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BACKGROUND: The reduction in cardiovascular disease (CVD) events with edetate disodium (EDTA) in the Trial to Assess Chelation Therapy (TACT) suggested that chelation of toxic metals might provide novel opportunities to reduce CVD in patients with diabetes. Lead and cadmium are vasculotoxic metals chelated by EDTA. We present baseline characteristics for participants in TACT2, a randomized, double-masked, placebo-controlled trial designed as a replication of the TACT trial limited to patients with diabetes. METHODS: TACT2 enrolled 1,000 participants with diabetes and prior myocardial infarction, age 50 years or older between September 2016 and December 2020. Among 959 participants with at least one infusion, 933 had blood and/or urine metals measured at the Centers for Diseases Control and Prevention using the same methodology as in the National Health and Nutrition Examination Survey (NHANES). We compared metal levels in TACT2 to a contemporaneous subset of NHANES participants with CVD, diabetes and other inclusion criteria similar to TACT2's participants. RESULTS: At baseline, the median (interquartile range, IQR) age was 67 (60, 72) years, 27% were women, 78% reported white race, mean (SD) BMI was 32.7 (6.6) kg/m2, 4% reported type 1 diabetes, 46.8% were treated with insulin, 22.3% with GLP1-receptor agonists or SGLT-2 inhibitors, 90.2% with aspirin, warfarin or P2Y12 inhibitors, and 86.5% with statins. Blood lead was detectable in all participants; median (IQR) was 9.19 (6.30, 13.9) µg/L. Blood and urine cadmium were detectable in 97% and median (IQR) levels were 0.28 (0.18, 0.43) µg/L and 0.30 (0.18, 0.51) µg/g creatinine, respectively. Metal levels were largely similar to those in the contemporaneous NHANES subset. CONCLUSIONS: TACT2 participants were characterized by high use of medication to treat CVD and diabetes and similar baseline metal levels as in the general US population. TACT2 will determine whether chelation therapy reduces the occurrence of subsequent CVD events in this high-risk population. CLINICAL TRIALS REGISTRATION: ClinicalTrials.gov. Identifier: NCT02733185. https://clinicaltrials.gov/study/NCT02733185.
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Terapia por Quelación , Humanos , Femenino , Masculino , Persona de Mediana Edad , Anciano , Terapia por Quelación/métodos , Método Doble Ciego , Ácido Edético/uso terapéutico , Plomo/sangre , Plomo/orina , Cadmio/orina , Cadmio/sangre , Quelantes/uso terapéutico , Enfermedades Cardiovasculares/prevención & control , Enfermedades Cardiovasculares/sangreRESUMEN
Importance: In 2013, the Trial to Assess Chelation Therapy (TACT) reported that edetate disodium (EDTA)-based chelation significantly reduced cardiovascular disease (CVD) events by 18% in 1708 patients with a prior myocardial infarction (MI). Objective: To replicate the finding of TACT in individuals with diabetes and previous MI. Design, Setting, and Participants: A 2 × 2 factorial, double-masked, placebo-controlled, multicenter trial at 88 sites in the US and Canada, involving participants who were 50 years or older, had diabetes, and had experienced an MI at least 6 weeks before recruitment compared the effect of EDTA-based chelation vs placebo infusions on CVD events and compared the effect of high doses of oral multivitamins and minerals with oral placebo. This article reports on the chelation vs placebo infusion comparisons. Interventions: Eligible participants were randomly assigned to 40 weekly infusions of an EDTA-based chelation solution or matching placebo and to twice daily oral, high-dose multivitamin and mineral supplements or matching placebo for 60 months. This article addresses the chelation study. Main Outcomes and Measures: The primary end point was the composite of all-cause mortality, MI, stroke, coronary revascularization, or hospitalization for unstable angina. Median follow-up was 48 months. Primary comparisons were made from patients who received at least 1 assigned infusion. Results: Of the 959 participants (median age, 67 years [IQR, 60-72 years]; 27% females; 78% White, 10% Black, and 20% Hispanic), 483 received at least 1 chelation infusion and 476 at least 1 placebo infusion. A primary end point event occurred in 172 participants (35.6%) in the chelation group and in 170 (35.7%) in the placebo group (adjusted hazard ratio [HR], 0.93; 95% CI, 0.76-1.16; P = .53). The 5-year primary event cumulative incidence rates were 45.8% for the chelation group and 46.5% for the placebo group. CV death, MI, or stroke events occurred in 89 participants (18.4%) in the chelation group and in 94 (19.7%) in the placebo group (adjusted HR, 0.89; 95% CI, 0.66-1.19). Death from any cause occurred in 84 participants (17.4%) in the chelation group and in 84 (17.6%) in the placebo group (adjusted HR, 0.96; 95% CI, 0.71-1.30). Chelation reduced median blood lead levels from 9.03 µg/L at baseline to 3.46 µg/L at infusion 40 (P < .001). Corresponding levels in the placebo group were 9.3 µg/L and 8.7 µg/L, respectively. Conclusions and Relevance: Despite effectively reducing blood lead levels, EDTA chelation was not effective in reducing cardiovascular events in stable patients with coronary artery disease who have diabetes and a history of MI. Trial Registration: ClinicalTrials.gov Identifier: NCT02733185.
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BACKGROUND: Intravenous edetate disodium-based infusions reduced cardiovascular events in a prior clinical trial. The Trial to Assess Chelation Therapy 2 (TACT2) will replicate the initial study design. METHODS: TACT2 is an NIH-sponsored, randomized, 2x2 factorial, double masked, placebo-controlled, multicenter clinical trial testing 40 weekly infusions of a multi-component edetate disodium (disodium ethylenediamine tetra-acetic acid, or Na2EDTA)-based chelation solution and twice daily oral, high-dose multivitamin and mineral supplements in patients with diabetes and a prior myocardial infarction (MI). TACT2 completed enrollment of 1000 subjects in December 2020, and infusions in December 2021. Subjects are followed for 2.5 to 5 years. The primary endpoint is time to first occurrence of all-cause mortality, MI, stroke, coronary revascularization, or hospitalization for unstable angina. The trial has >;85% power to detect a 30% relative reduction in the primary endpoint. TACT2 also includes a Trace Metals and Biorepository Core Lab, to test whether benefits of treatment, if present, are due to chelation of lead and cadmium from patients. Design features of TACT2 were chosen to replicate selected features of the first TACT, which demonstrated a significant reduction in cardiovascular outcomes in the EDTA chelation arm compared with placebo among patients with a prior MI, with the largest effect in patients with diabetes. RESULTS: Results are expected in 2024. CONCLUSION: TACT2 may provide definitive evidence of the benefit of edetate disodiumbased chelation on cardiovascular outcomes, as well as the clinical importance of longitudinal changes in toxic metal levels of participants.
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Diabetes Mellitus , Infarto del Miocardio , Quelantes/uso terapéutico , Terapia por Quelación/métodos , Diabetes Mellitus/tratamiento farmacológico , Método Doble Ciego , Ácido Edético/uso terapéutico , Humanos , Infarto del Miocardio/tratamiento farmacológico , Infarto del Miocardio/epidemiología , VitaminasRESUMEN
The National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) Diabetic Foot Consortium (DFC) was established in September 2018 by the NIDDK to build an organization to facilitate the highest quality of clinical research on diabetic foot ulcers (DFUs) that will answer clinically significant questions to improve DFU healing and prevent amputations. The initial focus of the DFC is to develop and validate biomarkers for DFUs that can be used in clinical care and research. The DFC consists of a data coordinating center (DCC) for operational oversight and statistical analysis, clinical sites for participant recruitment and evaluation, and biomarker analysis units (BAUs). The DFC is currently studying biomarkers to predict wound healing and recurrence and is collecting biosamples for future studies through a biorepository. The DFC plans to address the challenges of recruitment and eligibility criteria for DFU clinical trials by taking an approach of "No DFU Patient Goes Unstudied." In this platform approach, clinical history, DFU outcome, wound imaging, and biologic measurements from a large number of patients will be captured and the in-depth longitudinal data set will be analyzed to develop a computational-based DFU risk factor profile to facilitate scientifically sound clinical trial design. The DFC will expand its platform to include studies of the role of social determinants of health, such as food insecurity, housing instability, limited health literacy, and poor social support. The DFC is starting partnerships with the broad group of stakeholders in the wound care community.
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Diabetes Mellitus , Pie Diabético , Nefropatías Diabéticas , Estados Unidos , Humanos , Pie Diabético/terapia , National Institute of Diabetes and Digestive and Kidney Diseases (U.S.) , Factores de Riesgo , BiomarcadoresRESUMEN
BACKGROUND: Optimal treatment for patients with both type 2 diabetes mellitus and stable ischemic heart disease has not been established. METHODS: We randomly assigned 2368 patients with both type 2 diabetes and heart disease to undergo either prompt revascularization with intensive medical therapy or intensive medical therapy alone and to undergo either insulin-sensitization or insulin-provision therapy. Primary end points were the rate of death and a composite of death, myocardial infarction, or stroke (major cardiovascular events). Randomization was stratified according to the choice of percutaneous coronary intervention (PCI) or coronary-artery bypass grafting (CABG) as the more appropriate intervention. RESULTS: At 5 years, rates of survival did not differ significantly between the revascularization group (88.3%) and the medical-therapy group (87.8%, P=0.97) or between the insulin-sensitization group (88.2%) and the insulin-provision group (87.9%, P=0.89). The rates of freedom from major cardiovascular events also did not differ significantly among the groups: 77.2% in the revascularization group and 75.9% in the medical-treatment group (P=0.70) and 77.7% in the insulin-sensitization group and 75.4% in the insulin-provision group (P=0.13). In the PCI stratum, there was no significant difference in primary end points between the revascularization group and the medical-therapy group. In the CABG stratum, the rate of major cardiovascular events was significantly lower in the revascularization group (22.4%) than in the medical-therapy group (30.5%, P=0.01; P=0.002 for interaction between stratum and study group). Adverse events and serious adverse events were generally similar among the groups, although severe hypoglycemia was more frequent in the insulin-provision group (9.2%) than in the insulin-sensitization group (5.9%, P=0.003). CONCLUSIONS: Overall, there was no significant difference in the rates of death and major cardiovascular events between patients undergoing prompt revascularization and those undergoing medical therapy or between strategies of insulin sensitization and insulin provision. (ClinicalTrials.gov number, NCT00006305.)
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Angioplastia Coronaria con Balón , Puente de Arteria Coronaria , Enfermedad Coronaria/terapia , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Hipoglucemiantes/uso terapéutico , Insulina/uso terapéutico , Terapia Combinada , Angiografía Coronaria , Enfermedad Coronaria/complicaciones , Enfermedad Coronaria/cirugía , Diabetes Mellitus Tipo 2/complicaciones , Femenino , Hemoglobina Glucada , Humanos , Hipoglucemiantes/efectos adversos , Insulina/efectos adversos , Estimación de Kaplan-Meier , Masculino , Metformina/uso terapéutico , Persona de Mediana Edad , Infarto del Miocardio/epidemiología , Infarto del Miocardio/prevención & control , Accidente Cerebrovascular/epidemiología , Accidente Cerebrovascular/prevención & control , Compuestos de Sulfonilurea/uso terapéutico , Tiazolidinedionas/uso terapéuticoRESUMEN
G alpha 13 stimulates the guanine nucleotide exchange factors (GEFs) for Rho, such as p115Rho-GEF. Activated Rho induces numerous cellular responses, including actin polymerization, serum response element (SRE)-dependent gene transcription and transformation. p115Rho-GEF contains a Regulator of G protein Signalling domain (RGS box) that confers GTPase activating protein (GAP) activity towards G alpha 12 and G alpha 13 (ref. 3). In contrast, classical RGS proteins (such as RGS16 and RGS4) exhibit RGS domain-dependent GAP activity on G alpha i and G alpha q, but not G alpha 12 or G alpha 13 (ref 4). Here, we show that RGS16 inhibits G alpha 13-mediated, RhoA-dependent reversal of stellation and SRE activation. The RGS16 amino terminus binds G alpha 13 directly, resulting in translocation of G alpha 13 to detergent-resistant membranes (DRMs) and reduced p115Rho-GEF binding. RGS4 does not bind G alpha 13 or attenuate G alpha 13-dependent responses, and neither RGS16 nor RGS4 affects G alpha 12-mediated signalling. These results elucidate a new mechanism whereby a classical RGS protein regulates G alpha 13-mediated signal transduction independently of the RGS box.
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Subunidades alfa de la Proteína de Unión al GTP G12-G13/metabolismo , Proteínas/metabolismo , Proteínas RGS/metabolismo , Transducción de Señal/fisiología , Proteínas de Unión al GTP rho/metabolismo , Línea Celular Tumoral , Retroalimentación Fisiológica/genética , Regulación de la Expresión Génica/genética , Genes Reguladores/genética , Factores de Intercambio de Guanina Nucleótido/metabolismo , Humanos , Unión Proteica/genética , Estructura Terciaria de Proteína/genética , Factores de Intercambio de Guanina Nucleótido RhoRESUMEN
The arrest of meiotic prophase in mouse oocytes within antral follicles requires the G protein G(s) and an orphan member of the G protein-coupled receptor family, GPR3. To determine whether GPR3 activates G(s), the localization of Galpha(s) in follicle-enclosed oocytes from Gpr3(+/+) and Gpr3(-/-) mice was compared by using immunofluorescence and Galpha(s)GFP. GPR3 decreased the ratio of Galpha(s) in the oocyte plasma membrane versus the cytoplasm and also decreased the amount of Galpha(s) in the oocyte. Both of these properties indicate that GPR3 activates G(s). The follicle cells around the oocyte are also necessary to keep the oocyte in prophase, suggesting that they might activate GPR3. However, GPR3-dependent G(s) activity was similar in follicle-enclosed and follicle-free oocytes. Thus, the maintenance of prophase arrest depends on the constitutive activity of GPR3 in the oocyte, and the follicle cell signal acts by a means other than increasing GPR3 activity.
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Subunidades alfa de la Proteína de Unión al GTP Gs/metabolismo , Meiosis/fisiología , Oocitos/metabolismo , Profase/fisiología , Receptores Acoplados a Proteínas G/metabolismo , Animales , Células Cultivadas , Femenino , Subunidades alfa de la Proteína de Unión al GTP Gs/química , Proteínas Fluorescentes Verdes/química , Proteínas Fluorescentes Verdes/metabolismo , Inmunohistoquímica , Ratones , Ratones Noqueados , Oocitos/citología , Folículo Ovárico/citología , Folículo Ovárico/fisiologíaRESUMEN
We evaluated the associations between glycemic therapies and prevalence of diabetic peripheral neuropathy (DPN) at baseline among participants in the Bypass Angioplasty Revascularization Investigation 2 Diabetes (BARI 2D) trial on medical and revascularization therapies for coronary artery disease (CAD) and on insulin-sensitizing vs. insulin-providing treatments for diabetes. A total of 2,368 patients with type 2 diabetes and CAD was evaluated. DPN was defined as clinical examination score >2 using the Michigan Neuropathy Screening Instrument (MNSI). DPN odds ratios across different groups of glycemic therapy were evaluated by multiple logistic regression adjusted for multiple covariates including age, sex, hemoglobin A1c (HbA1c), and diabetes duration. Fifty-one percent of BARI 2D subjects with valid baseline characteristics and MNSI scores had DPN. After adjusting for all variables, use of insulin was significantly associated with DPN (OR = 1.57, 95% CI: 1.15-2.13). Patients on sulfonylurea (SU) or combination of SU/metformin (Met)/thiazolidinediones (TZD) had marginally higher rates of DPN than the Met/TZD group. This cross-sectional study in a cohort of patients with type 2 diabetes and CAD showed association of insulin use with higher DPN prevalence, independent of disease duration, glycemic control, and other characteristics. The causality between a glycemic control strategy and DPN cannot be evaluated in this cross-sectional study, but continued assessment of DPN and randomized therapies in BARI 2D trial may provide further explanations on the development of DPN.
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Neuropatías Diabéticas/tratamiento farmacológico , Neuropatías Diabéticas/epidemiología , Hipoglucemiantes/uso terapéutico , Adulto , Anciano , Estudios de Cohortes , Puente de Arteria Coronaria/métodos , Enfermedad de la Arteria Coronaria/complicaciones , Enfermedad de la Arteria Coronaria/terapia , Estudios Transversales , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Neuropatías Diabéticas/etiología , Femenino , Hemoglobina Glucada/metabolismo , Humanos , Insulina/uso terapéutico , Modelos Logísticos , Masculino , Metformina/uso terapéutico , Persona de Mediana Edad , Examen Neurológico , Prevalencia , Ensayos Clínicos Controlados Aleatorios como Asunto , Compuestos de Sulfonilurea/uso terapéutico , Tiazolidinedionas/uso terapéuticoRESUMEN
One in five Americans experiences disability that affects their daily function because of impairments in mobility, cognitive function, sensory impairment, or communication impairment. The need for rehabilitation strategies to optimize function and reduce disability is a clear priority for research to address this public health challenge. The National Institutes of Health (NIH) recently published a Research Plan on Rehabilitation that provides a set of priorities to guide the field over the next 5 years. The plan was developed with input from multiple Institutes and Centers within the NIH, the National Advisory Board for Medical Rehabilitation Research, and the public. This article provides an overview of the need for this research plan, an outline of its development, and a listing of six priority areas for research. The NIH is committed to working with all stakeholder communities engaged in rehabilitation research to track progress made on these priorities and to work to advance the science of medical rehabilitation.This article is being published almost simultaneously in the following six journals: American Journal of Occupational Therapy, American Journal of Physical Medicine and Rehabilitation, Archives of Physical Medicine and Rehabilitation, Neurorehabilitation and Neural Repair, Physical Therapy, and Rehabilitation Psychology. Citation information is as follows: NIH Medical Rehabilitation Coordinating Committee. Am J Phys Med Rehabil. 2017;97(4):404-407.
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Personas con Discapacidad/rehabilitación , Prioridades en Salud , National Institutes of Health (U.S.) , Investigación en Rehabilitación , Humanos , Objetivos Organizacionales , Estados UnidosRESUMEN
The acylation of proteins through the addition of palmitate to cysteine residues is a common posttranslational modification for a variety of proteins, but the enzymology of this reversible modification has resisted elucidation. We developed a strategy to purify protein fatty acyltransferase (PAT) activity from rat livers that took advantage of recent knowledge on the cellular location and inhibition of PAT activity. We determined that three different thiolases have PAT activity in the presence of imidazole and therefore started the purification with a plasma membrane fraction to minimize the contamination with these enzymes. After detergent extraction of the plasma membrane fraction, the PAT activity was enriched about 90-fold by sequential chromatography including affinity chromatography to a cerulenin-based inhibitor of palmitoylation. The partially purified PAT activity (1) was lost with treatments to degrade or denature proteins, (2) could acylate tubulin, Galpha(i) and RGS16 and (3) showed a preference for palmitate and to a lesser degree other long-chain fatty acids. This purification procedure is a significant advance over previous efforts at PAT purification and a starting point for a proteomic approach for identification of mammalian PAT.
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Aciltransferasas/aislamiento & purificación , Aciltransferasas/metabolismo , Hígado/enzimología , Acilación , Animales , Membrana Celular/enzimología , Cromatografía en Gel/métodos , Electroforesis en Gel de Poliacrilamida/métodos , Cinética , Masculino , Peso Molecular , Procesamiento Proteico-Postraduccional , Ratas , Especificidad por SustratoRESUMEN
Palmitoylation, the reversible, post-translational addition of palmitate to cysteine residues, occurs on several regulators of G-protein signaling (RGS) proteins. Palmitoylation can occur near the amino terminus, as for RGS4 and RGS16, but can also occur on a cysteine residue in the alpha4 helix of the RGS box, which is conserved in most RGS proteins. For some of the RGS proteins, palmitoylation is required to turn off G-protein signaling by accelerating GTP hydrolysis on the Galpha subunit. This article discusses the role of palmitoylation in RGS function and protocols are given for metabolic and in vitro labeling of RGS proteins with [3H]palmitate and measurement of GTP hydrolysis in membranes.
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Ácido Palmítico/metabolismo , Proteínas RGS/metabolismo , Secuencia de Aminoácidos , Animales , Membrana Celular/química , Membrana Celular/enzimología , Membrana Celular/metabolismo , Secuencia Conservada , Cisteína/metabolismo , GTP Fosfohidrolasas/metabolismo , Subunidades alfa de la Proteína de Unión al GTP Gi-Go/metabolismo , Proteínas Activadoras de GTPasa/metabolismo , Guanosina Trifosfato/metabolismo , Humanos , Modelos Moleculares , Procesamiento Proteico-Postraduccional , Estructura Secundaria de Proteína , Proteínas RGS/química , Proteínas RGS/genética , Transducción de SeñalRESUMEN
OBJECTIVE: The Bypass Angioplasty Revascularization Investigation 2 Diabetes (BARI 2D) trial demonstrated similar long-term clinical effectiveness of insulin-sensitizing (IS) versus insulin-providing (IP) treatments for type 2 diabetes on cardiovascular outcomes in a cohort with documented coronary artery disease. We evaluated the effects of randomized glycemic control strategy (IS vs. IP) on the prevalence and incidence of diabetic peripheral neuropathy (DPN). RESEARCH DESIGN AND METHODS: DPN (defined as Michigan Neuropathy Screening Instrument [MNSI] clinical examination score>2) was assessed at baseline and yearly for 4 years. DPN prevalence and incidence were compared by intention-to-treat modeling by logistic generalized estimating equation models for prevalence and Kaplan-Meier estimates and Cox regression models for incidence rates. RESULTS: Results are reported for 2,159 BARI 2D participants (70% males) with valid baseline and at least one follow-up MNSI score (mean age 62±9 years, mean HbA1c 7.7±1.6%, diabetes duration 10±9 years). There were no differences in the prevalence of DPN between the IS and the IP groups throughout the 4 years of follow-up. In 1,075 BARI 2D participants with no DPN at baseline, the 4-year cumulative incidence rate of DPN was significantly lower in the IS (66%) than in the IP (72%) strategy group (P=0.02), which remained significant after adjusting for the in-trial HbA1c (P=0.04). In subgroup analyses, IS strategy had a greater benefit in men (hazard ratio 0.75 [99% CI 0.58-0.99], P<0.01). CONCLUSIONS: Among patients with type 2 diabetes followed for up to 4 years during BARI 2D, a glycemic control therapy with IS significantly reduced the incidence of DPN compared with IP therapy and may add further benefit for men.
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Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/fisiopatología , Neuropatías Diabéticas/etiología , Neuropatías Diabéticas/fisiopatología , Enfermedades del Sistema Nervioso Periférico/etiología , Enfermedades del Sistema Nervioso Periférico/fisiopatología , Anciano , Glucemia/efectos de los fármacos , Glucemia/metabolismo , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Neuropatías Diabéticas/sangre , Neuropatías Diabéticas/tratamiento farmacológico , Femenino , Humanos , Hipoglucemiantes/uso terapéutico , Masculino , Persona de Mediana Edad , Enfermedades del Sistema Nervioso Periférico/sangre , Enfermedades del Sistema Nervioso Periférico/tratamiento farmacológicoRESUMEN
The relation between the metabolic syndrome (MetS) and resting heart rate (rHR) in patients with diabetes and coronary artery disease is unknown. The authors examined the cross-sectional association at baseline between components of the MetS and rHR and between rHR and left ventricular ejection fraction in the population from the Bypass Angioplasty Revascularization Investigation 2 Diabetes (BARI 2D) randomized clinical trial. The mean rHR in the MetS group was significantly higher than in those without (68.4+/-12.3 vs 65.6+/-11.8 beats per min, P=.0017). The rHR was higher (P<.001 for trend) with increasing number of components for MetS. Linear regression analyses demonstrated that as compared to individuals without MetS, rHR was significantly higher in participants with MetS (regression coefficient, 2.9; P=.0015). In patients with type 2 diabetes and coronary artery disease, the presence of higher rHR is associated with increasing number of criteria of MetS and the presence of ventricular dysfunction.
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Enfermedad de la Arteria Coronaria/fisiopatología , Diabetes Mellitus Tipo 2/fisiopatología , Frecuencia Cardíaca , Síndrome Metabólico/fisiopatología , Descanso , Angiografía Coronaria , Estudios Transversales , Femenino , Indicadores de Salud , Humanos , Modelos Lineales , Masculino , Persona de Mediana Edad , Modelos Estadísticos , Análisis Multivariante , Factores de Riesgo , Estadística como Asunto , Volumen Sistólico , Función Ventricular IzquierdaRESUMEN
Patients with diabetes continue to die of coronary artery disease (CAD) at rates 2 to 4 times higher than patients without diabetes, despite advances in treatment of cardiovascular disease. The role of glycemic control therapies, independent of their glucose-lowering effects, on cardiovascular disease is a recurring question. We examined the association of glycemic control therapies with extent of CAD as measured by coronary angiogram obtained at baseline in 1,803 subjects in the Bypass Angioplasty Revascularization Investigation 2 Diabetes (BARI 2D) trial who had type 2 diabetes mellitus, documented moderate to severe CAD, and no previous cardiac revascularization procedures. The association between glycemic control therapy use recorded at baseline and percent coronary artery stenosis and myocardial jeopardy index was analyzed by multiple regression models. Insulin use at study entry was associated with 23% fewer highly stenotic lesions (> or =70%) (p <0.001) and a significantly lower myocardial jeopardy index compared with subjects not on insulin, despite a worse cardiac risk factor profile, more unstable angina, and increased inflammatory markers in insulin users. Subjects taking thiazolidinediones (TZDs) for > or =6 months had 17% fewer highly stenotic lesions (p = 0.02) and significantly lower C-reactive protein, fibrinogen, and plasminogen activator inhibitor-1 levels compared with those not taking TZDs. In conclusion, this cross-sectional study of patients with type 2 diabetes mellitus and CAD showed that treatment with insulin or TZDs was associated with fewer highly stenotic lesions, independent of disease duration, glycemic control, and other risk factors.
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Enfermedad de la Arteria Coronaria/complicaciones , Vasos Coronarios/efectos de los fármacos , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Hipoglucemiantes/uso terapéutico , Insulina/uso terapéutico , Tiazolidinedionas/uso terapéutico , Angioplastia de Balón , Angiografía Coronaria , Enfermedad de la Arteria Coronaria/tratamiento farmacológico , Enfermedad de la Arteria Coronaria/terapia , Vasos Coronarios/patología , Estudios Transversales , Diabetes Mellitus Tipo 2/complicaciones , Progresión de la Enfermedad , Femenino , Humanos , Hipoglucemiantes/farmacología , Insulina/farmacología , Modelos Logísticos , Masculino , Persona de Mediana Edad , Oportunidad Relativa , Factores de Riesgo , Índice de Severidad de la Enfermedad , Tiazolidinedionas/farmacologíaRESUMEN
The heterotrimeric G proteins, G(12) and G(13), are closely related in their sequences, signaling partners, and cellular effects such as oncogenic transformation and cytoskeletal reorganization. Yet G(12) and G(13) can act through different pathways, bind different proteins, and show opposing actions on some effectors. We investigated the compartmentalization of G(12) and G(13) at the membrane because other G proteins reside in lipid rafts, membrane microdomains enriched in cholesterol and sphingolipids. Lipid rafts were isolated after cold, nonionic detergent extraction of cells and gradient centrifugation. Galpha(12) was in the lipid raft fractions, whereas Galpha(13) was not associated with lipid rafts. Mutation of Cys-11 on Galpha(12), which prevents its palmitoylation, partially shifted Galpha(12) from the lipid rafts. Geldanamycin treatment, which specifically inhibits Hsp90, caused a partial loss of wild-type Galpha(12) and a complete loss of the Cys-11 mutant from the lipid rafts and the appearance of a higher molecular weight form of Galpha(12) in the soluble fractions. These results indicate that acylation and Hsp90 interactions localized Galpha(12) to lipid rafts. Hsp90 may act as both a scaffold and chaperone to maintain a functional Galpha(12) only in discrete membrane domains and thereby explain some of the nonoverlapping functions of G(12) and G(13) and control of these potent cell regulators.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Proteínas HSP90 de Choque Térmico/metabolismo , Proteínas de Unión al GTP Heterotriméricas/metabolismo , Microdominios de Membrana/metabolismo , beta-Ciclodextrinas , Células 3T3 , Acilación , Animales , Benzoquinonas , Células COS , Línea Celular , Ciclodextrinas/farmacología , Cisteína/química , Detergentes/farmacología , Dimerización , Inhibidores Enzimáticos/farmacología , Subunidades alfa de la Proteína de Unión al GTP G12-G13 , Humanos , Immunoblotting , Lactamas Macrocíclicas , Ratones , Octoxinol/farmacología , Unión Proteica , Estructura Terciaria de Proteína , Transporte de Proteínas , Quinonas/farmacología , Transducción de Señal , Fracciones Subcelulares/metabolismo , TransfecciónRESUMEN
Most heterotrimeric G-protein alpha subunits are posttranslationally modified by palmitoylation, a reversible process that is dynamically regulated. We analyzed the effects of Galpha(s) palmitoylation for its intracellular distribution and ability to couple to the beta-adrenergic receptor (betaAR) and stimulate adenylyl cyclase. Subcellular fractionation and immunofluorescence microscopy of stably transfected cyc(-) cells, which lack endogenous Galpha(s), showed that wild-type Galpha(s) was predominantly localized at the plasma membrane, but the mutant C3A-Galpha(s), which does not incorporate [(3)H]palmitate, was mostly associated with intracellular membranes. In agreement with this mislocalization, C3A-Galpha(s) showed neither isoproterenol- or GTPgammaS-stimulated adenylyl cyclase activation nor GTPgammaS-sensitive high-affinity agonist binding, all of which were present in the wild-type Galpha(s) expressing cells. Fusion of C3A-Galpha(s) with the betaAR [betaAR-(C3A)Galpha(s)] partially rescued its ability to induce high-affinity agonist binding and to stimulate adenylyl cyclase activity after isoproterenol or GTPgammaS treatment. In comparison to results with the WT-Galpha(s) and betaAR (betaAR-Galpha(s)) fusion protein, the betaAR-(C3A)Galpha(s) fusion protein was about half as efficient at coupling to the receptor and effector. Chemical depalmitoylation by hydroxylamine of membranes expressing betaAR-Galpha(s) reduced the high-affinity agonist binding and adenylyl cyclase activation to a similar degree as that observed in betaAR-(C3A)Galpha(s) expressing membranes. Altogether, these findings indicate that palmitoylation ensured proper localization of Galpha(s) and facilitated bimolecular interactions of Galpha(s) with the betaAR and adenylyl cyclase.
Asunto(s)
Subunidades alfa de la Proteína de Unión al GTP Gs/química , Mutagénesis Sitio-Dirigida , Ácido Palmítico/metabolismo , Receptores Adrenérgicos beta 2/química , Proteínas Recombinantes de Fusión/química , Adenilil Ciclasas/metabolismo , Alanina/genética , Animales , Membrana Celular/efectos de los fármacos , Membrana Celular/genética , Membrana Celular/metabolismo , Cisteína/genética , Subunidades alfa de la Proteína de Unión al GTP Gs/deficiencia , Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Subunidades alfa de la Proteína de Unión al GTP Gs/metabolismo , Regulación de la Expresión Génica/genética , Humanos , Hidroxilamina/farmacología , Líquido Intracelular/metabolismo , Ratones , Unión Proteica/efectos de los fármacos , Unión Proteica/genética , Subunidades de Proteína/química , Subunidades de Proteína/deficiencia , Subunidades de Proteína/genética , Subunidades de Proteína/metabolismo , Transporte de Proteínas/efectos de los fármacos , Transporte de Proteínas/genética , Ratas , Receptores Adrenérgicos beta 2/biosíntesis , Receptores Adrenérgicos beta 2/genética , Receptores Adrenérgicos beta 2/metabolismo , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Transfección , Células Tumorales CultivadasRESUMEN
The mammalian ovarian follicle consists of a multilayered complex of somatic cells that surround the oocyte. A signal from the follicle cells keeps the oocyte cell cycle arrested at prophase of meiosis I until luteinizing hormone from the pituitary acts on the follicle cells to release the arrest, causing meiosis to continue. Here we show that meiotic arrest can be released in mice by microinjecting the oocyte within the follicle with an antibody that inhibits the stimulatory heterotrimeric GTP-binding protein Gs. This indicates that Gs activity in the oocyte is required to maintain meiotic arrest within the ovarian follicle and suggests that the follicle may keep the cell cycle arrested by activating Gs.