothelium-dependent microvascular vasomotion and meliorates iECs function due to anti-hyperglycemic and anti-oxidative effects, partly through mechanism involving regulation of eNOS and p-eNOSser1177.
Assuntos
Hipertensão/patologia , Ilhotas Pancreáticas/irrigação sanguínea , Microcirculação/fisiologia , Animais , Arginina/farmacologia , Glicemia/análise , Pressão Sanguínea , Movimento Celular/efeitos dos fármacos , Dimerização , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Glucose/farmacologia , Hipertensão/metabolismo , Insulina/metabolismo , Insulina/farmacologia , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/metabolismo , Malondialdeído/metabolismo , Mercaptoetanol/farmacologia , Microcirculação/efeitos dos fármacos , Óxido Nítrico Sintase Tipo III/química , Óxido Nítrico Sintase Tipo III/metabolismo , Nitritos/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKYRESUMO
OBJECTIVE: To observe the effect of Citrullus colocynthis on beta cell regeneration and intra-islet vasculature. METHODS: This experimental study was conducted at the University of Health Sciences, Lahore, Pakistan, from February 2013 to January 2014. It comprised male wistar rats weighing 100-150gand aged 6-8 weeks. The animals were divided into 6 groups. Group A1 served as control. Diabetes was induced in groups A2, B2 and C2 using single intravenous injection of 50mg/kg of alloxan. Animals having fasting blood glucose>250mg/dl were considered diabetic. Diabetic rats in groups B2 and C2 and their controls B1 and C1 were given 1ml/kg and 2ml/kg of Citrullus colocynthis aqueous seed extract orally per day for 14 days. Animals were sacrifised on day 15. RESULTS: Of the 48 rats, there were 8(16.7%) in each group. Citrullus colocynthis has stabilized the body weight of rats and difference was statistically significant on days 7(p<0.013) and 14(p<0.001). Citrullus colocynthis significantly reduced (p<0.001) the fasting blood sugar levels in a dose- and time-dependent manner. It increased the islet diameter (p<0.001) and beta cell count (p<0.001). The number of intra-islet capillaries was increased in group C2, but the difference was not statistically significant (p>0.05). CONCLUSIONS: Citrullus colocynthis aqueous seed extract stabilised animal body weight and ameliorated hyperglycaemia in a dose- and time-dependent manner which was attributable to regenerative effect on beta cells and intra-islet vasculature.
Assuntos
Vasos Sanguíneos/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Citrullus colocynthis , Diabetes Mellitus Experimental , Células Secretoras de Insulina/efeitos dos fármacos , Ilhotas Pancreáticas/irrigação sanguínea , Extratos Vegetais/farmacologia , Sementes , Animais , Diabetes Mellitus , Ilhotas Pancreáticas/diagnóstico por imagem , Masculino , Ratos , Regeneração/efeitos dos fármacosRESUMO
BACKGROUND: The transplantation of isolated pancreatic islets is a promising treatment for diabetes. 5,7-dihydroxy-3,4,6-trimethoxyflavone (Eupatilin), a pharmacologically active flavone derived from the Artemisia plant species, has been reported to have antioxidant and anti-inflammatory activities. This study examines the hypothesis that preoperative eupatilin treatment can attenuate ischemic damage and apoptosis before islet transplantation. METHODS: Islets isolated from Balb/c mice were randomly divided into 2 groups, and cultured in medium supplemented with or without eupatilin. In vitro islet viability and function were assessed. After treatment with a cytokine cocktail consisting of tumor necrosis factor (TNF)-α, interferon (INF)-γ, and interleukin (IL)-1ß, islet cell viability, function, and apoptotic status were determined. The glutathione (GSH) and nitrous oxide (NO) levels were also measured. Proteins related to apoptosis were analyzed using Western blotting. RESULTS: There was no difference in cell viability between the 2 groups. Islets cultured in the medium supplemented with eupatilin showed 1.4-fold higher glucose-induced insulin secretion than the islets cultured in the medium without eupatilin. After treatment with a cytokine cocktail, glucose-induced insulin release and the total insulin content of the islets were significantly improved in eupatilin-pretreated islets compared with islets not treated with eupatilin. Apoptosis was significantly decreased, and GSH levels were elevated in the eupatilin-pretreated group. Cytokine-only treated islets produced significantly higher levels of NO, iNOS, and caspase-3 than islets pretreated with eupatilin before cytokine treatment. CONCLUSIONS: These results suggest that preoperative eupatilin administration enhances islet function before transplantation and attenuates the cytokine-induced damage associated with NO production and apoptosis.
Assuntos
Apoptose/efeitos dos fármacos , Flavonoides/farmacologia , Transplante das Ilhotas Pancreáticas , Ilhotas Pancreáticas/irrigação sanguínea , Traumatismo por Reperfusão/prevenção & controle , Animais , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Medicamentos de Ervas Chinesas , Feminino , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Camundongos , Camundongos Endogâmicos BALB C , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologiaRESUMO
Pancreatic islet blood perfusion varies according to the needs for insulin secretion. We examined the effects of blood lipids on pancreatic islet blood flow in anesthetized rats. Acute administration of Intralipid to anesthetized rats increased both triglycerides and free fatty acids, associated with a simultaneous increase in total pancreatic and islet blood flow. A preceding abdominal vagotomy markedly potentiated this and led acutely to a 10-fold increase in islet blood flow associated with a similar increase in serum insulin concentrations. The islet blood flow and serum insulin response could be largely prevented by pretreatment with propranolol and the selective ß3-adrenergic inhibitor SR-59230A. The nitric oxide synthase inhibitor N(G)-nitro-l-arginine methyl ester prevented the blood flow increase but was less effective in reducing serum insulin. Increased islet blood flow after Intralipid administration was also seen in islet and whole pancreas transplanted rats, i.e., models with different degrees of chronic islet denervation, but the effect was not as pronounced. In isolated vascularly perfused single islets Intralipid dilated islet arterioles, but this was not affected by SR-59230A. Both the sympathetic and parasympathetic nervous system are important for the coordination of islet blood flow and insulin release during hyperlipidemia, with a previously unknown role for ß3-adrenoceptors.
Assuntos
Hiperlipidemias/fisiopatologia , Insulina/metabolismo , Ilhotas Pancreáticas/irrigação sanguínea , Receptores Adrenérgicos beta 3/metabolismo , Fluxo Sanguíneo Regional , Regulação para Cima , Nervo Vago/fisiopatologia , Antagonistas de Receptores Adrenérgicos beta 3/farmacologia , Animais , Emulsões/efeitos adversos , Ácidos Graxos não Esterificados/sangue , Ácidos Graxos não Esterificados/metabolismo , Hiperlipidemias/sangue , Hiperlipidemias/etiologia , Hiperlipidemias/metabolismo , Insulina/sangue , Secreção de Insulina , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/inervação , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Pâncreas/irrigação sanguínea , Pâncreas/efeitos dos fármacos , Pâncreas/inervação , Pâncreas/metabolismo , Perfusão , Fosfolipídeos/efeitos adversos , Propanolaminas/farmacologia , Ratos Endogâmicos WF , Receptores Adrenérgicos beta 3/química , Fluxo Sanguíneo Regional/efeitos dos fármacos , Óleo de Soja/efeitos adversos , Triglicerídeos/sangue , Triglicerídeos/metabolismo , Regulação para Cima/efeitos dos fármacos , Vagotomia Troncular , Nervo Vago/efeitos dos fármacos , Nervo Vago/cirurgiaRESUMO
OBJECTIVE: This study investigates the therapeutic potential of hyperbaric oxygen therapy (HBO) in reducing hypoxia and improving engraftment of intraportal islet transplants by promoting angiogenesis. METHODS: Diabetic BALB/c mice were transplanted with 500 syngeneic islets intraportally and received six consecutive twice-daily HBO treatments (n = 9; 100% oxygen for 1 h at 2.5 atmospheres absolute) after transplantation. Dynamic contrast-enhanced magnetic resonance imaging (DCE MRI) was used to assess new vessel formation at postoperative days (POD) 3, 7, and 14. Liver tissue was recovered at the same time points for correlative histology, including: hematoxylin and eosin, hypoxia-inducible factor (HIF1α), Terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL), vascular endothelial growth factor (VEGF), and von Willebrand Factor immunohistochemistry. RESULTS: HBO therapy significantly reduced HIF-1α, TUNEL and VEGF expression in islets at POD 7. In the non-HBO transplants, liver enhancement on DCE MRI peaked at POD 7 consistent with less mature vasculature but this enhancement was suppressed at POD 7 in the HBO-treated group. The number of new peri-islet vessels at POD 7 was not significantly different between HBO and control groups. CONCLUSION: These results are consistent with a hyperbaric oxygen-mediated decrease in hypoxia that appeared to enhance vessel maturation in the critical days following intraportal islet transplantation.
Assuntos
Diabetes Mellitus Experimental/terapia , Oxigenoterapia Hiperbárica , Transplante das Ilhotas Pancreáticas/métodos , Neovascularização Fisiológica/efeitos dos fármacos , Animais , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Marcação In Situ das Extremidades Cortadas , Ilhotas Pancreáticas/irrigação sanguínea , Ilhotas Pancreáticas/fisiologia , Imageamento por Ressonância Magnética , Camundongos , Camundongos Endogâmicos BALB C , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Previous studies have shown that the Goto-Kakizaki (GK) rat, a nonobese type 2 diabetes model, has an increased white adipose tissue (WAT) and islet blood flow when compared with control rats. The aim of the study was to examine if these increased blood flow values in GK rats could be affected by the beta(3)-adrenoceptor antagonist SR-59230A. We measured organ blood flow with a microsphere technique 10 min after administration of SR-59230A (1 mg/kg body wt), or the corresponding volume of 0.9% NaCl solution (1 ml/kg body wt) in rats anaesthetized with thiobutabarbital. The GK rat had an increased blood flow in all intra-abdominal adipose tissue depots except for the sternal fat pad compared with Wistar-Furth (WF) rats. However, no differences were seen in the blood perfusion of subcutaneous white or brown adipose tissue. The blood flow was also increased in both the pancreas and in the islets in the GK rat compared with WF rats. SR-59230A treatment affected neither WAT nor pancreatic blood flow in WF rats. In GK rats, on the other hand, SR-59230A decreased both WAT and islet blood flow values to values similar to those seen in control WF rats. The whole pancreatic blood flow was not affected by SR-59230A administration in GK rats. Interestingly, the brown adipose tissue blood flow in GK rats increased after SR-59230A administration. These results suggest that beta(3)-adrenoceptors are involved in regulation of blood flow both in islet and in adipose tissue.
Assuntos
Tecido Adiposo Branco/efeitos dos fármacos , Inibidores da Angiogênese/uso terapêutico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Ilhotas Pancreáticas/efeitos dos fármacos , Propanolaminas/uso terapêutico , Fluxo Sanguíneo Regional/efeitos dos fármacos , Tecido Adiposo Marrom/irrigação sanguínea , Tecido Adiposo Marrom/efeitos dos fármacos , Tecido Adiposo Branco/irrigação sanguínea , Antagonistas de Receptores Adrenérgicos beta 3 , Estruturas Animais/irrigação sanguínea , Estruturas Animais/efeitos dos fármacos , Animais , Avaliação Pré-Clínica de Medicamentos , Expressão Gênica/efeitos dos fármacos , Ilhotas Pancreáticas/irrigação sanguínea , Masculino , Ratos , Ratos Endogâmicos , Ratos Endogâmicos WF , Receptores Adrenérgicos beta 3/genética , Receptores Adrenérgicos beta 3/metabolismoRESUMO
BACKGROUND/PURPOSE: Apoptosis resulting from disruption of the normal cell-matrix relationship (anoikis) during islet isolation, and the reactive oxygen and nitrogen species generated following hypoxia/reoxygenation (H/R) can lead to a loss of islet tissue in culture and the reduced survival of transplanted pancreatic islets. The aim of this study was to investigate the effect of (-)-epigallocatechin-3-gallate (EGCG), a well-known antiapoptotic agent, on inhibiting anoikis and H/R injury in an in vitro islet culture system. METHODS: Islets were isolated from F344 rats and cultured under normal or H/R condition with/without EGCG. RESULTS: EGCG inhibited apoptosis and lactate-dehydrogenase leakage from anoikis and H/R in a dose-dependent manner. Further, EGCG prevent increases in 8-hydroxy-2'-deoxyguanosine content and inhibited the decline of insulin secretory function induced by H/R. CONCLUSIONS: These results suggest that the addition of EGCG to an islet culture system may improve the survival rate of isolated islets and reduce the loss of functional islet mass that compromises the stable reversal of diabetes after islet transplantation.
Assuntos
Anoikis/efeitos dos fármacos , Antioxidantes/farmacologia , Catequina/análogos & derivados , Hipóxia Celular/efeitos dos fármacos , Ilhotas Pancreáticas/fisiologia , Traumatismo por Reperfusão/tratamento farmacológico , 8-Hidroxi-2'-Desoxiguanosina , Animais , Catequina/farmacologia , Hipóxia Celular/fisiologia , Desoxiguanosina/análogos & derivados , Desoxiguanosina/análise , Insulina/metabolismo , Ilhotas Pancreáticas/irrigação sanguínea , Ilhotas Pancreáticas/efeitos dos fármacos , L-Lactato Desidrogenase/análise , Ratos , Ratos Endogâmicos F344 , Chá/químicaRESUMO
A technique to measure angiogenesis and revascularization in pancreatic islets transplanted at the renal subcapsular site in the rat has been developed. In-vivo imaging of the microcirculation of transplanted pancreatic islets was conducted using a confocal scanning laser microscope (CSLM) to achieve optical sectioning through the graft in order to perform a computer reconstruction of the three-dimensional neovascular morphology. Individual islets were harvested by enzymatic digestion of excised pancreas from Fischer 344 rats. Isolated islets were cultured for 24 h, and approximately 300-350 islets were transplanted at the renal subcapsular site of the left kidney in an anaesthetized rat. Six to 14 days post-transplantation, the animal was anaesthetized and prepared for in-vivo imaging of the microvasculature on a Zeiss LSM-10. Optical contrast of the microvasculature was enhanced by the administration of fluorescein-labelled dextran into the circulating blood. The transplant site was identified and serial sections were obtained through the vascular bed at varying z-intervals. Complementary fluorescence video images were also obtained via a silicon intensifier tube camera mounted on the CSLM. At completion of the imaging procedure, the kidney was returned into the body cavity, the area was sutured and the animal was allowed to recuperate for subsequent examinations. Image processing algorithms, such as grey-level thresholding, median filtering, skeletonization and template matching, were applied to compute the vessel density and diameters and extrapolated to measure 3-D vessel lengths and the tortousity index of the neovasculature.
Assuntos
Processamento de Imagem Assistida por Computador/métodos , Transplante das Ilhotas Pancreáticas , Ilhotas Pancreáticas/irrigação sanguínea , Microscopia Confocal/métodos , Neovascularização Patológica , Algoritmos , Animais , Rim/irrigação sanguínea , Microcirculação , Ratos , Ratos Endogâmicos F344 , Transplante HeterotópicoRESUMO
Hyperinsulinemia, a main feature of both human and animal obesity, has been demonstrated to be due to both an increased sensitivity to nutrient secretagogues and an impairment of the nervous regulation of insulin secretion. Recent studies have shown that pancreatic islet blood flow increases under conditions associated with an enhanced insulin secretion. The aim of this study was to determine whether or not changes in islet blood flow are present in hyperinsulinemic obese rats. Using the nonradioactive microsphere technique, we were able to show a significantly higher islet blood flow in obese rats either of the Zucker strain or Wistar rats after lesion of the ventromedial hypothalamus than in their respective lean controls. Subdiaphragmatic vagotomy had no significant effect on basal islet blood flow of lean rats, whereas it decreased significantly that of obese Zucker rats. Conversely, clonidine, an alpha 2-adrenergic agonist, induced a higher decrease of islet blood flow in obese than in lean Zucker rats. The injection of an intravenous bolus of glucose (375 mg/kg iv) increased significantly more islet blood flow in obese than in lean Zucker rats. It is concluded that obese rats present an increased pancreatic islet blood flow, which may result, at least in part, from exaggerated parasympathetic activity and lower than normal sympathetic activity. This could participate in the hyperinsulinemia observed in these rats.