Publications (8)27.86 Total impact
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Article: Altered NAD(P)H production in neonatal rat islets resistant to H2O2.
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ABSTRACT: We determined the involvement of NAD(P)H generation ability on the resistance of pancreatic islets B-cells to oxidative stress caused by culture exposition to H2O2. We cultured isolated neonatal Wistar rat islets for four days in medium containing 5.6 or 20 mM glucose, with or without H2O2 (200 microM), and analyzed several parameters associated with islet survival in different media. High glucose was used since it protects neonatal islets against the loss of GSIS. While none of the culture conditions increased the rate of NAD(P)H content at 16.7 mM glucose, the islets resistant to H2O2 and those exposed to 20 mM glucose showed a greater use of the pentose phosphate pathway and increased ATP synthesis from glucose. Oxidative stress contributes to the loss of glucose-induced insulin secretion (GSIS) during the onset of diabetes mellitus. Although immature rat islets have reduced GSIS compared to mature islets, they adapt better to oxidative stress and are a good model for understanding the causes involved in the destruction or survival of islet cells. These data support the idea that GSIS and resistance against oxidative stress in immature islets rely on NADH shuttle activities, with little contribution of reduced equivalents from the tricarboxylic acid cycle (TCAC).Life Sciences 10/2008; 83(21-22):709-16. · 2.53 Impact Factor -
Article: INGAP-PP up-regulates the expression of genes and proteins related to K+ ATP channels and ameliorates Ca2+ handling in cultured adult rat islets.
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ABSTRACT: Islet Neogenesis Associated Protein (INGAP) increases pancreatic beta-cell mass and potentiates glucose-induced insulin secretion. Here, we investigated the effects of the pentadecapeptide INGAP-PP in adult cultured rat islets upon the expression of proteins constitutive of the K(+)(ATP) channel, Ca(2+) handling, and insulin secretion. The islets were cultured in RPMI medium with or without INGAP-PP for four days. Thereafter, gene (RT-PCR) and protein expression (Western blotting) of Foxa2, SUR1 and Kir6.2, cytoplasmic Ca(2+) ([Ca(2+)](i)), static and dynamic insulin secretion, and (86)Rb efflux were measured. INGAP-PP increased the expression levels of Kir6.2, SUR1 and Foxa2 genes, and SUR1 and Foxa2 proteins. INGAP-PP cultured islets released significantly more insulin in response to 40 mM KCl and 100 muM tolbutamide. INGAP-PP shifted to the left the dose-response curve of insulin secretion to increasing concentrations of glucose (EC(50) of 10.0+/-0.4 vs. 13.7+/-1.5 mM glucose of the controls). It also increased the first phase of insulin secretion elicited by either 22.2 mM glucose or 100 microM tolbutamide and accelerated the velocity of glucose-induced reduction of (86)Rb efflux in perifused islets. These effects were accompanied by a significant increase in [Ca(2+)](i) and the maintenance of a considerable degree of [Ca(2+)](i) oscillations. These results confirm that the enhancing effect of INGAP-PP upon insulin release, elicited by different secretagogues, is due to an improvement of the secretory function in cultured islets. Such improvement is due, at least partly, to an increased K(+)(ATP) channel protein expression and/or changing in the kinetic properties of these channels and augmented [Ca(2+)](i) response. Accordingly, INGAP-PP could potentially be used to maintain the functional integrity of cultured islets and eventually, for the prevention and treatment of diabetes.Regulatory Peptides 07/2008; 148(1-3):39-45. · 2.11 Impact Factor -
Article: Impaired insulin secretion and decreased expression of the nutritionally responsive ribosomal kinase protein S6K-1 in pancreatic islets from malnourished rats.
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ABSTRACT: Low protein diet has been shown to affect the levels and activities of several enzymes from pancreatic islets. To further extend the knowledge on how malnutrition affects insulin secretion pathway, we investigated in this work the insulin release induced by glucose or leucine, an insulin secretagogue, and the expression of insulin receptor (IR), insulin receptor substrate 1 (IRS1), phosphatidylinositol 3-kinase (PI3K), and p70S6K1 (S6K-1) proteins from pancreatic islets of rats fed a normal (17%; NP) or a low (6%; LP) protein diet for 8 weeks. Isolated islets were incubated for 1 h in Krebs-bicarbonate solution containing 16.7 mmol/L of glucose, or 2.8 mmol/L of glucose in the presence or absence of 20 mmol/L of leucine. Glucose- and leucine-induced insulin secretions were higher in NP than in LP islets. Western blotting analysis showed an increase in the expression of IR and PI3K protein levels whereas IRS1 and S6K-1 protein expression were lower in LP compared to NP islets. In addition, S6K-1 mRNA expression was also reduced in islets from LP rats. Our data indicate that a low protein diet modulates the levels of several proteins involved in the insulin secretion pathway. Particularly, the decrease in S6K-1 expression might be an important factor affecting either glucose- or leucine-induced insulin secretion.Life Sciences 03/2008; 82(9-10):542-8. · 2.53 Impact Factor -
Article: Ciliary neurotrophic factor promotes survival of neonatal rat islets via the BCL-2 anti-apoptotic pathway.
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ABSTRACT: Ciliary neurotrophic factor (CNTF) belongs to the cytokine family and increases neuron differentiation and/or survival. Pancreatic islets are richly innervated and express receptors for nerve growth factors (NGFs) and may undergo neurotypic responses. CNTF is found in pancreatic islets and exerts paracrine effects in neighboring cells. The aim of this study was to investigate possible effects of CNTF on neonatal rat pancreatic islet differentiation and/or survival. For this purpose, we isolated pancreatic islets from neonatal rats (1-2 days old) by the collagenase method and cultured for 3 days in RPMI medium with (CNTF) or without (CTL) 1 nM CNTF. Thereafter, glucose-stimulated insulin secretion (RIA), general metabolism by (NAD(P)H production; MTS), glucose metabolism ((14)CO(2) production), gene (RT-PCR), protein expression (western blotting), caspase-3 activity (Asp-Glu-Val-Asp (DEVD)), and apoptosis (DNA fragmentation) were analyzed. Our results showed that CNTF-treated islets demonstrated reduced glucose-induced insulin secretion. CNTF treatment did not affect glucose metabolism, as well as the expression of mRNAs and proteins that are crucial for the secretory process. Conversely, CNTF significantly increased mRNA and protein levels related to cell survival, such as Cx36, PAX4, and BCL-2, reduced caspase-3 activity, and islet cells apoptosis, suggesting that CNTF does not affect islet cell differentiation and, instead, acts as a survival factor reducing apoptosis by increasing the expression of the anti-apoptotic BCL-2 protein and decreasing caspase-3 activity.Journal of Endocrinology 11/2007; 195(1):157-65. · 3.55 Impact Factor -
Article: Inhibition of UCP2 expression reverses diet-induced diabetes mellitus by effects on both insulin secretion and action.
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ABSTRACT: Recent characterization of the ability of uncoupling protein 2 (UCP2) to reduce ATP production and inhibit insulin secretion by pancreatic beta-cells has placed this mitochondrial protein as a candidate target for therapeutics in diabetes mellitus. In the present study we evaluate the effects of short-term treatment of two animal models of type 2 diabetes mellitus with an antisense oligonucleotide to UCP2. In both models, Swiss mice (made obese and diabetic by a hyperlipidic diet) and ob/ob mice, the treatment resulted in a significant improvement in the hyperglycemic syndrome. This effect was due not only to an improvement of insulin secretion, but also to improved peripheral insulin action. In isolated pancreatic islets, the partial inhibition of UCP2 increased ATP content, followed by increased glucose-stimulated insulin secretion. This was not accompanied by increased expression of enzymes involved in protection against oxidative stress. The evaluation of insulin action in peripheral tissues revealed that the inhibition of UCP2 expression significantly improved insulin signal transduction in adipose tissue. In conclusion, short-term inhibition of UCP2 expression ameliorates the hyperglycemic syndrome in two distinct animal models of obesity and diabetes. Metabolic improvement is due to a combined effect on insulin-producing pancreatic islets and in at least one peripheral tissue that acts as a target for insulin.The FASEB Journal 05/2007; 21(4):1153-63. · 5.71 Impact Factor -
Article: INGAP-related pentadecapeptide: its modulatory effect upon insulin secretion.
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ABSTRACT: We examined the effects of a pentadecapeptide having the 104-118 aminoacid sequence of islet neogenesis-associated protein (INGAP-PP) on insulin secretion, and the morphological characteristics of adult and neonatal pancreatic rat islets cultured in RPMI and 10 mM glucose for 4 days, with or without different INGAP-PP concentrations (0.1-100 mug/ml). A scrambled 15 aminoacid peptide was used as control for the specificity of INGAP-PP effect. Cultured neonatal and adult islets released insulin in response to glucose (2.8-16.7 mM) in a dose-dependent manner, and to leucine and arginine (10 mM). In all cases, the response was greater in adult islets. INGAP-PP added to the culture medium significantly enhanced glucose- and aminoacid-induced insulin release in both adult and newborn rats; however, no changes were observed with the scrambled peptide. Similar results were obtained incubating freshly isolated adult rat islets with INGAP-PP. Whereas INGAP-PP did not induce significant changes in islet survival rate or proportion/number of islet cells, it increased significantly beta-cell size. This first demonstration of the enhancing effect of INGAP-PP on the beta-cell secretory response of adult and newborn islets opens a new avenue to study its production mechanism and potential use to increase the secretory capacity of endogenous islets in intact animals or of islets preserved for future transplants.Regulatory Peptides 12/2005; 131(1-3):97-102. · 2.11 Impact Factor -
Article: Characterization of the peroxidase system at low H2O2 concentrations in isolated neonatal rat islets.
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ABSTRACT: B cell destruction during the onset of diabetes mellitus is associated with oxidative stress. In this work, we attempted to further trace the fate of H2O2 inside the pancreatic islets and determine whether it is mediated by enzymatic (peroxidase) activity or by chemical reaction with thiols from any protein chain. Our results suggest that the islet cells have a very similar peroxidase activity at the hydrophilic (cytoplasm) and hydrophobic compartments (organelles and nucleus), independent of the catalase content of the samples. This activity is composed of sacrificial thiols and by proteins with Fe3+/Mn3+ ions at non-heme catalytic sites. The capacity of the hydrophobic fraction to scavenge O2- was increased in the presence of high concentrations of NADP* and RS* and was highly dependent on RSH. On the contrary, the hydrophilic fraction exhibited a low RSH-dependent activity where the O2- scavenging is related to metal Cu2+/Fe3+/Mn3+ ions attached to the protein molecules.Biochimica et Biophysica Acta 11/2004; 1690(2):159-68. · 4.66 Impact Factor -
Article: Protective effect of D-glucose, L-leucine and fetal calf serum against oxidative stress in neonatal pancreatic islets.
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ABSTRACT: B-cell destruction during the onset of diabetes mellitus is associated with oxidative stress. In this work, we investigated the mechanisms of defense against oxidative stress present in neonatal islets and their modulation by D-glucose, L-leucine and fetal calf serum (FCS). Culturing neonatal rat islets in the presence of low D-glucose concentrations (2.8-5.6 mmol/l) and 1 mmol/l H(2)O(2) increased the D-glucose uptake by islets sixfold compared to control levels. This effect was dose-dependently inhibited by D-glucose or FCS and by high concentrations of L-leucine. These supplements allowed islets to increase cytoplasmic catalase (CAT) activity only in response to H(2)O(2), with no decrease in NO formation. Although L-leucine increased CAT activity and restored D-glucose uptake, it did not prevent damage to the islets. These data indicate that the most important H(2)O(2) scavenger system in the islets is CAT and that this system can be modulated by metabolic substrates.Biochimica et Biophysica Acta 12/2002; 1588(2):113-8. · 4.66 Impact Factor
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Institutions
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2002–2004
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Universidade Estadual de Campinas
- Instituto de Biologia (IB)
Campinas, Estado de Sao Paulo, Brazil
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