RESUMO
PURPOSE: To evaluate whether a second biopsy, following a first diagnostic failure on blastocysts tested for preimplantation genetic testing for monogenic diseases (PGT-M), allows to obtain genetic diagnosis and to what extent this procedure can influence clinical pregnancy and live birth rates compared to the PGT-M process with a successful genetic diagnosis from the first biopsy. METHODS: Embryos from women who underwent PGT-M in an infertility centre and who had been transferred after two biopsies for genetic analysis (n = 27) were matched in a 1:1 ratio accordingly to women's age (± 1 year) and fertility status (fertile vs infertile), as well as with the study period, with embryos who were transferred after receiving a conclusive PGT result straight after the first biopsy (n = 27). The main evaluated outcome was clinical pregnancy rate following embryo transfers in which healthy embryos were transferred after only one biopsy and those in which an embryo was transferred after being re-biopsied. Live birth rate was the secondary outcome. RESULTS: Clinical pregnancy rate was 52% (95% CI: 34-69) following the transfer of a single-biopsy blastocyst and 30% (95% CI: 16-48) following the transfer of a re-biopsied blastocyst. The likelihood to have a healthy baby was 33% (95% CI: 19-52) following the transfer of a blastocyst biopsied once and 22% (95% CI: 11-41) following the transfer of a re-biopsied blastocyst. CONCLUSIONS: The re-biopsy intervention seems to considerably reduce the pregnancy potential of a blastocyst. However, a greater sample size is necessary to clarify this issue definitively.
Assuntos
Embrião de Mamíferos , Humanos , Biópsia , Embrião de Mamíferos/metabolismo , Implantação do Embrião , Testes Genéticos , Gravidez , Masculino , Adulto , Técnicas de Reprodução Assistida , Estudos de Casos e Controles , Resultado da Gravidez , Infertilidade FemininaRESUMO
Glutamate is the major excitatory neurotransmitter of the central nervous system (CNS) and may induce cytotoxicity through persistent activation of glutamate receptors and oxidative stress. Its extracellular concentration is maintained at physiological concentrations by high affinity glutamate transporters of the solute carrier 1 family (SLC1). Glutamate is also present in islet of Langerhans where it is secreted by the α-cells and acts as a signaling molecule to modulate hormone secretion. Whether glutamate plays a role in islet cell viability is presently unknown. We demonstrate that chronic exposure to glutamate exerts a cytotoxic effect in clonal ß-cell lines and human islet ß-cells but not in α-cells. In human islets, glutamate-induced ß-cell cytotoxicity was associated with increased oxidative stress and led to apoptosis and autophagy. We also provide evidence that the key regulator of extracellular islet glutamate concentration is the glial glutamate transporter 1 (GLT1). GLT1 localizes to the plasma membrane of ß-cells, modulates hormone secretion, and prevents glutamate-induced cytotoxicity as shown by the fact that its down-regulation induced ß-cell death, whereas GLT1 up-regulation promoted ß-cell survival. In conclusion, the present study identifies GLT1 as a new player in glutamate homeostasis and signaling in the islet of Langerhans and demonstrates that ß-cells critically depend on its activity to control extracellular glutamate levels and cellular integrity.
Assuntos
Transportador 2 de Aminoácido Excitatório/biossíntese , Regulação da Expressão Gênica , Proteínas de Transporte de Glutamato da Membrana Plasmática/biossíntese , Células Secretoras de Insulina/citologia , Animais , Apoptose , Autofagia , Sobrevivência Celular , Transportador 2 de Aminoácido Excitatório/fisiologia , Proteínas de Transporte de Glutamato da Membrana Plasmática/fisiologia , Ácido Glutâmico/química , Ácido Glutâmico/metabolismo , Homeostase , Humanos , Ilhotas Pancreáticas/citologia , Camundongos , Modelos Biológicos , Estresse OxidativoRESUMO
BACKGROUND: Several studies have associated elevated liver enzymes (LFTs), obesity, and type 2 diabetes (T2DM), and a link has been established between insulin resistance (IR) and elevated ferritin concentrations. We examined the relationship between LFTs, ferritin, and IR in morbid obese subjects and the effect of weight loss after bariatric surgery. METHODS: We measured liver enzymes, ferritin, insulin resistance, and glucose tolerance (by OGTT) in 159 morbid obese subjects (BMI = 44.4 +/- 0.4 kg/m(2)) at baseline, 6 months and 1 year after laparoscopic-adjustable-gastric banding (LAGB). Subjects were divided in two groups: increased LFTs (ALT > 30; AST/ALT < 1) vs. normal LFTs. RESULTS: A large proportion of morbid obese subjects had increased LFTs (44%) which were associated with increased IR and ferritin, suggesting potential liver disease. A majority of the morbidly obese with increased LFTs, IGT, and T2DM, were male and had almost double ferritin concentrations, strongly correlated with ALT (r = 0.43, p < 0.0001). Both ferritin and ALT correlated with waist circumference and IR. One year after, LAGB glucose tolerance improved, LFTs and IR were reduced; ferritin did not change significantly, but was still correlated with IR. CONCLUSIONS: Ferritin may be an additional useful marker for more severe hepatic IR.
Assuntos
Ferritinas/sangue , Gastroplastia , Resistência à Insulina/fisiologia , Obesidade Mórbida/metabolismo , Transaminases/sangue , Redução de Peso/fisiologia , Adulto , Índice de Massa Corporal , Estudos de Coortes , Feminino , Humanos , Laparoscopia , Testes de Função Hepática , Masculino , Obesidade Mórbida/cirurgia , Estudos RetrospectivosRESUMO
Ac-SDKP (N-acetyl-seryl-aspartyl-lysyl-proline) is a physiological tetrapeptide hydrolysed by ACE (angiotensin-converting enzyme). In experimental models of hypertension, Ac-SDKP has antifibrotic effects in the heart; however, the role of Ac-SDKP in diabetic cardiomyopathy is currently unknown. The aim of the present study was to evaluate the effect of Ac-SDKP on cardiac systolic and diastolic function, and interstitial and perivascular fibrosis in the heart of diabetic rats.Diabetes was induced in 55 Sprague-Dawley rats by streptozotocin injection. Control rats (n=18)underwent only buffer injection.Out of the 55 diabetic rats, 19 were chronically treated with insulin and 13 with the ACEI (ACE inhibitor) ramipril (3 mg x kg(-1 )of body weight x day(-1)). At 2 months after the onset of diabetes, Ac-SDKP (1 mg x kg(-1) of body weight x day(-1)) was administered by osmotic minipumps for 8 weeks to eight control rats, 13 diabetic rats, seven diabetic rats treated with ramipril and nine insulin-treated diabetic rats. Diabetic rats had a significant increase in blood glucose levels. Left ventricular interstitial and perivascular fibrosis, and TGF-beta1 (transforming growth factor-beta1) protein levels were increased in diabetic rats, but not in insulin-treated diabetic rats and ramipril-treated diabetic rats, compared with control rats. Ac-SDKP administration significantly reduced left ventricular interstitial and perivascular fibrosis in diabetic rats and in diabetic rats treated with ramipril. This was accompanied by a significant reduction in active TGF-beta1 and phospho-Smad2/3 protein levels in myocardial tissue of diabetic rats. Echocardiography showed that diabetes was associated with increased end-systolic diameters, and depressed global systolic function and diastolic dysfunction, as assessed by transmitral Doppler velocity profile. These changes were completely reversed by insulin or ramipril treatment. Ac-SDKP treatment partially restored diastolic function in diabetic rats. In conclusion, Ac-SDKP administration in diabetic rats reduces left ventricular interstitial and perivascular fibrosis, active TGF-beta1 and phospho-Smad2/3levels, and improves diastolic function. Taken together, these findings suggest that, by inhibiting theTGF-beta/Smad pathway, Ac-SDKP protects against the development of diabetic cardiomyopathy
Assuntos
Complicações do Diabetes/prevenção & controle , Diabetes Mellitus Experimental/fisiopatologia , Ventrículos do Coração/patologia , Miocárdio/patologia , Oligopeptídeos/farmacologia , Inibidores da Enzima Conversora de Angiotensina/administração & dosagem , Animais , Cardiomiopatias/prevenção & controle , Fibrose , Ventrículos do Coração/efeitos dos fármacos , Hipoglicemiantes/administração & dosagem , Insulina/administração & dosagem , Masculino , Oligopeptídeos/administração & dosagem , Ramipril/administração & dosagem , Ratos , Ratos Sprague-Dawley , Proteínas Smad/efeitos dos fármacos , Proteínas Smad/metabolismo , Fator de Crescimento Transformador beta1/efeitos dos fármacos , Fator de Crescimento Transformador beta1/metabolismoRESUMO
BACKGROUND: Volume reduction of cord blood units decreases the cost of cryogenic storage. This study reports the analysis of a 10-year quality control program of a semiautomated cord blood volume reduction procedure. STUDY DESIGN AND METHODS: Cord blood was collected in a plastic bag containing 29 mL citrate-phosphate-dextrose, centrifuged at 2124 x g for 12 minutes, and processed with a semiautomated device. The procedure was aimed at removing most red blood cells and plasma and concentrating hematopoietic progenitors in the buffy coat (BC), thus reducing the unit volume and saving cryogenic space. Finally, the BC was cryopreserved with an equal volume of 20 percent dimethyl sulfoxide. Total nucleated cells (TNCs) were counted before and after processing in the 4311 units banked from 1998 through 2007, whereas CD34+ cells and colony-forming units-granulocyte-macrophage (CFU-GM) were counted in 420 random units from 2001 through 2007. RESULTS: Mean postvolume reduction annual recoveries of TNCs, CD34+ cells, and CFU-GM ranged from 82.8 +/- 12.3 (standard deviation) to 91.4 +/- 6.4 percent, from 87.8 +/- 14.1 to 95.2 +/- 23.8 percent, and from 101.5 +/- 51.4 to 117.8 +/- 59.5 percent, respectively. Very strong correlations were found (r > 0.87) between postprocessing versus preprocessing TNCs, CD34+ cells, and CFU-GM; a moderate correlation between initial TNC count and unit's volume (r = 0.51); and no correlation between TNC percentage of recovery in the BC and initial unit's volume. The latter data indicate that most TNCs concentrate in the BC. CONCLUSIONS: The semiautomated procedure of cord blood unit volume reduction used in this study provides high and stable cellular recoveries during several years of routine cord blood banking.
Assuntos
Preservação de Sangue/métodos , Sangue Fetal , Automação , Preservação de Sangue/economia , Ensaio de Unidades Formadoras de Colônias , Humanos , Controle de Qualidade , Fatores de TempoRESUMO
Essential hypertension is an insulin resistant state. Early insulin signaling steps are impaired in essential hypertension and a large body of data suggests that there is a crosstalk at multiple levels between the signal transduction pathways that mediate insulin and angiotensin II actions. At the extracellular level the angiotensin converting enzyme (ACE) regulates the synthesis of angiotensin II and bradykinin that is a powerful vasodilator. At early intracellular level angiotensin II acts on JAK-2/IRS1-IRS2/PI3-kinase, JNK and ERK to phosphorylate serine residues of key elements of insulin signaling pathway therefore inhibiting signaling by the insulin receptor. On another level angiotensin II inhibits the insulin signaling inducing the regulatory protein SOCS 3. Angiotensin II acting through the AT1 receptor can inhibit insulin-induced nitric oxide (NO) production by activating ERK 1/2 and JNK and enhances the activity of NADPH oxidase that leads to an increased reactive oxygen species generation. From the clinical standpoint, the inhibition of the renin angiotensin system improves insulin sensitivity and decreases the incidence of Type 2 Diabetes Mellitus (T2DM). This might represent an alternative approach to prevent type 2 diabetes in patients with hypertension and metabolic syndrome, (i.e. insulin resistant patients). This review will discuss: a) the molecular mechanisms of the crosstalk between the insulin and angiotensin II signaling systems b) the results of clinical studies employing drugs targeting the renin-angiotensin II-aldosterone systems and their role in glucose metabolism and diabetes prevention.
Assuntos
Glicemia/metabolismo , Diabetes Mellitus Tipo 2/prevenção & controle , Insulina/metabolismo , Sistema Renina-Angiotensina , Transdução de Sinais , Bloqueadores do Receptor Tipo 1 de Angiotensina II/uso terapêutico , Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Animais , Glicemia/efeitos dos fármacos , Diabetes Mellitus Tipo 2/metabolismo , Humanos , Antagonistas de Receptores de Mineralocorticoides/uso terapêutico , Sistema Renina-Angiotensina/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacosRESUMO
To learn more about nonimmune-mediated islet graft failure, we transplanted different preparations (preps) of isolated human islets under the kidney capsule of streptozotocin (STZ)-diabetic nude mice. One month after the implantation of 1,000 or 2,000 islets, grafts were harvested for morphological, immunohistochemical, and ultrastructural analysis. Only a single islet prep cured the diabetes out of all the recipients, while the remaining preps showed only partial function after the implantation of 2,000 islets. Transplanted mice showed high circulating proinsulin levels but, with the exclusion of those bearing curative grafts, relatively low mature insulin levels. Engrafted beta-cells showed positive carboxypeptidase E (CPE) and prohormone convertase 1 (PC1) staining, while prohormone convertase 2 (PC2) was undetectable. In contrast, PC2 was abundantly expressed by engrafted alpha-cells. Moreover, engrafted beta-cells did not show evidence of replication, and preapoptotic beta-cells, with intra- and extracellular amyloid deposition, were detected with electron microscopy. Cell cycle inhibitors p16(INK4), p21(WAF1), and p27(Kip1) were abundantly expressed in the islet grafts and showed a predominant nuclear localization. In conclusion, diabetic nude mice transplanted with human islets showed disproportionate hyperproinsulinemia and graft evidence of beta-cell restricted PC2 depletion, amyloid deposition and beta-cell death, and lack of beta-cell replication with nuclear translocation of p27(Kip1) and p21(WAF1) that together may contribute to delayed graft failure.
Assuntos
Ciclo Celular/fisiologia , Diabetes Mellitus Experimental/cirurgia , Hiperinsulinismo/etiologia , Transplante das Ilhotas Pancreáticas/efeitos adversos , Ilhotas Pancreáticas/enzimologia , Ilhotas Pancreáticas/metabolismo , Proinsulina/metabolismo , Pró-Proteína Convertase 2/deficiência , Transplante Heterólogo/efeitos adversos , Animais , Autopsia , Humanos , Células Secretoras de Insulina/citologia , Células Secretoras de Insulina/enzimologia , Células Secretoras de Insulina/patologia , Ilhotas Pancreáticas/patologia , Transplante das Ilhotas Pancreáticas/patologia , Masculino , Camundongos , Camundongos Nus , Pâncreas/patologia , Proinsulina/sangue , Valores de Referência , Falha de TratamentoRESUMO
Information about the mechanisms of meiotic arrest and resumption of meiosis in feline oocytes is still limited. The aim of this study was to investigate the effect of the presence of gonadotropins during IVM, on meiotic progression in relation to the status of gap junction mediated communications between oocyte and cumulus cells, to the cAMP intracellular content, and to the intra-oocyte concentration of glutathione (GSH) in feline oocytes. Our results indicated that about 50% of cumulus-oocyte complexes (COCs) showed functionally open communications at the time of collection, while the remainder were partially or totally closed. After 3h of culture, the percentage of COCs with functional gap junctions was significantly greater in the group matured in the presence of gonadotropins than in those matured without them, where an interruption of communications was observed. Moreover, this precocious uncoupling was associated with a moderate increase of cAMP concentration in the oocyte, lower than in the group exposed to gonadotropins. Intra-oocyte glutathione levels decreased significantly after 24h of IVM, whether gonadotropins were present or absent during the culturing process. The presence of thiol compounds in the IVM medium induced an intra-oocyte GSH concentration significantly higher than that found in oocytes cultured without these compounds, and similar to the GSH content of immature oocytes. Moreover, the intracellular GSH concentration increased as meiosis progressed. The present study suggests that in feline oocytes, gonadotropins affect the dynamic changes in communications between oocyte and cumulus cells during IVM. However, the intracellular concentration of GSH is not influenced by the gonadotropin stimulation. Moreover, the presence of gonadotropins and thiol compounds results in an increase of GSH levels along with meiotic progression of the oocytes.
Assuntos
Gatos , Comunicação Celular/efeitos dos fármacos , Glutationa/análise , Gonadotropinas/farmacologia , Oócitos/citologia , Folículo Ovariano/citologia , Animais , Células Cultivadas , AMP Cíclico/análise , Feminino , Junções Comunicantes , Meiose , Oócitos/química , Oócitos/efeitos dos fármacosRESUMO
OBJECTIVE: Lifestyle modifications and pharmacological interventions can prevent type 2 diabetes in obese subjects with impaired glucose tolerance. The aim of this study was to compare laparoscopic adjustable gastric banding (LAGB) and conventional diet (No-LAGB) in the prevention (primary intervention study; 56 vs. 29 patients) and remission (secondary intervention study; 17 vs. 20 patients) of type 2 diabetes and hypertension in grade 3 obesity in a 4-year study. RESEARCH DESIGN AND METHODS: The subjects (n = 122; age 48.5 +/- 1.05 years; BMI 45.7 +/- 0.67 kg/m2) underwent a diagnostic workup, including psychological and psychiatric assessments, in preparation for the LAGB procedure. Of the 122 subjects, 73 had the surgery (LAGB group). The control group (No-LAGB group) consisted of the 49 subjects who refused the surgery but agreed to be followed up; 6 of these subjects dropped out by the 2nd year of the study, so that the final number of patients was 73 and 43 in the LAGB and No-LAGB groups, respectively. All patients had a yearly visit and oral glucose tolerance test. RESULTS: From baseline to the end of the 4-year follow-up, BMI decreased from 45.9 +/- 0.89 at baseline to 37.7 +/- 0.71 kg/m2 in the LAGB group and remained steady in the No-LAGB group (from 45.2 +/- 1.04 to 46.5 +/- 1.37 kg/m2), with no significant differences between the primary and secondary intervention groups. In the primary intervention study, five of the No-LAGB subjects (17.2%) and none of the LAGB subjects (0.0%; P = 0.0001) progressed to type 2 diabetes; in the secondary intervention study, type 2 diabetes remitted in one No-LAGB patient (4.0%) and seven LAGB patients (45.0%; P = 0.0052). Hypertension occurred in 11 No-LAGB patients (25.6%) and 1 LAGB patient (1.4%; P = 0.0001) and remitted in 1 No-LAGB (2.3%) and 15 LAGB patients (20.5%; P = 0.0001). A study of body mass composition revealed a significant reduction of fat mass and a transitory, but not significant, decrease of fat-free mass in LAGB patients. CONCLUSIONS: In morbid obesity, sustained and long-lasting weight loss obtained through LAGB prevents the occurrence of type 2 diabetes and hypertension and decreases the prevalence of these disorders.
Assuntos
Diabetes Mellitus Tipo 2/prevenção & controle , Gastroplastia/métodos , Hipertensão/prevenção & controle , Laparoscopia , Obesidade Mórbida/metabolismo , Obesidade Mórbida/cirurgia , Adulto , Índice de Massa Corporal , Estudos de Casos e Controles , Feminino , Seguimentos , Humanos , Hipertensão/epidemiologia , Itália/epidemiologia , Masculino , Pessoa de Meia-Idade , Obesidade Mórbida/diagnóstico , Prevalência , Fatores de Tempo , Resultado do Tratamento , Redução de PesoRESUMO
OBJECTIVE: The C-174G promoter polymorphism of the interleukin (IL)-6 gene was found to influence transcriptional activity and plasma IL-6 levels in humans. We addressed the question of whether the C-174G IL-6 polymorphism contributes to variation of insulin sensitivity. RESEARCH DESIGN AND METHODS: Two cohorts of subjects were genotyped. Cohort 1 includes 275 nondiabetic subjects who underwent a euglycemic-hyperinsulinemic clamp. Cohort 2 includes 77 patients with morbid obesity who underwent laparoscopic adjustable gastric banding (LAGB). RESULTS: The genotypes were consistent with Hardy-Weinberg equilibrium proportions. In cohort 1, insulin sensitivity was reduced in carriers of the -174G/G genotype as compared with subjects carrying the C allele (P = 0.004). Carriers of -174G/G displayed significantly higher plasma IL-6 levels in comparison with carriers of the C allele. In a stepwise linear regression analysis, the C-174G polymorphism was independently associated with insulin sensitivity; however, after inclusion of plasma IL-6 concentrations, the polymorphism was excluded from the model explaining insulin sensitivity variability, thus suggesting that the polymorphism was affecting insulin sensitivity by regulating IL-6 plasma levels. IL-6 mRNA levels were measured by real-time RT-PCR in subcutaneous fat obtained from obese patients of cohort 2 during LAGB. Carriers of -174G/G showed increased IL-6 expression compared with subjects carrying the C allele (P = 0.04). There was a significant correlation between adipose IL-6 mRNA expression and insulin resistance assessed by homeostasis model assessment (rho = 0.28, P = 0.014). CONCLUSIONS: These results indicate that the -174G/G genotype of the IL-6 gene may contribute to variations in insulin sensitivity.
Assuntos
Resistência à Insulina/genética , Interleucina-6/genética , Obesidade Mórbida/fisiopatologia , Polimorfismo de Nucleotídeo Único , Regiões Promotoras Genéticas , Tecido Adiposo/anatomia & histologia , Adulto , Estudos de Coortes , Feminino , Derivação Gástrica , Genótipo , Humanos , Resistência à Insulina/imunologia , Masculino , Obesidade Mórbida/genética , Obesidade Mórbida/imunologia , Obesidade Mórbida/cirurgia , Valores de ReferênciaRESUMO
CONTEXT: Obesity is frequently associated with left ventricular hypertrophy, even when uncomplicated by hypertension or diabetes mellitus. Left ventricular hypertrophy is an important risk factor for congestive heart failure. OBJECTIVE: The objective of this study was to evaluate the relationship between leptin and left ventricular mass in uncomplicated, morbid (grade 3) obesity and the existence of leptin receptors and intracellular signaling proteins in the human heart. DESIGN: Left ventricular mass (LVM) was calculated through electrocardiogram reading in normotensive grade III obese patients (World Health Organization classification) undergoing bariatric surgery [laparoscopic adjustable gastric banding (LAGB)] at baseline and 1 yr later. The control group was composed of healthy lean normotensive subjects. Leptin receptors were detected by PCR and immunocytochemistry in human heart biopsies. SETTING: This study was performed at university hospitals. PATIENTS: Thirty-one grade 3 obese patients and 30 healthy nonobese normotensive, age- and sex-matched control subjects were studied. INTERVENTION: Obese subjects underwent LAGB to induce weight loss and were evaluated at baseline and after 1 yr. RESULTS: LVM, plasma leptin, glucose, insulin levels, and homeostasis model assessment index were higher in obese than in lean controls (P < 0.01); at univariate regression analysis, LVM correlated with body mass index, leptin, and homeostasis model assessment index; at multiple regression analysis, LVM only correlated with leptin levels (P = 0.001). Obese subjects were reevaluated 1 yr after LAGB, when their body mass index changed from 46.2 +/- 1.24 to 36.6 +/- 1.05 kg/m(2) (P < 0.01); the decrease in LVM correlated only with the decrease in leptin levels (P < 0.01). We demonstrated that long and short isoforms of the leptin receptor and intracellular proteins mediating leptin signaling were expressed in human heart by RT-PCR, immunocytochemistry, or both methods. CONCLUSIONS: These data suggest that leptin could contribute to the left ventricular hypertrophy in humans.
Assuntos
Hipertrofia Ventricular Esquerda/etiologia , Leptina/sangue , Obesidade Mórbida/sangue , Redução de Peso , Adulto , Índice de Massa Corporal , Feminino , Humanos , Leptina/fisiologia , Masculino , Obesidade Mórbida/cirurgia , Receptores de Superfície Celular/análise , Receptores para Leptina , Análise de RegressãoRESUMO
CONTEXT: It is unknown whether genetic factors that play an important role in body weight homeostasis influence the response to laparoscopic adjustable gastric banding (LAGB). OBJECTIVE: We investigated the impact of common polymorphisms in four candidate genes for insulin resistance on weight loss after LAGB. DESIGN: The design was a 6-month follow-up study. SETTING: The study setting was hospitalized care. PATIENTS: A total of 167 unrelated morbidly obese subjects were recruited according to the following criteria: age, 18-66 yr inclusive; and body mass index greater than 40 kg/m2 or greater than 35.0 kg/m2 in the presence of comorbidities. INTERVENTION: LAGB was used as an intervention. MAIN OUTCOME MEASURE: Measure of correlation between weight loss and common polymorphisms in candidate genes for insulin resistance and obesity was the main outcome measure. RESULTS: The following single nucleotide polymorphisms were detected by digestion of PCR products with appropriate restriction enzymes: Gly972Arg of the insulin receptor substrate-1 gene, Pro12Ala of the proliferator-activated receptor-gamma gene, C-174G in the promoter of IL-6 gene, and G-866A in the promoter of uncoupling protein 2 gene. Baseline characteristics including body mass index did not differ between the genotypes. At the 6-month follow-up after LAGB, carriers of G-174G IL-6 genotype had lost more weight than G-174C or C-174C genotype (P = 0.037), and carriers of A-866A uncoupling protein 2 genotype had lost more weight as compared with G-866G (P = 0.018) and G-866A (P = 0.035) genotype, respectively. Weight loss was lower in carriers of Gly972Arg insulin receptor substrate-1 genotype than Gly972Gly carriers, but not statistically significant (P = 0.06). No difference between carriers of Pro12Ala and Pro12Pro proliferator-activated receptor-gamma genotype was observed. CONCLUSIONS: These data demonstrate that genetic factors, which play an important role in the regulation of body weight, may account for differences in the therapeutic response to LAGB.
Assuntos
Dieta Redutora , Gastroplastia , Resistência à Insulina/genética , Obesidade Mórbida/genética , Obesidade Mórbida/cirurgia , Polimorfismo de Nucleotídeo Único , Redução de Peso/genética , Adulto , Feminino , Seguimentos , Heterozigoto , Humanos , Proteínas Substratos do Receptor de Insulina , Interleucina-6/genética , Canais Iônicos , Masculino , Proteínas de Membrana Transportadoras/genética , Pessoa de Meia-Idade , Proteínas Mitocondriais/genética , PPAR gama/genética , Fosfoproteínas/genética , Período Pós-Operatório , Proteína Desacopladora 2RESUMO
Uncoupling protein (UCP)-2 is a member of the mitochondrial inner membrane carriers that uncouple pro-ton entry in the mitochondrial matrix from ATP synthesis. The -866G/A polymorphism in the UCP2 gene, which enhances its transcriptional activity, was associated with enhanced risk for type 2 diabetes in obese subjects. We addressed the question of whether the -866G/A polymorphism contributes to variation in insulin sensitivity by genotyping 181 nondiabetic offspring of type 2 diabetic patients. Insulin sensitivity, assessed by the hyperinsulinemic-euglycemic clamp, was reduced in -866A/A carriers compared with -866A/G or -866G/G carriers (P = 0.01). To directly investigate the correlation between UCP2 expression and insulin resistance, UCP2 mRNA levels were measured by real-time RT-PCR in subcutaneous fat obtained from 100 obese subjects who underwent laparoscopic adjustable gastric banding. UCP2 mRNA expression was significantly correlated with insulin resistance as assessed by the homeostasis model assessment index (r = 0.27, P = 0.007). We examined the association of the -866A/A genotype in a case-control study including 483 type 2 diabetic subjects and 565 control subjects. The -866A/A genotype was associated with diabetes in women (odds ratio 1.84, 95% CI 1.03-3.28; P = 0.037), but not in men. These results indicate that the -866A/A genotype of the UCP2 gene may contribute to diabetes susceptibility by affecting insulin sensitivity.
Assuntos
Diabetes Mellitus Tipo 2/fisiopatologia , Resistência à Insulina/genética , Proteínas de Membrana Transportadoras/genética , Proteínas Mitocondriais/genética , Regiões Promotoras Genéticas , Tecido Adiposo/metabolismo , Adulto , Alanina , Estudos de Casos e Controles , Diabetes Mellitus Tipo 2/genética , Feminino , Genótipo , Técnica Clamp de Glucose , Glicina , Heterozigoto , Homeostase , Humanos , Canais Iônicos , Masculino , Pessoa de Meia-Idade , Modelos Biológicos , Obesidade/genética , Obesidade/fisiopatologia , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Caracteres Sexuais , Tela Subcutânea/metabolismo , Proteína Desacopladora 2RESUMO
Recent evidence suggests that insulin signaling through the insulin receptor A type (Ex11-), regulates insulin gene transcription. Because chronic hyperglycemia negatively affects insulin receptor function and regulates alternative splicing of the insulin receptor, we inquired whether chronic exposure of pancreatic beta-cells to high glucose results in alterations in insulin signaling due to changes in insulin receptor expression and relative abundance of its spliced isoforms. Our results demonstrate that the insulin receptor is localized in insulin secretory vescicles in human pancreatic beta-cells. Furthermore, we find that alterations in insulin expression and secretion caused by chronic exposure to high glucose are paralleled by decreased insulin receptor expression and increased relative abundance of the Ex11+ isoform in both human islets and RIN beta-cells. PDX-1 and HMGI(Y) transcription factors are down-regulated by high glucose. These changes are associated with defects in insulin signaling involving insulin receptor-associated PI 3-kinase/Akt/PHAS-I pathway in RIN beta-cells. Re-expression in RIN beta-cells chronically exposed to high glucose of the Ex11-, but not the Ex11+, isoform restored insulin mRNA expression. These data suggest that changes in early steps of insulin receptor signaling may play a role in determining beta-cell dysfunction caused by chronic hyperglycemia.
Assuntos
Glucose/farmacologia , Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Proteínas Serina-Treonina Quinases , Receptor de Insulina/genética , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal , Proteínas de Transporte/metabolismo , Proteínas de Ciclo Celular , Linhagem Celular , Proteína HMGA1a/metabolismo , Humanos , Insulina/biossíntese , Proteínas Substratos do Receptor de Insulina , Secreção de Insulina , Peptídeos e Proteínas de Sinalização Intracelular , Ilhotas Pancreáticas/efeitos dos fármacos , Modelos Biológicos , Pâncreas/química , Pâncreas/ultraestrutura , Fosfatidilinositol 3-Quinases/metabolismo , Fosfoproteínas/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-akt , Splicing de RNA , Receptor de Insulina/análise , Receptor de Insulina/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Transcrição GênicaRESUMO
Weight loss in metabolically healthy obese (MHO) subjects may result in deterioration of cardio-metabolic risk profile. We analyzed the effects of weight loss induced by laparoscopic adjustable gastric banding (LAGB) on cardio-metabolic risk factors in MHO and insulin resistant obese (IRO) individuals. This study included 190 morbidly obese non-diabetic subjects. Obese individuals were stratified on the basis of their insulin sensitivity index (ISI), estimated from an OGTT, into MHO (ISI index in the upper quartile) and IRO (ISI in the three lower quartiles). Anthropometric and cardio-metabolic variables were measured at baseline and 6-months after LAGB. Six months after LAGB, anthropometric measures were significantly reduced in both MHO and IRO. Percent changes in body weight, BMI, and waist circumference did not differ between the two groups. Fasting glucose and insulin levels, triglycerides, AST, and ALT were significantly reduced, and HDL cholesterol significantly increased, in both MHO and IRO subjects with no differences in percent changes from baseline. Insulin sensitivity increased in both MHO and IRO group. Insulin secretion was significantly reduced in the IRO group only. However, the disposition index significantly increased in both MHO and IRO individuals with no differences in percent changes from baseline between the two groups. The change in insulin sensitivity correlated with the change in BMI (râ=â-0.43; P<0.0001). In conclusion, our findings reinforce the recommendation that weight loss in response to LAGB intervention should be considered an appropriate treatment option for morbidly obese individuals regardless of their metabolic status, i.e. MHO vs. IRO subjects.
Assuntos
Dieta Redutora , Gastroplastia , Saúde , Laparoscopia , Obesidade/fisiopatologia , Obesidade/cirurgia , Redução de Peso/fisiologia , Adulto , Índice de Massa Corporal , Feminino , Seguimentos , Humanos , Resistência à Insulina/fisiologia , Masculino , Obesidade/metabolismoRESUMO
Insulin resistance, reduced ß-cell mass, and hyperglucagonemia are consistent features in type 2 diabetes mellitus (T2DM). We used pancreas and islets from humans with T2DM to examine the regulation of insulin signaling and cell-cycle control of islet cells. We observed reduced ß-cell mass and increased α-cell mass in the Type 2 diabetic pancreas. Confocal microscopy, real-time PCR and western blotting analyses revealed increased expression of PCNA and down-regulation of p27-Kip1 and altered expression of insulin receptors, insulin receptor substrate-2 and phosphorylated BAD. To investigate the mechanisms underlying these findings, we examined a mouse model of insulin resistance in ß-cells--which also exhibits reduced ß-cell mass, the ß-cell-specific insulin receptor knockout (ßIRKO). Freshly isolated islets and ß-cell lines derived from ßIRKO mice exhibited poor cell-cycle progression, nuclear restriction of FoxO1 and reduced expression of cell-cycle proteins favoring growth arrest. Re-expression of insulin receptors in ßIRKO ß-cells reversed the defects and promoted cell cycle progression and proliferation implying a role for insulin-signaling in ß-cell growth. These data provide evidence that human ß- and α-cells can enter the cell-cycle, but proliferation of ß-cells in T2DM fails due to G1-to-S phase arrest secondary to defective insulin signaling. Activation of insulin signaling, FoxO1 and proteins in ß-cell-cycle progression are attractive therapeutic targets to enhance ß-cell regeneration in the treatment of T2DM.
Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/patologia , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Receptor de Insulina/metabolismo , Transdução de Sinais , Idoso , Idoso de 80 Anos ou mais , Animais , Moléculas de Adesão Celular/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Feminino , Fase G1/genética , Regulação da Expressão Gênica , Humanos , Insulina/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Modelos Biológicos , Antígeno Nuclear de Célula em Proliferação/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor de Insulina/deficiência , Fase S/genética , Transdução de Sinais/genética , Doadores de TecidosRESUMO
OBJECTIVE: To evaluate oxidative stress in uremia and dialysis and chromogranin A, a stress hormone that could be related to oxidative processes. METHODS: Plasma oxidative stress biomarkers (-SH, 8-OHdG, and ox-LDL) and chromogranin A were measured in 89 outpatients (21 uremic patients, 17 in peritoneal dialysis, and 51 in haemodialysis), and in 18 subjects with normal renal function. RESULTS: -SH groups were significantly reduced in heamodialysis, peritoneal, and uremic patients as compared with the control group (p=0.01), while 8-OHdG was increased (p<0.01). No differences were observed for ox-LDL. Chromogranin A was increased in uremic, peritoneal and haemodialysis patients (p<0.01), showing a positive correlation to 8-OHdG (p<0.01). CONCLUSION: Oxidative stress biomarkers and chromogranin A levels differ between control subjects when compared to both uremic and dialysis patients. No differences were observed between uremic and dialysis patients, suggesting that uremia is the major source of the increase in oxidative stress and CgA levels in patients with end stage renal disease.
Assuntos
Biomarcadores/sangue , Cromogranina A/sangue , Estresse Oxidativo , Diálise Peritoneal , Diálise Renal , Uremia/sangue , Idoso , Idoso de 80 Anos ou mais , Animais , Humanos , Masculino , Pessoa de Meia-Idade , Uremia/fisiopatologiaRESUMO
BACKGROUND: In type 1 diabetes (T1D) vascular complications such as accelerated atherosclerosis and diffused macro-/microangiopathy are linked to chronic hyperglycemia with a mechanism that is not yet well understood. End-stage renal disease (ESRD) worsens most diabetic complications, particularly, the risk of morbidity and mortality from cardiovascular disease is increased several fold. METHODS AND FINDINGS: We evaluated protein regulation and expression in skin biopsies obtained from T1D patients with and without ESRD, to identify pathways of persistent cellular changes linked to diabetic vascular disease. We therefore examined pathways that may be normalized by restoration of normoglycemia with kidney-pancreas (KP) transplantation. Using proteomic and ultrastructural approaches, multiple alterations in the expression of proteins involved in oxidative stress (catalase, superoxide dismutase 1, Hsp27, Hsp60, ATP synthase delta chain, and flavin reductase), aerobic and anaerobic glycolysis (ACBP, pyruvate kinase muscle isozyme, and phosphoglycerate kinase 1), and intracellular signaling (stratifin-14-3-3, S100-calcyclin, cathepsin, and PPI rotamase) as well as endothelial vascular abnormalities were identified in T1D and T1D+ESRD patients. These abnormalities were reversed after KP transplant. Increased plasma levels of malondialdehyde were observed in T1D and T1D+ESRD patients, confirming increased oxidative stress which was normalized after KP transplant. CONCLUSIONS: Our data suggests persistent cellular changes of anti-oxidative machinery and of aerobic/anaerobic glycolysis are present in T1D and T1D+ESRD patients, and these abnormalities may play a key role in the pathogenesis of hyperglycemia-related vascular complications. Restoration of normoglycemia and removal of uremia with KP transplant can correct these abnormalities. Some of these identified pathways may become potential therapeutic targets for a new generation of drugs.
Assuntos
Diabetes Mellitus Tipo 1/metabolismo , Regulação da Expressão Gênica , Transplante de Rim/métodos , Oxigênio/química , Transplante de Pâncreas/métodos , Proteômica/métodos , Pele/metabolismo , Adulto , Estudos de Casos e Controles , Diabetes Mellitus Tipo 1/terapia , Feminino , Glicólise , Humanos , Falência Renal Crônica/metabolismo , Masculino , Pessoa de Meia-Idade , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
Insulin and angiotensin II are hormones that play pivotal roles in the control of two vital and closely related systems, the metabolic and the circulatory systems, respectively. A failure in the proper action of each of these hormones results, to a variable degree, in the development of two highly prevalent and commonly overlapping diseases-diabetes mellitus and hypertension. In recent years, a series of studies has revealed a tight connection between the signal transduction pathways that mediate insulin and angiotensin II actions in target tissues. This molecular cross-talk occurs at multiple levels and plays an important role in phenomena that range from the action of anti-hypertensive drugs to cardiac hypertrophy and energy acquisition by the heart. At the extracellular level, the angiotensin-converting enzyme controls angiotensin II synthesis but also interferes with insulin signaling through the proper regulation of angiotensin II and through the accumulation of bradykinin. At an early intracellular level, angiotensin II, acting through JAK-2/IRS-1/PI3-kinase, JNK and ERK, may induce the serine phosphorylation and inhibition of key elements of the insulin-signaling pathway. Finally, by inducing the expression of the regulatory protein SOCS-3, angiotensin II may impose a late control on the insulin signal. This review will focus on the main advances obtained in this field and will discuss the implications of this molecular cross-talk in the common clinical association between diabetes mellitus and hypertension.
Assuntos
Angiotensina II/fisiologia , Insulina/fisiologia , Transdução de Sinais/fisiologia , Diabetes Mellitus/fisiopatologia , Angiopatias Diabéticas/fisiopatologia , Humanos , Hipertensão/fisiopatologia , Modelos Biológicos , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina/fisiologiaRESUMO
End-stage renal disease (ESRD) is characterized by several atherothrombotic abnormalities, and kidney transplant seems to improve most of them. However, because it is not clear which mechanism is responsible for such improvement, our purpose was to clarify that point.We conducted a cross sectional study involving 30 ESRD patients, 30 ESRD kidney-transplanted patients (Ktx) and 30 healthy controls (C) to evaluate platelet morphology and function, atherothrombotic profile, endothelial abnormalities and cytokine levels involved in the insulin resistance/endothelial dysfunction. (i) Platelet morphology: The ESRD group showed platelet size similar to the other two groups (ESRD=3518x10(3)+/-549x10(3) nm2, C=3075x10(3)+/-197x10(3) nm2, Ktx=2862x10(3)+/-205x10(3) nm2) with similar platelet granules and number. (ii) Platelet surface glycoprotein: The CD41 and P-Selectin were similar between groups. (iii) Platelet intracellular calcium: Resting intracellular calcium was statistically higher in ESRD compared to the C group (ESRD=182.1+/-34.5, Ktx=126.7+/-14.1, C=72.0+/-11.0 nM, P<0.01). (iv) Hypercoagulability markers and natural anticoagulants: The Ktx and ESRD groups showed higher levels of hypercoagulability markers compared to the C group. A reduction in antithrombin activity was evident in ESRD compared to the Ktx group (P=0.03). (v) Endothelial morphology: The ESRD group showed a thickened vessel basal membrane compared to the Ktx and C groups with more endothelial sufference. (vi) Insulin resistance and pro-inflammatory cytokine profile: The ESRD showed a higher homeostasis model assessment provided equations for estimating insulin resistance (HOMA-IR) compared to the Ktx and C groups (ESRD=2.6+/-0.3, Ktx=1.8+/-0.2, C=1.1+/-0.1, P=0.005) and increased soluble tumor neurosis factor alpha (sTNFalpha) (P<0.05) and soluble vascular cell adhesion molecule (sVCAM) levels (P<0.01). Positive correlations were evident among HOMA-IR and sTNFalpha (P<0.001) and sVCAM (P=0.01), respectively. In a small subgroup of ESRD who underwent Ktx (five pts), our findings were confirmed at 1 year of follow-up, suggesting an improvement of almost haemostatic abnormalities. Kidney transplant is associated with a better atherothrombotic profile in ESRD, platelet intracellular calcium and cytokines seem to be most influenced by the transplant, while most morphological abnormalities are retained.