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1.
Toxicol Lett ; 320: 19-27, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31778773

RESUMO

The deleterious effects of glucocorticoids on glucose homeostasis limit their clinical use. There is substantial evidence demonstrating that islet function impaired by long-term glucocorticoids exposure is a core defect in the progression of impaired glucose tolerance to diabetes. The activity of heat-shock protein (Hsp) 90 is required to maintain the hormone-binding activity and stability of glucocorticoid receptor (GR). In the present study, Hsp90 inhibition by 17-DMAG counteracted dexamethasone-mediated inhibition of glucose-stimulated insulin secretion in isolated rat islets as well as expressions of neuropeptide Y (NPY) and somatostatin receptor 3 (SSTR3), two negative regulators of insulin secretion. Like 17-DMAG, both the pan-histone deacetylase (HDAC) inhibitor TSA and HDAC6 inhibitor Tubacin exhibited a similar action in protecting islet function against dexamethasone-induced injury, along with the downregulation of NPY and SSTR3 expressions. The hyperacetylation of Hsp90 by TSA and Tubacin disrupted its binding ability to GR and blocked dexamethasone-elicited nuclear translocation of GR in INS-1 ß-cell lines. In addition, Tubacin treatment triggered the GR protein degradation through the ubiquitin-proteasome pathway. These findings suggest that Hsp90 acetylation by inhibiting HDAC6 activity may be a potential strategy to prevent the development of steroid diabetes mellitus via alleviating glucocorticoid-impaired islet function.


Assuntos
Anilidas/farmacologia , Benzoquinonas/farmacologia , Dexametasona/toxicidade , Glucocorticoides/toxicidade , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Desacetilase 6 de Histona/antagonistas & inibidores , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , Insulina/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Lactamas Macrocíclicas/farmacologia , Acetilação , Animais , Linhagem Celular , Proteínas de Choque Térmico HSP90/metabolismo , Desacetilase 6 de Histona/metabolismo , Ilhotas Pancreáticas/metabolismo , Masculino , Complexo de Endopeptidases do Proteassoma/metabolismo , Processamento de Proteína Pós-Traducional , Proteólise , Ratos Sprague-Dawley , Via Secretória , Técnicas de Cultura de Tecidos
2.
Life Sci ; 237: 116913, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31622609

RESUMO

AIMS: To explore the impact of GC administration periconceptionally on the glucose metabolism of adult offspring (male and female) and whether this periconception exposure might influence the metabolic outcomes when the offspring are also treated with dexamethasone in adult life. MATERIALS AND METHODS: Rats received a daily injection of dexamethasone (1 mg/kg, body mass) or saline solution (1 mL/kg body mass) for 7 consecutive days prior became pregnant. Male and female offspring had glucose homeostasis assessed at 3- and 6-month-old and after dexamethasone treatment (1 mg/kg, body mass) or vehicle for 5 consecutive days. Then, murinometric, functional, biochemical, and histomorphometric analyses were performed. KEY FINDINGS: Male and female offspring born from rats treated with GC prior to becoming pregnant had none of the murinometric and metabolic outcomes (i.e., body mass, food intake, blood glucose, plasma triacylglycerol, and glucose tolerance) changed up to 6-month-old. None of the expected diabetogenic effects caused by dexamethasone treatment at 6-month of age (i.e., elevation in fasting blood glucose, plasma insulin, triacylglycerol, and albumin, glucose intolerance, insulin insensitivity, augmentation in hepatic glycogen content, and increase in pancreatic islet mass) was observed in offspring born from rats treated with dexamethasone in the prepregnancy period. However, periconceptional exposure to GC predisposed the offspring of both sexes to a higher prevalence of augmented fed blood glucose values. SIGNIFICANCE: These results give validity for the use of GC as anti-inflammatory purposes in this critical periconceptional period, but highlight the importance to consider all parental habits when interpreting adult outcomes.


Assuntos
Dexametasona/administração & dosagem , Intolerância à Glucose/tratamento farmacológico , Homeostase , Secreção de Insulina/efeitos dos fármacos , Cuidado Pré-Concepcional , Animais , Glicemia/análise , Peso Corporal , Feminino , Glucocorticoides/administração & dosagem , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/crescimento & desenvolvimento , Masculino , Estresse Oxidativo/efeitos dos fármacos , Gravidez , Prenhez , Ratos , Ratos Wistar
3.
Cell Physiol Biochem ; 53(3): 573-586, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31529929

RESUMO

BACKGROUND/AIMS: In our recent work, the importance of GSK3ß-mediated phosphorylation of presenilin-1 as crucial process to establish a Ca2+ leak in the endoplasmic reticulum and, subsequently, the pre-activation of resting mitochondrial activity in ß-cells was demonstrated. The present work is a follow-up and reveals the importance of GSK3ß-phosphorylated presenilin-1 for responsiveness of pancreatic islets and ß-cells to elevated glucose in terms of cytosolic Ca2+ spiking and insulin secretion. METHODS: Freshly isolated pancreatic islets and the two pancreatic ß-cell lines INS-1 and MIN-6 were used. Cytosolic Ca2+ was fluorometrically monitored using Fura-2/AM and cellular insulin content and secretion were measured by ELISA. RESULTS: Our data strengthened our previous findings of the existence of a presenilin-1-mediated ER-Ca2+ leak in ß-cells, since a reduction of presenilin-1 expression strongly counteracted the ER Ca2+ leak. Furthermore, our data revealed that cytosolic Ca2+ spiking upon administration of high D-glucose was delayed in onset time and strongly reduced in amplitude and frequency upon siRNA-mediated knock-down of presenilin-1 or the inhibition of GSK3ß in the pancreatic ß-cells. Moreover, glucose-triggered initial insulin secretion disappeared by depletion from presenilin-1 and inhibition of GSK3ß in the pancreatic ß-cells and isolated pancreatic islets, respectively. CONCLUSION: These data complement our previous work and demonstrate that the sensitivity of pancreatic islets and ß-cells to glucose illustrated as glucose-triggered cytosolic Ca2+ spiking and initial but not long-lasting insulin secretion crucially depends on a strong ER Ca2+ leak that is due to the phosphorylation of presenilin-1 by GSK3ß, a phenomenon that might be involved in the development of type 2 diabetes.


Assuntos
Retículo Endoplasmático/metabolismo , Glucose/farmacologia , Células Secretoras de Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Presenilina-1/metabolismo , Animais , Antracenos/farmacologia , Cálcio/metabolismo , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Humanos , Secreção de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/efeitos dos fármacos , Ilhotas Pancreáticas/efeitos dos fármacos , MAP Quinase Quinase 4/antagonistas & inibidores , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo
4.
J Diabetes Res ; 2019: 2813489, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31467926

RESUMO

While the autoimmune character of T1D (type 1 diabetes) is being challenged, it is currently recognized that inflammation plays a key role in its development. We hypothesized that glucotoxicity could contribute to ß-cell mass destruction through participation in islet inflammation. We evaluated the potential of empagliflozin (EMPA) and GABA (gamma-aminobutyric acid) to protect ß-cell mass against glucotoxicity and to increase ß-cell mass after diagnosis of T1D. Empagliflozin is a SGLT2 (sodium-dependent glucose cotransporter) inhibitor which thereby blocks glucose recapture by the kidney and promotes glucose excretion in urine. GABA is an inhibitory neurotransmitter, which stimulates α-to-ß cell transdifferentiation. In streptozotocin-treated mice, empagliflozin and/or GABA were delivered for a period of five days or three weeks. As compared to untreated T1D mice, EMPA-treated T1D mice had decreased FFA (free fatty acid) levels and improved glucose homeostasis. EMPA-treated T1D mice had higher islet density, with preserved architecture, compared to T1D mice, and EMPA-treated T1D mice also differed from T1D mice by the total absence of immune cell infiltration within islets. Islets from EMPA-treated mice were also less subjected to ER (endoplasmic reticulum) stress and inflammation, as shown by qPCR analysis. Glucose homeostasis parameters and islet area/pancreas area ratio improved, as compared to diabetic controls, when T1D mice were treated for three weeks with GABA and EMPA. T1D EMPA+GABA mice had higher glucagon levels than T1D mice, without modifications of glucagon area/islet area ratios. In conclusion, empagliflozin and GABA, used in monotherapy in streptozotocin-induced diabetic mice, have positive effects on ß-cell mass preservation or proliferation through an indirect effect on islet cell inflammation and ER stress. Further research is mandatory to evaluate whether empagliflozin and GABA may be a potential therapeutic target for the protection of ß-cell mass after new-onset T1D.


Assuntos
Compostos Benzidrílicos/farmacologia , Glicemia/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/metabolismo , Glucosídeos/farmacologia , Células Secretoras de Insulina/efeitos dos fármacos , Ácido gama-Aminobutírico/farmacologia , Animais , Compostos Benzidrílicos/administração & dosagem , Glicemia/metabolismo , Contagem de Células , Proliferação de Células/efeitos dos fármacos , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patologia , Esquema de Medicação , Quimioterapia Combinada , Intervenção Médica Precoce , Teste de Tolerância a Glucose , Glucosídeos/administração & dosagem , Injeções Intraperitoneais , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/fisiologia , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/fisiologia , Masculino , Camundongos , Estreptozocina , Fatores de Tempo , Ácido gama-Aminobutírico/administração & dosagem
5.
Phytomedicine ; 63: 153002, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31301539

RESUMO

BACKGROUND: Type 1 diabetes is an autoimmune disease directed to the pancreatic islets where inflammation leads to the death of insulin-producing ß cells and insulin deficiency. Type 2 diabetes, which is closely related to overweight, is characterized by insulin resistance. In both cases, proinflammatory cytokines play an important role by causing insulitis and insulin resistance. The gum resin of Boswellia species and its pharmacologically active compounds, including 11-keto-ß-boswellic acids have been shown to suppress the expression of proinflammatory cytokines in various immune-competent cells. PURPOSE: To review the present evidence of the therapeutic effects of boswellic extracts (BE) and/or 11-keto-ß-boswellic acids in the prevention/treatment of diabetes mellitus and to provide comprehensive insights into the underlying molecular mechanisms. METHODS: This review considers all available informations from preclinical and clinical studies concerning BEs, 11-keto-ß-boswellic acids, proinflammatory cytokines and diabetes mellitus collected via electronic search (PubMed) and related publications of the author. RESULTS: Type 1 diabetes: Studies in mice with autoimmune diabetes revealed that in the model of multiple injections of low doses of streptozotocin (MLD-STZ), an extract of the gum resin of Boswellia serrata and 11-keto-ß-boswellic acid (KBA) suppressed the increase in proinflammatory cytokines in the blood, infiltration of lymphocytes into pancreatic islets and increase in blood glucose. In a second model, i.e. the nonobese diabetic (NOD) mouse, KBA prevented the infiltration of lymphocytes into pancreatic islets. Regarding the clinical effects, a case report provided evidence that BE suppressed the blood levels of tyrosine phosphatase antibody (IA2-A), a marker for insulitis, in a patient with late-onset autoimmune diabetes of the adult (LADA). Type 2 diabetes: In a preclinical study in rats where obesity was alimentary induced, the administration of BE significantly reduced food intake, overweight, proinflammatory cytokines such as interleukin-1ß (IL-1ß), and tumor necrosis factor-α (TNF-α) and ameliorated the parameters of glucose and lipid metabolism. Similar results were obtained in a second animal study, where type 2 diabetes was induced by a combination of a high-fat/high-fructose diet and a single dose of streptozotocin. Two clinical trials with patients with type 2 diabetes receiving the resin of Boswellia serrata demonstrated improvement in the blood glucose, HbA1c and lipid parameters. CONCLUSION: Preclinical and clinical data suggest that BE and/or 11-keto-ß-boswellic acids by inhibiting the expression of proinflammatory cytokines from immune-competent cells, may prevent insulitis and insulin resistance in type 1 and type 2 diabetes, respectively, and therefore may be an option in the treatment/prevention of type 1 and type 2 diabetes. It is hypothesized that molecularly, BE and 11-keto-ß-boswellic acids act via interference with the IκB kinase/Nuclear Transcription Factor-κB (IKK/NF-κB) signaling pathway through inhibition of the phosphorylation activity of IKK. However, further investigations and well-designed clinical studies are required.


Assuntos
Boswellia/química , Diabetes Mellitus Tipo 1/prevenção & controle , Diabetes Mellitus Tipo 2/prevenção & controle , Extratos Vegetais/farmacologia , Animais , Citocinas/sangue , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patologia , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patologia , Humanos , Insulina/farmacologia , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/patologia , Linfócitos/efeitos dos fármacos , Linfócitos/patologia , Camundongos , Camundongos Endogâmicos NOD , NF-kappa B/metabolismo , Extratos Vegetais/química , Ratos , Resinas Vegetais/química , Resinas Vegetais/farmacologia , Triterpenos/farmacologia
6.
Eur J Pharmacol ; 858: 172518, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31265840

RESUMO

Minocycline functions as a therapeutic drug in different diseases because of its cytoprotective properties. In the present study, we examined the potential of minocycline to decrease the islet loss in pre-transplantation culture stage. Pancreatic islets were isolated from the deceased donors and treated by 0, 2, 10, and 20 µM minocycline for 24 and 72 h. After that, the incubated islets were evaluated for viability and function. Apoptosis markers including Bax, Bcl2, and caspase-3 were determined at gene and protein level. On the other hand, TUNEL assay was used to confirm apoptosis. The functionality of the islets was investigated using glucose-induced insulin and c-peptide secretion assay. After 72 h of incubation, the viability of human islet was drastically decreased, whereas supplementation with minocycline inhibited the cells death. In this regard, the expression of Bax and active Caspase-3 was downregulated, whereas the expression of Bcl2 was upregulated. These consequences suggest that pancreatic islets undergo apoptosis in vitro and minocycline can decelerate or inhibit this process. Our findings identified minocycline as a cytoprotective molecule for preventing human islets death in pre-transplantation culture.


Assuntos
Apoptose/efeitos dos fármacos , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/efeitos dos fármacos , Minociclina/farmacologia , Caspase 3/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Humanos , Ilhotas Pancreáticas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteína X Associada a bcl-2/metabolismo
7.
Eur J Pharmacol ; 858: 172514, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31265841

RESUMO

Recently, we reported the role of coixol (6-methoxy-2(3H)-benzoxazolone), an alkaloid from Scoparia dulcis, in glucose-dependent insulin secretion; however, its insulin secretory mechanism(s) remained unknown. Here, we explored the insulinotropic mechanism(s) of coixol in vitro and in vivo. Mice islets were batch incubated, perifused with coixol in the presence of agonists/antagonists, and insulin secretion was measured by ELISA. Intracellular cAMP levels were measured using enzyme immunoassay. K+- and Ca2+-currents were recorded in MIN6 cells using whole-cell patch-clamp technique. The in vivo glucose tolerance and the insulinogenic index were evaluated in diabetic rats treated with coixol at 25 and 50 mg/kg, respectively. Coixol, unlike sulfonylurea, enhanced insulin secretion in batch incubated and perifused islets at high glucose, with no effect at basal glucose concentrations. Coixol showed no pronounced effect on the inward rectifying K+- and Ca2+-currents in whole-cell patch recordings. Moreover, coixol-induced insulin secretion was further amplified in the depolarized islets. Coixol showed an additive effect with forskolin (10 µM)-induced cAMP level, and in insulin secretion; however, no additive effect was observed with isobutylmethylxanthine (IBMX, 100 µM)-induced cAMP level, nor in insulin secretion. The PKA inhibitor H-89 (50 µM), and Epac2 inhibitor MAY0132 (50 µM) significantly inhibited the coixol-induced insulin secretion (P < 0.01). Furthermore, insulin secretory kinetics revealed that coixol potentiates insulin secretion in both early and late phases of insulin secretion. In diabetic animals, coixol showed significant improvement in glucose tolerance and on fasting blood glucose levels. These data suggest that coixol amplifies glucose-stimulated insulin secretion by cAMP-mediated signaling pathways.


Assuntos
Benzoxazóis/farmacologia , AMP Cíclico/metabolismo , Glucose/farmacologia , Secreção de Insulina/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Glicemia/metabolismo , Canais de Cálcio/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Experimental/fisiopatologia , Fenômenos Eletrofisiológicos/efeitos dos fármacos , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Potássio/metabolismo , Ratos
8.
J Pharmacol Exp Ther ; 370(2): 172-181, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31182471

RESUMO

The GPR40/FFA1 receptor is a G-protein-coupled receptor expressed in the pancreatic islets and enteroendocrine cells. Here, we report the pharmacological profiles of (3S)-3-cyclopropyl-3-{2-[(1-{2-[(2,2-dimethylpropyl)(6-methylpyridin-2-yl)carbamoyl]-5-methoxyphenyl}piperidin-4-yl)methoxy]pyridin-4-yl}propanoic acid (SCO-267), a novel full agonist of GPR40. Ca2+ signaling and insulin and glucagon-like peptide-1 (GLP-1) secretion were evaluated in GPR40-expressing CHO, MIN6, and GLUTag cells. Hormone secretions and effects on fasting glucose were tested in rats. Single or repeated dosing effects were evaluated in neonatally streptozotocin-induced diabetic rats (N-STZ-1.5 rats), diet-induced obese (DIO) rats, and GPR40-knockout (Ffar1-/- ) mice. Treatment with SCO-267 activated Gq signaling in both high- and low-FFAR1-expressing CHO cells, stimulated insulin secretion in MIN6 cells, and induced GLP-1 release in GLUTag cells. When administered to normal rats, SCO-267 increased insulin, glucagon, GLP-1, glucose-dependent insulinotropic peptide, and peptide YY (PYY) secretions under nonfasting conditions. These results show the full agonistic property of SCO-267 against GPR40. Hypoglycemia was not induced in SCO-267-treated rats during the fasting condition. In diabetic N-STZ-1.5 rats, SCO-267 was highly effective in improving glucose tolerance in single and 2-week dosing studies. DIO rats treated with SCO-267 for 2 weeks showed elevated plasma GLP-1 and PYY levels, reduced food intake, and decreased body weight. In wild-type mice, SCO-267 induced GLP-1 secretion, food intake inhibition, and body weight reduction; however, these effects were abolished in Ffar1-/- mice, indicating a GPR40-dependent mechanism. In conclusion, SCO-267 stimulated islet and gut hormone secretion, improved glycemic control in diabetic rats, and decreased body weight in obese rats. These data suggest the therapeutic potential of SCO-267 for the treatment of diabetes and obesity.


Assuntos
Glicemia/metabolismo , Peso Corporal/efeitos dos fármacos , Ciclopropanos/farmacologia , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/tratamento farmacológico , Obesidade/complicações , Piperidinas/farmacologia , Propionatos/farmacologia , Receptores Acoplados a Proteínas-G/agonistas , Animais , Células CHO , Cricetulus , Ciclopropanos/uso terapêutico , Diabetes Mellitus Experimental/complicações , Modelos Animais de Doenças , Cães , Ingestão de Alimentos/efeitos dos fármacos , Peptídeo 1 Semelhante ao Glucagon/metabolismo , Humanos , Secreção de Insulina/efeitos dos fármacos , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Piperidinas/uso terapêutico , Propionatos/uso terapêutico , Ratos
9.
Islets ; 11(3): 51-64, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31084524

RESUMO

The aim of the study was to determine the acute contribution of fuel oxidation in mediating the increase in insulin secretion rate (ISR) in response to fatty acids. Measures of mitochondrial metabolism, as reflected by oxygen consumption rate (OCR) and cytochrome c reduction, calcium signaling, and ISR by rat islets were used to evaluate processes stimulated by acute exposure to palmitic acid (PA). The contribution of mitochondrial oxidation of PA was determined in the presence and absence of a blocker of mitochondrial transport of fatty acids (etomoxir) at different glucose concentrations. Subsequent to increasing glucose from 3 to 20 mM, PA caused small increases in OCR and cytosolic calcium (about 20% of the effect of glucose). In contrast, the effect of PA on ISR was almost 3 times that by glucose, suggesting that the metabolism of PA is not the dominant mechanism mediating PA's effect on ISR. This was further supported by lack of inhibition of PA-stimulated OCR and ISR when blocking entry of PA into mitochondria (with etomoxir), and PA's lack of stimulation of reduced cytochrome c in the presence of high glucose. Consistent with the lack of metabolic stimulation by PA, an inhibitor of calcium release from the endoplasmic reticulum, but not a blocker of L-type calcium channels, abolished the PA-induced elevation of cytosolic calcium. Notably, ISR was unaffected by thapsigargin showing the dissociation of endoplasmic reticulum calcium release and second phase insulin secretion. In conclusion, stimulation of ISR by PA was mediated by mechanisms largely independent of the oxidation of the fuel.


Assuntos
Cálcio/metabolismo , Retículo Endoplasmático/metabolismo , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Palmitatos/metabolismo , Animais , Retículo Endoplasmático/efeitos dos fármacos , Insulina/agonistas , Ilhotas Pancreáticas/efeitos dos fármacos , Masculino , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Oxirredução/efeitos dos fármacos , Consumo de Oxigênio/efeitos dos fármacos , Palmitatos/farmacologia , Ratos , Ratos Sprague-Dawley
10.
Transplant Proc ; 51(5): 1451-1457, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31079939

RESUMO

BACKGROUND: Quercetin (QE) is an antioxidant, anti-inflammatory, flavonoid compound. It was shown that islets are susceptible to oxidative stress due to their inherent low antioxidant capacity. In the present study, we investigated whether treatment of mouse islets with QE could enhance their function before transplantation. METHODS: Balb/c mouse islets were treated with various concentrations of QE and their viability, function, and nitric oxide (NO) and the expression of inducible NO synthase (iNOS) were determined before and after cytokine treatment. The expression of antioxidant genes was determined. Apoptosis and apoptosis-associated gene expression was measured using INS-1 cells with or without QE treatment before and after cytokine treatment. RESULTS: The QE-treated islets and INS-1 cells showed higher cell function compared to untreated control. The expression of heme oxygenase-1, manganese-dependent superoxide dismutase, and B-cell lymphoma-2 (Bcl-2) were enhanced, and the expression of NO, iNOS, and Bcl-2-associated X protein were reduced before and after cytokine treatment. CONCLUSIONS: Our results show that QE could enhance the viability and reduce apoptosis of mouse islets and improve their function before transplantation.


Assuntos
Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Transplante das Ilhotas Pancreáticas , Ilhotas Pancreáticas/efeitos dos fármacos , Quercetina/farmacologia , Animais , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Camundongos , Camundongos Endogâmicos BALB C , Estresse Oxidativo/efeitos dos fármacos
11.
Ann Anat ; 224: 153-160, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31108190

RESUMO

The effect of empagliflozin (EMPA), a sodium-glucose cotransporter 2 inhibitor (SGLT2i), on the structure of endocrine pancreas in pre-diabetes (Pre-DM) is not yet elucidated. In the current study the relatively enlarged islets of Langerhans seen in the Pre-DM group was restored to control size by administration of EMPA. In addition the disbalance in the percentage of ß-cells and α-cells in islets of the Pre-DM was corrected in the Pre-DM + EMPA group with reversal of the significantly increased islet mass, ß-cell mass and neogenesis. Administrating EMPA in Pre-DM decreased level of caspase-3, increased that of Bcl-2 to control level and reduced the significantly increased inflammatory cytokines to levels approximated to those of the control group. In Pre-DM + EMPA group, EMPA had efficiently restored the significantly impaired glucose hemostasis to levels nearly similar to those of the control animals. This may indicate that the modulatory effect of EMPA on cells of the islets in Pre-DM is associated with a local pleotropic effect on inflammatory cytokines.


Assuntos
Compostos Benzidrílicos/uso terapêutico , Diabetes Mellitus Experimental/patologia , Glucosídeos/uso terapêutico , Ilhotas Pancreáticas/efeitos dos fármacos , Estado Pré-Diabético/patologia , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Animais , Área Sob a Curva , Compostos Benzidrílicos/farmacologia , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Caspase 3/efeitos dos fármacos , Caspase 3/metabolismo , Proliferação de Células , Citocinas/efeitos dos fármacos , Citocinas/metabolismo , Diabetes Mellitus Experimental/tratamento farmacológico , Glucagon/sangue , Células Secretoras de Glucagon/citologia , Células Secretoras de Glucagon/efeitos dos fármacos , Teste de Tolerância a Glucose , Glucosídeos/farmacologia , Homeostase , Imuno-Histoquímica , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/efeitos dos fármacos , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/patologia , Masculino , Estado Pré-Diabético/tratamento farmacológico , Proteínas Proto-Oncogênicas c-bcl-2/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos , Ratos Sprague-Dawley , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia
12.
Nat Immunol ; 20(6): 677-686, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31110312

RESUMO

Consumption of a high-energy Western diet triggers mild adaptive ß cell proliferation to compensate for peripheral insulin resistance; however, the underlying molecular mechanism remains unclear. In the present study we show that the toll-like receptors TLR2 and TLR4 inhibited the diet-induced replication of ß cells in mice and humans. The combined, but not the individual, loss of TLR2 and TLR4 increased the replication of ß cells, but not that of α cells, leading to enlarged ß cell area and hyperinsulinemia in diet-induced obesity. Loss of TLR2 and TLR4 increased the nuclear abundance of the cell cycle regulators cyclin D2 and Cdk4 in a manner dependent on the signaling mediator Erk. These data reveal a regulatory mechanism controlling the proliferation of ß cells in diet-induced obesity and suggest that selective targeting of the TLR2/TLR4 pathways may reverse ß cell failure in patients with diabetes.


Assuntos
Células Secretoras de Insulina/metabolismo , Obesidade/etiologia , Obesidade/metabolismo , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética , Animais , Proliferação de Células , Ciclina D2/metabolismo , Quinase 4 Dependente de Ciclina/metabolismo , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Feminino , Humanos , Insulina/sangue , Insulina/metabolismo , Células Secretoras de Insulina/ultraestrutura , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/metabolismo , Sistema de Sinalização das MAP Quinases , Masculino , Camundongos , Camundongos Knockout , Complexos Multiproteicos/metabolismo , Obesidade/tratamento farmacológico , Parabiose , Ligação Proteica , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo
13.
Morphologie ; 103(341 Pt 2): 80-93, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31101500

RESUMO

This study evaluated the antidiabetic potentials of flavonoid-rich aqueous fraction of methanolic extract of Hibiscus sabdariffa calyx (HSCE) on the pancreatic ß-cells of experimental type I diabetic model rats. Type 1 diabetes mellitus was induced in Wistar rats by a single intraperitoneal injection of 80mg/kg b/w streptozotocin (STZ) dissolved in 0.1M citrate buffer (pH 6.3). The rats were divided into five groups (n=12) including normal control group, test group I, diabetic negative control, test group II, and diabetic positive control. The test groups received 1.75g/kg b/w of HSCE by gavage for 15 days. Animals were sacrificed; the splenic portion of their pancreas and serum were evaluated for histopathological and biochemical parameters respectively. The regenerative effects of the extract on STZ-diabetes ß-cells damage was evident from the results of the histopathological analysis and the biochemical parameters evaluated in the serum. Reduced levels of glutathione, catalase and superoxide dismutase in the serum of diabetic rats were significantly improved in the H. sabdariffa-treated rats (P<0.05). Histological examination of pancreatic islet sections revealed degenerative and necrotic changes (D) in the pancreatic islet of Langerhans, ß-cell degranulation, pyknotic ß-cell nuclei, decreased islet cellular density, and severe vacuolation (V) in the islet of STZ-diabetic negative control group. The morphology of the pancreas of HSCE-treated diabetic rats (test group II) revealed remarkable improvements in the islet of Langerhans. Stereological studies also revealed that HSCE-treatment remarkably improved volume of the pancreatic islets and the numerical density of ß-cell (number of ß-cells per unit area of islet) depleted by STZ diabetes. The study concluded that possible antidiabetic mechanism of Hibiscus sabdariffa in STZ diabetes is through induction of ß-cell regeneration and its strong antioxidant potential.


Assuntos
Antioxidantes/farmacologia , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Tipo 1/tratamento farmacológico , Hibiscus/química , Extratos Vegetais/farmacologia , Administração Oral , Animais , Antioxidantes/uso terapêutico , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 1/induzido quimicamente , Diabetes Mellitus Tipo 1/patologia , Feminino , Flores/química , Humanos , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/patologia , Masculino , Extratos Vegetais/uso terapêutico , Ratos , Ratos Wistar , Regeneração/efeitos dos fármacos , Estreptozocina/toxicidade , Resultado do Tratamento
14.
Biomater Sci ; 7(6): 2308-2316, 2019 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-31011732

RESUMO

Islet transplantation is one of the most promising therapeutic options that could restore euglycaemia in type 1 diabetic individuals. The currently implemented alginate microsphere islet encapsulation approach has led to positive outcomes in improving intraportal islet delivery in rodents. However, results obtained from human clinical trials remain disappointing. The less than satisfactory clinical outcome is mainly attributable to the increased size of the encapsulated islet and alginate-elicited host immune responses. In order to achieve islet encapsulation without significant alteration of the islet size, we designed and prepared a chondroitin sulfate (CS)-incorporated starPEG nanocoating (CS-PEG) that conjugates covalently to the pancreatic islet cell surface amine groups via pseudo-orthogonal chemistry for islet surface engineering. CS-PEG surface engineering incurred minimal alteration of the islet volume. Enhanced in situ revascularization, which is protective against extracellular matrix disruption, was also observed from CS-PEG islets in addition to robust islet viability and uncompromised insulin secretory ability. More importantly, CS-PEG surface engineering reduced blood coagulation while facilitating islet cell survival under pro-inflammatory conditions. Given that host immune rejection and an instant blood-mediated inflammatory reaction (IBMIR) are the primary factors detrimental to islet engraftment and survival, often resulting in rapid cell loss and graft failure, CS-PEG surface engineering provides an "easy-to-adopt" approach for cell surface engineering that could potentially improve the clinical efficacy of islet transplantation and other types of cell therapies.


Assuntos
Sulfatos de Condroitina/química , Sulfatos de Condroitina/farmacologia , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/efeitos dos fármacos , Nanoestruturas/química , Polietilenoglicóis/química , Engenharia Tecidual/métodos , Animais , Sobrevivência Celular/efeitos dos fármacos , Masculino , Camundongos , Neovascularização Fisiológica/efeitos dos fármacos
15.
Nutrients ; 11(4)2019 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-30987324

RESUMO

Alcalase- generated potato protein hydrolysate (APPH) is a potential bioactive peptide against diabetes mellitus (DM) and DM-associated secondary effects in animal models. The aim of the present study was to find the efficiency of a deca-peptide DIKTNKPVIF (DF) from APPH against DM. Six-week-old male ICR mice were divided into the following groups: Control, Control+DF (received 50 mg/kg DF), streptozotocin (STZ)-induced DM group, DM+Acarbose group (20 mg/kg of acarbose), DM+DF-L (25 mg/kg of DF), DM+DF-H (50 mg/kg of DF), and DM+APPH (50 mg/kg of APPH). Comparable to APPH, treatment with DF effectively regulated blood glucose level and also controlled plasma total glycerol (TG), total cholesterol (TC), insulin, and HbA1c levels in DM animals. DF treatment also showed evidence of ameliorating DM-associated damages in the pancreatic islets and in the liver, heart, and kidney tissues. Therefore, the results demonstrate that the short synthetic peptide-DF may effectively provide protection against DM-associated damages.


Assuntos
Glicemia/efeitos dos fármacos , Diabetes Mellitus Experimental/prevenção & controle , Hipoglicemiantes/farmacologia , Ilhotas Pancreáticas/efeitos dos fármacos , Oligopeptídeos/farmacologia , Proteínas de Plantas/metabolismo , Hidrolisados de Proteína/metabolismo , Solanum tuberosum/metabolismo , Animais , Biomarcadores/sangue , Glicemia/metabolismo , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/patologia , Hemoglobina A Glicada/metabolismo , Hipoglicemiantes/metabolismo , Insulina/sangue , Ilhotas Pancreáticas/metabolismo , Ilhotas Pancreáticas/patologia , Lipídeos/sangue , Masculino , Camundongos Endogâmicos ICR , Oligopeptídeos/metabolismo , Estreptozocina , Subtilisinas/metabolismo
16.
Cell Physiol Biochem ; 52(3): 486-502, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30873823

RESUMO

BACKGROUND/AIMS: Cross-talk between different pancreatic islet cell types regulates islet function and somatostatin (SST) released from pancreatic delta cells inhibits insulin secretion from pancreatic beta cells. In other tissues SST exhibits both protective and pro-apoptotic properties in a tissue-specific manner, but little is known about the impact of the peptide on beta cell survival. Here we investigate the specific role of SST in the regulation of beta cell survival in response to physiologically relevant inducers of cellular stress including palmitate, cytokines and glucose. METHODS: Pancreatic MIN6 beta cells and primary mouse islet cells were pre-treated with SST with or without the Gi/o signalling inhibitor, pertussis toxin, and exposed to different cellular stress factors. Apoptosis and proliferation were assessed by measurement of caspase 3/7 activity, TUNEL and BrdU incorporation, respectively, and expression of target genes was measured by qPCR. RESULTS: SST partly alleviated upregulation of cellular stress markers (Hspa1a and Ddit3) and beta cell apoptosis in response to factors such as lipotoxicity (palmitate), pro-inflammatory cytokines (IL1ß and TNFα) and low glucose levels. This effect was mediated via a Gi/o protein-dependent pathway, but did not modify transcriptional upregulation of the specific NFκB-dependent genes, Nos2 and Ccl2, nor was it associated with transcriptional changes in SST receptor expression. CONCLUSION: Our results suggest an underlying protective effect of SST which modulates the beta cell response to ER stress and apoptosis induced by a range of cellular stressors associated with type 2 diabetes.


Assuntos
Proliferação de Células/efeitos dos fármacos , Glucose/antagonistas & inibidores , Células Secretoras de Insulina/efeitos dos fármacos , Ilhotas Pancreáticas/efeitos dos fármacos , Toxina Pertussis/antagonistas & inibidores , Somatostatina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Caspase 3/genética , Caspase 3/metabolismo , Caspase 7/genética , Caspase 7/metabolismo , Linhagem Celular , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Regulação da Expressão Gênica , Glucose/farmacologia , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/metabolismo , Interleucina-1beta/antagonistas & inibidores , Interleucina-1beta/farmacologia , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos ICR , NF-kappa B/genética , NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Ácido Palmítico/antagonistas & inibidores , Ácido Palmítico/farmacologia , Toxina Pertussis/farmacologia , Técnicas de Cultura de Tecidos , Fator de Transcrição CHOP/genética , Fator de Transcrição CHOP/metabolismo , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/farmacologia
17.
Islets ; 11(4): 77-88, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30849280

RESUMO

A potentiating effect of medium-chain triglycerides on glucose-stimulated insulin secretion (GSIS) has been observed since the 1960s. Subsequent observations identified octanoic acid (OA), the main component of medium-chain triglyceride, as the potentiator of GSIS, but the mechanism was unclear. We used wild-type (WT), short-chain 3-hydroxyacyl-CoA dehydrogenase knockout (Hadh-/-), and sulfonylurea receptor 1 knockout (Sur1-/-) mouse islets to define the mechanism of OA potentiation of insulin secretion. Application of OA alone induced a 2- to 3- fold increase of insulin secretion with an apparent threshold of 3 mM in WT mouse islets, suggesting that OA itself is a weak insulin secretagogue. However, OA at 1 mM strongly potentiated fuel-stimulated insulin secretion, especially GSIS. The potentiating effect on fuel-stimulated insulin secretion by OA did not require fatty acid ß-oxidation because OA also potentiated amino acid-stimulated insulin secretion in islets isolated from Hadh-/- mice, which cannot fully oxidize OA. Measurements using Sur1-/- islets indicated that the potentiating effect of OA on fuel-stimulated insulin secretion is Ca2+ dependent and is often accompanied by ß-cell membrane potential depolarization, and may also involve the Ca2+/calmodulin complex. Experiments using DCPIB, an ethacrynic acid derivative, to inhibit volume-sensitive anion channels (VSACs) in Sur1-/- islets demonstrated that the potentiation effects of OA on insulin secretion are in part medicated by activation of VSAC. In addition, inhibition of IP3 receptor also abolishes the OA-induced intracellular Ca2+ increase in Sur1-/- islets.


Assuntos
Caprilatos/farmacologia , Secreção de Insulina/efeitos dos fármacos , Ilhotas Pancreáticas/efeitos dos fármacos , Animais , Células Cultivadas , Sinergismo Farmacológico , Glucose/farmacologia , Insulina/metabolismo , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais/efeitos dos fármacos
18.
Amino Acids ; 51(4): 727-738, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30830312

RESUMO

Obesity in fathers leads to DNA damage and epigenetic changes in sperm that may carry potential risk factors for metabolic diseases to the next generation. Taurine (TAU) supplementation has demonstrated benefits against testicular dysfunction and pancreatic islet impairments induced by obesity, but it is not known if these protective actions prevent the propagation of metabolic disruptions to the next generation; as such, we hypothesized that paternal obesity may increase the probability of endocrine pancreatic dysfunction in offspring, and that this could be prevented by TAU supplementation in male progenitors. To test this, male C57Bl/6 mice were fed on a control diet (CTL) or a high-fat diet (HFD) without or with 5% TAU in their drinking water (CTAU and HTAU) for 4 months. Subsequently, all groups of mice were mated with CTL females, and the F1 offspring were identified as: CTL-F1, CTAU-F1, HFD-F1, and HTAU-F1. HFD-fed mice were normoglycemic, but glucose intolerant and their islets hypersecreted insulin. However, at 90 days of age, HFD-F1 offspring displayed normal glucose homeostasis and adiposity, but reduced glucose-induced insulin release. HFD-F1 islets also exhibited ß- and α-cell hypotrophy, and lower δ-cell number per islet. Paternal TAU supplementation prevented the decrease in glucose-induced insulin secretion and normalized ß-cell size and δ-cell number, and increased α-cell size/islet in HTAU-F1 mice. In conclusion, HFD consumption by male founders decreases ß-cell secretion and islet-cell distribution in their offspring. TAU attenuates the deleterious effects of paternal obesity on insulin secretion and islet-cell morphology in F1 offspring.


Assuntos
Dieta Hiperlipídica/efeitos adversos , Suplementos Nutricionais , Sistema Endócrino/efeitos dos fármacos , Intolerância à Glucose/tratamento farmacológico , Ilhotas Pancreáticas/efeitos dos fármacos , Pancreatopatias/tratamento farmacológico , Taurina/administração & dosagem , Animais , Sistema Endócrino/fisiopatologia , Intolerância à Glucose/etiologia , Intolerância à Glucose/patologia , Homeostase , Secreção de Insulina , Ilhotas Pancreáticas/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/fisiopatologia , Pancreatopatias/etiologia , Pancreatopatias/patologia
19.
J Diabetes Res ; 2019: 5245063, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30863781

RESUMO

Increasing evidence shows that statins increase the risk of new-onset diabetes mellitus, but the exact mechanism is not clearly known. Free fatty acid receptor 1 (FFA1) has been recognized to mediate insulin secretion, and pioglitazone has direct effects on glucose-stimulated insulin secretion in addition to the reversion of insulin resistance. In this study, we found that atorvastatin decreased potassium-stimulated insulin secretion and inhibited the expression of FFA1, PDX-1, and BETA2/NeuroD in INS-1 cells. Further study demonstrated that pioglitazone prevented the impairment of insulin secretion induced by atorvastatin and enhanced the expression of FFA1, PDX-1, and BETA2/NeuroD reduced by atorvastatin in INS-1 cells. In addition, the preventive effect of pioglitazone on atorvastatin-induced impairment of insulin secretion and the enhancement of the expression of PDX-1 and BETA2/NeuroD was abolished by knockdown of FFA1 using siRNA or the PLC inhibitor, U-73122, respectively. Ultimately, FFA1 may mediate the atorvastatin-induced pancreatic ß-cell dysfunction and pioglitazone may ameliorate this deleterious effect through the upregulation of FFA1 expression.


Assuntos
Atorvastatina/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Hipoglicemiantes/farmacologia , Células Secretoras de Insulina/efeitos dos fármacos , Ilhotas Pancreáticas/efeitos dos fármacos , Pioglitazona/farmacologia , Receptores Acoplados a Proteínas-G/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Linhagem Celular , Proteínas de Homeodomínio/metabolismo , Insulina/metabolismo , Resistência à Insulina , Células Secretoras de Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Ratos , Transativadores/metabolismo
20.
Nutrients ; 11(3)2019 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-30871106

RESUMO

Low birth weight is a risk factor for gestational and type 2 diabetes (T2D). Since mammalian target of rapamycin (mTOR) controls pancreatic ß-cell mass and hormone release, we hypothesized that nutritional insult in utero might permanently alter mTOR signaling. Mice were fed a low-protein (LP, 8%) or control (C, 20%) diet throughout pregnancy, and offspring examined until 130 days age. Mice receiving LP were born 12% smaller and ß-cell mass was significantly reduced throughout life. Islet mTOR levels were lower in LP-exposed mice and localized predominantly to α-rather than ß-cells. Incubation of isolated mouse islets with rapamycin significantly reduced cell proliferation while increasing apoptosis. mRNA levels for mTORC complex genes mTOR, Rictor and Raptor were elevated at 7 days in LP mice, as were the mTOR and Raptor proteins. Proglucagon gene expression was similarly increased, but not insulin or the immune/metabolic defense protein STING. In human and mouse pancreas STING was strongly associated with islet ß-cells. Results support long-term changes in islet mTOR signaling in response to nutritional insult in utero, with altered expression of glucagon and insulin and a reduced ß-cell mass. This may contribute to an increased risk of gestational or type 2 diabetes.


Assuntos
Dieta com Restrição de Proteínas , Proteínas na Dieta/administração & dosagem , Glucagon/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Fenômenos Fisiológicos da Nutrição Pré-Natal , Serina-Treonina Quinases TOR/metabolismo , Animais , Linhagem Celular , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Glucagon/genética , Insulina/genética , Insulina/metabolismo , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Gravidez , Distribuição Aleatória , Serina-Treonina Quinases TOR/genética
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