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1.
iScience ; 27(1): 108681, 2024 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-38269100

RESUMO

Aging increases the risk of age-related diseases, imposing substantial healthcare and personal costs. Targeting fundamental aging mechanisms pharmacologically can promote healthy aging and reduce this disease susceptibility. In this work, we employed transcriptome-based drug screening to identify compounds emulating transcriptional signatures of long-lived genetic interventions. We discovered compound 60 (Cmpd60), a selective histone deacetylase 1 and 2 (HDAC1/2) inhibitor, mimicking diverse longevity interventions. In extensive molecular, phenotypic, and bioinformatic assessments using various cell and aged mouse models, we found Cmpd60 treatment to improve age-related phenotypes in multiple organs. Cmpd60 reduces renal epithelial-mesenchymal transition and fibrosis in kidney, diminishes dementia-related gene expression in brain, and enhances cardiac contractility and relaxation for the heart. In sum, our two-week HDAC1/2 inhibitor treatment in aged mice establishes a multi-tissue, healthy aging intervention in mammals, holding promise for therapeutic translation to promote healthy aging in humans.

2.
Placenta ; 145: 19-26, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38011757

RESUMO

INTRODUCTION: Hypertensive disorders of pregnancy (HDP) and fetal growth restriction (FGR) are common obstetrical complications, often with pathological features of maternal vascular malperfusion (MVM) in the placenta. Currently, clinical placental pathology methods involve a manual visual examination of histology sections, a practice that can be resource-intensive and demonstrates moderate-to-poor inter-pathologist agreement on diagnostic outcomes, dependant on the degree of pathologist sub-specialty training. METHODS: This study aims to apply machine learning (ML) feature extraction methods to classify digital images of placental histopathology specimens, collected from cases of HDP [pregnancy induced hypertension (PIH), preeclampsia (PE), PE + FGR], normotensive FGR, and healthy pregnancies, according to the presence or absence of MVM lesions. 159 digital images were captured from histological placental specimens, manually scored for MVM lesions (MVM- or MVM+) and used to develop a support vector machine (SVM) classifier model, using features extracted from pre-trained ResNet18. The model was trained with data augmentation and shuffling, with the performance assessed for patch-level and image-level classification through measurements of accuracy, precision, and recall using confusion matrices. RESULTS: The SVM model demonstrated accuracies of 70 % and 79 % for patch-level and image-level MVM classification, respectively, with poorest performance observed on images with borderline MVM presence, as determined through post hoc observation. DISCUSSION: The results are promising for the integration of ML methods into the placental histopathological examination process. Using this study as a proof-of-concept will lead our group and others to carry ML models further in placental histopathology.


Assuntos
Hipertensão Induzida pela Gravidez , Pré-Eclâmpsia , Gravidez , Feminino , Humanos , Placenta/patologia , Resultado da Gravidez , Estudos Retrospectivos , Pré-Eclâmpsia/patologia , Hipertensão Induzida pela Gravidez/patologia , Retardo do Crescimento Fetal/diagnóstico por imagem , Retardo do Crescimento Fetal/patologia
3.
Autophagy ; 2023 Nov 29.
Artigo em Inglês | MEDLINE | ID: mdl-38018843

RESUMO

CARM1 (coactivator associated arginine methyltransferase 1) has recently emerged as a powerful regulator of skeletal muscle biology. However, the molecular mechanisms by which the methyltransferase remodels muscle remain to be fully understood. In this study, carm1 skeletal muscle-specific knockout (mKO) mice exhibited lower muscle mass with dysregulated macroautophagic/autophagic and atrophic signaling, including depressed AMP-activated protein kinase (AMPK) site-specific phosphorylation of ULK1 (unc-51 like autophagy activating kinase 1; Ser555) and FOXO3 (forkhead box O3; Ser588), as well as MTOR (mechanistic target of rapamycin kinase)-induced inhibition of ULK1 (Ser757), along with AKT/protein kinase B site-specific suppression of FOXO1 (Ser256) and FOXO3 (Ser253). In addition to lower mitophagy and autophagy flux in skeletal muscle, carm1 mKO led to increased mitochondrial PRKN/parkin accumulation, which suggests that CARM1 is required for basal mitochondrial turnover and autophagic clearance. carm1 deletion also elicited PPARGC1A (PPARG coactivator 1 alpha) activity and a slower, more oxidative muscle phenotype. As such, these carm1 mKO-evoked adaptations disrupted mitophagy and autophagy induction during food deprivation and collectively served to mitigate fasting-induced muscle atrophy. Furthermore, at the threshold of muscle atrophy during food deprivation experiments in humans, skeletal muscle CARM1 activity decreased similarly to our observations in mice, and was accompanied by site-specific activation of ULK1 (Ser757), highlighting the translational impact of the methyltransferase in human skeletal muscle. Taken together, our results indicate that CARM1 governs mitophagic, autophagic, and atrophic processes fundamental to the maintenance and remodeling of muscle mass. Targeting the enzyme may provide new therapeutic approaches for mitigating skeletal muscle atrophy.

4.
Front Endocrinol (Lausanne) ; 14: 1219205, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37842294

RESUMO

Preeclampsia (PE), a hypertensive pregnancy disorder, can originate from varied etiology. Placenta malperfusion has long been considered the primary cause of PE. However, we and others have showed that this disorder can also result from heightened inflammation at the maternal-fetal interface. To advance our understanding of this understudied PE subtype, it is important to establish validated rodent models to study the pathophysiology and test therapies. We evaluated three previously described approaches to induce inflammation-mediated PE-like features in pregnant rats: 1) Tumor necrosis factor-α (TNF-α) infusion via osmotic pump from gestational day (GD) 14-19 at 50ng/day/animal; 2) Polyinosinic:polycytidylic acid (Poly I:C) intraperitoneal (IP) injections from GD 10-18 (alternate days) at 10mg/kg/day/animal; and, 3) Lipopolysaccharide (LPS) IP injections from GD 13-18 at 20ug-70ug/kg/day per animal. Maternal blood pressure was measured by tail-cuff. Upon sacrifice, fetal and placenta weights were recorded. Placenta histomorphology was assessed using H&E sections. Placenta inflammation was determined by quantifying TNF-α levels and inflammatory gene expression. Placenta metabolic and mitochondrial health were determined by measuring mitochondrial respiration rates and placenta NAD+/NADH content. Of the three rodent models tested, we found that Poly I:C and LPS decreased both fetal weight and survival; and correlated with a reduction in region specific placenta growth. As the least effective model characterized, TNF-α treatment resulted in a subtle decrease in fetal/placenta weight and placenta mitochondrial respiration. Only the LPS model was able to induce maternal hypertension and exhibited pronounced placenta metabolic and mitochondrial dysfunction, common features of PE. Thus, the rat LPS model was most effective for recapitulating features observed in cases of human inflammatory PE. Future mechanistic and/or therapeutic intervention studies focuses on this distinct PE patient population may benefit from the employment of this rodent model of PE.


Assuntos
Hipertensão , Pré-Eclâmpsia , Gravidez , Feminino , Humanos , Ratos , Animais , Pré-Eclâmpsia/metabolismo , Fator de Necrose Tumoral alfa , Lipopolissacarídeos , Inflamação/metabolismo , Poli I
5.
Sci Rep ; 13(1): 8391, 2023 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-37225705

RESUMO

Maintaining mitochondrial function is critical to an improved healthspan and lifespan. Introducing mild stress by inhibiting mitochondrial translation invokes the mitochondrial unfolded protein response (UPRmt) and increases lifespan in several animal models. Notably, lower mitochondrial ribosomal protein (MRP) expression also correlates with increased lifespan in a reference population of mice. In this study, we tested whether partially reducing the gene expression of a critical MRP, Mrpl54, reduced mitochondrial DNA-encoded protein content, induced the UPRmt, and affected lifespan or metabolic health using germline heterozygous Mrpl54 mice. Despite reduced Mrpl54 expression in multiple organs and a reduction in mitochondrial-encoded protein expression in myoblasts, we identified few significant differences between male or female Mrpl54+/- and wild type mice in initial body composition, respiratory parameters, energy intake and expenditure, or ambulatory motion. We also observed no differences in glucose or insulin tolerance, treadmill endurance, cold tolerance, heart rate, or blood pressure. There were no differences in median life expectancy or maximum lifespan. Overall, we demonstrate that genetic manipulation of Mrpl54 expression reduces mitochondrial-encoded protein content but is not sufficient to improve healthspan in otherwise healthy and unstressed mice.


Assuntos
Longevidade , Ribossomos , Feminino , Masculino , Animais , Camundongos , Longevidade/genética , Composição Corporal , Expectativa de Vida , Expressão Gênica
6.
Int J Mol Sci ; 24(4)2023 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-36835587

RESUMO

The placenta is a vital organ of pregnancy, regulating adaptation to pregnancy, gestational parent/fetal exchange, and ultimately, fetal development and growth. Not surprisingly, in cases of placental dysfunction-where aspects of placental development or function become compromised-adverse pregnancy outcomes can result. One common placenta-mediated disorder of pregnancy is preeclampsia (PE), a hypertensive disorder of pregnancy with a highly heterogeneous clinical presentation. The wide array of clinical characteristics observed in pregnant individuals and neonates of a PE pregnancy are likely the result of distinct forms of placental pathology underlying the PE diagnosis, explaining why no one common intervention has proven effective in the prevention or treatment of PE. The historical paradigm of placental pathology in PE highlights an important role for utero-placental malperfusion, placental hypoxia and oxidative stress, and a critical role for placental mitochondrial dysfunction in the pathogenesis and progression of the disease. In the current review, the evidence of placental mitochondrial dysfunction in the context of PE will be summarized, highlighting how altered mitochondrial function may be a common feature across distinct PE subtypes. Further, advances in this field of study and therapeutic targeting of mitochondria as a promising intervention for PE will be discussed.


Assuntos
Hipertensão , Doenças Placentárias , Pré-Eclâmpsia , Recém-Nascido , Gravidez , Feminino , Humanos , Pré-Eclâmpsia/patologia , Placenta/patologia , Resultado da Gravidez , Hipertensão/patologia , Mitocôndrias/patologia , Doenças Placentárias/patologia
7.
Mol Metab ; 64: 101555, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-35872306

RESUMO

OBJECTIVE: Coactivator-associated arginine methyltransferase 1 (CARM1) catalyzes the methylation of arginine residues on target proteins to regulate critical processes in health and disease. A mechanistic understanding of the role(s) of CARM1 in skeletal muscle biology is only gradually emerging. The purpose of this study was to elucidate the function of CARM1 in regulating the maintenance and plasticity of skeletal muscle. METHODS: We used transcriptomic, methylproteomic, molecular, functional, and integrative physiological approaches to determine the specific impact of CARM1 in muscle homeostasis. RESULTS: Our data defines the occurrence of arginine methylation in skeletal muscle and demonstrates that this mark occurs on par with phosphorylation and ubiquitination. CARM1 skeletal muscle-specific knockout (mKO) mice displayed altered transcriptomic and arginine methylproteomic signatures with molecular and functional outcomes confirming remodeled skeletal muscle contractile and neuromuscular junction characteristics, which presaged decreased exercise tolerance. Moreover, CARM1 regulates AMPK-PGC-1α signalling during acute conditions of activity-induced muscle plasticity. CONCLUSIONS: This study uncovers the broad impact of CARM1 in the maintenance and remodelling of skeletal muscle biology.


Assuntos
Arginina , Transcriptoma , Animais , Arginina/metabolismo , Biologia , Camundongos , Músculo Esquelético/metabolismo , Proteína-Arginina N-Metiltransferases
8.
J Cell Biol ; 221(2)2022 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-35024765

RESUMO

Protein lysine acetylation is a post-translational modification that regulates protein structure and function. It is targeted to proteins by lysine acetyltransferases (KATs) or removed by lysine deacetylases. This work identifies a role for the KAT enzyme general control of amino acid synthesis protein 5 (GCN5; KAT2A) in regulating muscle integrity by inhibiting DNA binding of the transcription factor/repressor Yin Yang 1 (YY1). Here we report that a muscle-specific mouse knockout of GCN5 (Gcn5skm-/-) reduces the expression of key structural muscle proteins, including dystrophin, resulting in myopathy. GCN5 was found to acetylate YY1 at two residues (K392 and K393), disrupting the interaction between the YY1 zinc finger region and DNA. These findings were supported by human data, including an observed negative correlation between YY1 gene expression and muscle fiber diameter. Collectively, GCN5 positively regulates muscle integrity through maintenance of structural protein expression via acetylation-dependent inhibition of YY1. This work implicates the role of protein acetylation in the regulation of muscle health and for consideration in the design of novel therapeutic strategies to support healthy muscle during myopathy or aging.


Assuntos
Distrofina/genética , Músculos/metabolismo , Fator de Transcrição YY1/metabolismo , Fatores de Transcrição de p300-CBP/metabolismo , Acetilação , Envelhecimento/metabolismo , Animais , DNA/metabolismo , Distrofina/metabolismo , Regulação da Expressão Gênica , Humanos , Lisina/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Contração Muscular/genética , Fibras Musculares Esqueléticas/metabolismo , Músculos/patologia , Músculos/ultraestrutura , Atrofia Muscular/patologia , Distrofias Musculares/patologia , Transcriptoma/genética , Fatores de Transcrição de p300-CBP/deficiência
9.
Geroscience ; 44(1): 311-327, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34661816

RESUMO

Aging is associated with impaired vascular repair following ischemic insult, largely due to reparative dysfunctions of progenitor cells. Activation of Mas receptor (MasR) was shown to reverse aging-associated vasoreparative dysfunction. This study tested the impact of MasR-deficiency on mobilization and vasoreparative functions with aging. Wild type (WT) or MasR-deficient mice (MasR-/- or MasR+/-) at 12-14 weeks (young) or middle age (11-12 months) (MA) were used in the study. Mobilization of lineage-negative, Sca-1-positive cKit-positive (LSK) cells in response to G-CSF or plerixafor was determined. Hindlimb ischemia (HLI) was induced by femoral artery ligation. Mobilization and blood flow recovery were monitored post-HLI. Radiation chimeras were made by lethal irradiation of WT or MasR-/- mice followed by administration of bone marrow cells from MasR-/- or WT mice, respectively. Nitric oxide (NO) generation by stromal-derived factor-1α (SDF) and mitochondrial reactive oxygen species (mitoROS) levels were determined by flow cytometry. Effect of A779 treatment on mobilization, blood flow recovery, and NO and ROS levels were determined in young WT and MasR+/- mice. Circulating LSK cells in basal or in response to plerixafor or G-CSF or in response to ischemic injury were lower in MasR-/- mice compared to the WT. Responses in MasR+/- mice were similar to the WT at young age but at the middle age, impairments were observed. Impaired mobilization to ischemia or G-CSF was rescued in WT → MasR-/- chimeras. NO levels were lower and mitoROS were higher in MasR-/- LSK cells compared to WT cells. A779 precipitated dysfunctions in young-MasR+/- mice similar to that observed in MA-MasR+/-, and this accompanied decreased NO generation by SDF and enhanced mitoROS levels. This study shows that mice at MA do not exhibit vasoreparative dysfunction. Either partial or total loss of MasR precipitates advanced-aging phenotype likely due to lack of NO and oxidative stress.


Assuntos
Mobilização de Células-Tronco Hematopoéticas , Compostos Heterocíclicos , Envelhecimento , Animais , Camundongos , Fenótipo , Células-Tronco/fisiologia
10.
Nutrients ; 13(10)2021 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-34684467

RESUMO

Mitochondrial dysfunction is widely reported in various diseases and contributes to their pathogenesis. We assessed the effect of cocoa flavanols supplementation on mitochondrial function and whole metabolism, and we explored whether the mitochondrial deacetylase sirtuin-3 (Sirt3) is involved or not. We explored the effects of 15 days of CF supplementation in wild type and Sirt3-/- mice. Whole-body metabolism was assessed by indirect calorimetry, and an oral glucose tolerance test was performed to assess glucose metabolism. Mitochondrial respiratory function was assessed in permeabilised fibres and the pyridine nucleotides content (NAD+ and NADH) were quantified. In the wild type, CF supplementation significantly modified whole-body metabolism by promoting carbohydrate use and improved glucose tolerance. CF supplementation induced a significant increase of mitochondrial mass, while significant qualitative adaptation occurred to maintain H2O2 production and cellular oxidative stress. CF supplementation induced a significant increase in NAD+ and NADH content. All the effects mentioned above were blunted in Sirt3-/- mice. Collectively, CF supplementation boosted the NAD metabolism that stimulates sirtuins metabolism and improved mitochondrial function, which likely contributed to the observed whole-body metabolism adaptation, with a greater ability to use carbohydrates, at least partially through Sirt3.


Assuntos
Cacau/química , Suplementos Nutricionais , Metabolismo Energético/efeitos dos fármacos , Flavonoides/farmacologia , Mitocôndrias Musculares/efeitos dos fármacos , Mitocôndrias Musculares/metabolismo , Extratos Vegetais/farmacologia , Animais , Biomarcadores , Composição Corporal , Flavonoides/química , Glucose/metabolismo , Masculino , Camundongos , Camundongos Knockout , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Oxirredução/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/química , Sirtuína 3/genética , Sirtuína 3/metabolismo
11.
Commun Biol ; 4(1): 989, 2021 08 19.
Artigo em Inglês | MEDLINE | ID: mdl-34413467

RESUMO

Mouse models of genetic mitochondrial disorders are generally used to understand specific molecular defects and their biochemical consequences, but rarely to map compensatory changes allowing survival. Here we took advantage of the extraordinary mitochondrial resilience of hepatic Lrpprc knockout mice to explore this question using native proteomics profiling and lipidomics. In these mice, low levels of the mtRNA binding protein LRPPRC induce a global mitochondrial translation defect and a severe reduction (>80%) in the assembly and activity of the electron transport chain (ETC) complex IV (CIV). Yet, animals show no signs of overt liver failure and capacity of the ETC is preserved. Beyond stimulation of mitochondrial biogenesis, results show that the abundance of mitoribosomes per unit of mitochondria is increased and proteostatic mechanisms are induced in presence of low LRPPRC levels to preserve a balance in the availability of mitochondrial- vs nuclear-encoded ETC subunits. At the level of individual organelles, a stabilization of residual CIV in supercomplexes (SCs) is observed, pointing to a role of these supramolecular arrangements in preserving ETC function. While the SC assembly factor COX7A2L could not contribute to the stabilization of CIV, important changes in membrane glycerophospholipid (GPL), most notably an increase in SC-stabilizing cardiolipins species (CLs), were observed along with an increased abundance of other supramolecular assemblies known to be stabilized by, and/or participate in CL metabolism. Together these data reveal a complex in vivo network of molecular adjustments involved in preserving mitochondrial integrity in energy consuming organs facing OXPHOS defects, which could be therapeutically exploited.


Assuntos
Mitocôndrias/metabolismo , Proteínas Mitocondriais/genética , Proteínas de Neoplasias/genética , Animais , Feminino , Masculino , Camundongos , Camundongos Knockout , Proteínas Mitocondriais/metabolismo , Proteínas de Neoplasias/metabolismo , Biossíntese de Proteínas
12.
FASEB J ; 35(4): e21278, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33769614

RESUMO

Mitochondria share attributes of vesicular transport with their bacterial ancestors given their ability to form mitochondrial-derived vesicles (MDVs). MDVs are involved in mitochondrial quality control and their formation is enhanced with stress and may, therefore, play a potential role in mitochondrial-cellular communication. However, MDV proteomic cargo has remained mostly undefined. In this study, we strategically used an in vitro MDV budding/reconstitution assay on cardiac mitochondria, followed by graded oxidative stress, to identify and characterize the MDV proteome. Our results confirmed previously identified cardiac MDV markers, while also revealing a complete map of the MDV proteome, paving the way to a better understanding of the role of MDVs. The oxidative stress vulnerability of proteins directed the cargo loading of MDVs, which was enhanced by antimycin A (Ant-A). Among OXPHOS complexes, complexes III and V were found to be Ant-A-sensitive. Proteins from metabolic pathways such as the TCA cycle and fatty acid metabolism, along with Fe-S cluster, antioxidant response proteins, and autophagy were also found to be Ant-A sensitive. Intriguingly, proteins containing hyper-reactive cysteine residues, metabolic redox switches, including professional redox enzymes and those that mediate iron metabolism, were found to be components of MDV cargo with Ant-A sensitivity. Last, we revealed a possible contribution of MDVs to the formation of extracellular vesicles, which may indicate mitochondrial stress. In conclusion, our study provides an MDV proteomics signature that delineates MDV cargo selectivity and hints at the potential for MDVs and their novel protein cargo to serve as vital biomarkers during mitochondrial stress and related pathologies.


Assuntos
Mitocôndrias Cardíacas/fisiologia , Estresse Oxidativo , Vesículas Transportadoras/fisiologia , Animais , Linhagem Celular , Regulação da Expressão Gênica , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Mioblastos , Proteômica , Ratos
13.
Biochim Biophys Acta Mol Basis Dis ; 1865(4): 782-796, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30472159

RESUMO

Recent studies have led to the discovery of multiple mitochondrial quality control (mQC) processes that operate at various scales, ranging from the degradation of proteins by mitochondrial proteases to the degradation of selected cargos or entire organelles in lysosomes. While the mechanisms governing these mQC processes are progressively being delineated, their role and importance remain unclear. Converging evidence however point to a complex system whereby multiple and partly overlapping processes are recruited to orchestrate a cell type specific mQC response that is adapted to the physiological state and level of stress encountered. Knowledge gained from basic model systems of mQC therefore need to be integrated within organ-specific (patho)physiological frameworks. Building on this notion, this article focuses on mQC in the heart, where developmental metabolic reprogramming, sustained contraction, and multiple pathophysiological conditions pose broadly different constraints. We provide an overview of current knowledge of mQC processes, and discuss their implication in cardiac mQC under normal and diseased conditions.


Assuntos
Mitocôndrias Cardíacas/metabolismo , Miocárdio/metabolismo , Proteostase , Animais , Humanos , Estresse Oxidativo , Peptídeo Hidrolases/metabolismo
14.
Diabetes ; 66(2): 505-518, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27856608

RESUMO

The angiotensin (ANG)-(1-7)/Mas receptor (MasR) pathway activates vascular repair-relevant functions of bone marrow progenitor cells. We tested the effects of ANG-(1-7) on mobilization and vasoreparative functions of progenitor cells that are impaired in diabetes. The study was performed in streptozotocin-induced diabetic (db/db) mice. Diabetes resulted in a decreased number of Lineage-Sca-1+c-Kit+ (LSK) cells in the circulation, which was normalized by ANG-(1-7). Diabetes-induced depletion of LSK cells in the bone marrow was reversed by ANG-(1-7). ρ-Kinase (ROCK) activity was increased specifically in bone marrow LSK cells by ANG-(1-7) in diabetes, and the beneficial effects of ANG-(1-7) were prevented by fasudil. ANG-(1-7) increased Slit3 levels in the bone marrow supernatants, which activated ROCK in LSK cells and sensitized them for stromal-derived factor-1α (SDF)-induced migration. Diabetes prevented the mobilization of LSK cells in response to ischemia and impaired the recovery of blood flow, both of which were reversed by ANG-(1-7) in both models of diabetes. Genetic ablation of MasR prevented ischemia-induced mobilization of LSK cells and impaired blood flow recovery, which was associated with decreased proliferation and migration of LSK cells in response to SDF or vascular endothelial growth factor. These results suggest that MasR is a promising target for the treatment of diabetic bone marrow mobilopathy and vascular disease.


Assuntos
Angiotensina I/farmacologia , Vasos Sanguíneos/efeitos dos fármacos , Células da Medula Óssea/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Diabetes Mellitus Experimental/fisiopatologia , Neovascularização Fisiológica/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Células-Tronco/efeitos dos fármacos , Vasodilatadores/farmacologia , 1-(5-Isoquinolinasulfonil)-2-Metilpiperazina/análogos & derivados , 1-(5-Isoquinolinasulfonil)-2-Metilpiperazina/farmacologia , Animais , Vasos Sanguíneos/fisiopatologia , Medula Óssea/efeitos dos fármacos , Medula Óssea/fisiopatologia , Linhagem da Célula , Quimiocina CXCL12/efeitos dos fármacos , Quimiocina CXCL12/metabolismo , Isquemia , Masculino , Proteínas de Membrana/efeitos dos fármacos , Proteínas de Membrana/metabolismo , Camundongos , Inibidores de Proteínas Quinases/farmacologia , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas/genética , Receptores Acoplados a Proteínas G/genética , Recuperação de Função Fisiológica/efeitos dos fármacos , Regeneração , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismo , Quinases Associadas a rho/efeitos dos fármacos , Quinases Associadas a rho/metabolismo
15.
Sci Rep ; 6: 26131, 2016 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-27188595

RESUMO

Diabetes is associated with impaired mobilization of bone marrow stem/progenitor cells that accelerate vascularization of ischemic areas. This study characterized mobilization of vascular reparative bone marrow progenitor cells in mouse models of diabetes. Age-matched control or streptozotocin (STZ)-induced diabetic, and db/db mice with lean-controls were studied. Mobilization induced by G-CSF, AMD3100 or ischemia was evaluated by flow cytometric enumeration of circulating Lin(-)Sca-1(+)cKit(+) (LSK) cells, and by colony forming unit (CFU) assay. The circulating WBCs and LSKs, and CFUs were reduced in both models with a shorter duration (10-12 weeks) of diabetes compared to their respective controls. Longer duration of STZ-diabetes (≥20 weeks) induced impairment of G-CSF- or AMD3100-mobilization (P < 0.01, n = 8). In db/db mice, mobilization by G-CSF or AMD3100 was either increased or unaffected (P < 0.05, n = 6 to 8). Proliferation, migration, and ischemia-induced mobilization, of LSK cells were impaired in both models. Leptin receptor antagonist, PESLAN-1, increased G-CSF- or AMD3100-mobilization of WBCs and LSKs, compared to the untreated. Leptin increased basal WBCs, decreased basal and AMD3100-mobilized LSK cells, and had no effect on G-CSF. These results suggest that mobilopathy is apparent in STZ-diabetes but not in db/db mice. Leptin receptor antagonism would be a promising approach for reversing diabetic bone marrow mobilopathy.


Assuntos
Células da Medula Óssea/fisiologia , Diabetes Mellitus Experimental/patologia , Neovascularização Fisiológica , Receptores para Leptina/deficiência , Células-Tronco/fisiologia , Animais , Movimento Celular , Proliferação de Células , Ensaio de Unidades Formadoras de Colônias , Citometria de Fluxo , Camundongos
16.
Eur J Pharmacol ; 774: 25-33, 2016 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-26851370

RESUMO

Angiotensin-converting enzymes, ACE and ACE2, are key members of renin angiotensin system. Activation of ACE2/Ang-(1-7) pathway enhances cardiovascular protective functions of bone marrow-derived stem/progenitor cells. The current study evaluated the selectivity of ACE2 inhibitors, MLN-4760 and DX-600, and ACE and ACE2 activities in human (hu) and murine (mu) bone marrow cells. Assays were carried out in hu and mu mononuclear cells (MNCs) and huCD34(+) cells or mu-lineage-depleted (muLin(-)) cells, human-recombinant (rh) enzymes, and mu-heart with enzyme-specific substrates. ACE or ACE2 inhibition by racemic MLN-4760, its isomers MLN-4760-A and MLN-4760-B, DX600 and captopril were characterized. MLN-4760-B is relatively less efficacious and less-selective than the racemate or MLN-4760-A at hu-rhACE2, and all three of them inhibited 43% rhACE. In huMNCs, MLN-4760-B detected 63% ACE2 with 28-fold selectivity over ACE. In huCD34(+) cells, MLN-4760-B detected 38% of ACE2 activity with 63-fold selectivity. In mu-heart and muMNCs, isomer B was 100- and 228-fold selective for ACE2, respectively. In muLin(-) cells, MLN-4760-B detected 25% ACE2 activity with a pIC50 of 6.3. The racemic mixture and MLN-4760-A showed lower efficacy and poor selectivity for ACE2 in MNCs and mu-heart. ACE activity detected by captopril was 32% and 19%, respectively, in huCD34(+) and muLin(-) cells. DX600 was less efficacious, and more selective for ACE2 compared to MLN-4760-B in all samples tested. These results suggest that MLN-4760-B is a better antagonist of ACE2 than DX600 at 10 µm concentration in human and murine bone marrow cells, and that these cells express more functional ACE2 than ACE.


Assuntos
Inibidores da Enzima Conversora de Angiotensina/metabolismo , Células da Medula Óssea/citologia , Imidazóis/metabolismo , Leucina/análogos & derivados , Peptídeos/metabolismo , Peptidil Dipeptidase A/metabolismo , Idoso , Sequência de Aminoácidos , Enzima de Conversão de Angiotensina 2 , Animais , Células da Medula Óssea/metabolismo , Feminino , Humanos , Leucina/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Miocárdio/citologia , Miocárdio/metabolismo , Peptídeos/química , Especificidade por Substrato
17.
J Sex Med ; 11(9): 2153-63, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24953642

RESUMO

INTRODUCTION: Angiotensin (Ang)-(1-7) is a recently identified vasoprotective heptapeptide, and it appears to activate the reparative functions of bone marrow-derived stem/progenitor cells (BMPCs). AIM: This study evaluated the effect of Ang-(1-7) in the angiogenic function of cavernosum in type 1 diabetes (T1D) and delineated the role of BMPCs in this protective function. METHODS: T1D was induced by streptozotocin in mice, and mice with 20-24 weeks of diabetes were used for the study. Ang-(1-7) was administered subcutaneously by using osmotic pumps. Cavernosa, and BMPCs from peripheral blood and bone marrow were evaluated in different assay systems. MAIN OUTCOME MEASURES: Angiogenic function was determined by endothelial tube formation in matrigel assay. Circulating BMPCs were enumerated by flow cytometry and proliferation was determined by BrdU incorporation. Cell-free supernatant of BMPCs were collected and tested for paracrine angiogenic effect. Expression of angiogenic factors in BMPCs and cavernosa were determined by real-time polymerase chain reaction. RESULTS: Ang-(1-7) (100 nM) stimulated angiogenesis in mouse cavernosum that was partially inhibited by Mas1 receptor antagonist, A779 (10 µM) (P < 0.05). In cavernosa of T1D, the angiogenic responses to Ang-(1-7) (P < 0.005) and VEGF (100 nM) (P < 0.03) were diminished. Ang-(1-7) treatment for 4 weeks reversed T1D-induced decrease in the VEGF-mediated angiogenesis. Ang-(1-7) treatment increased the circulating number of BMPCs and proliferation that were decreased in T1D (P < 0.02). Paracrine angiogenic function of BMPCs was reduced in diabetic BMPCs, which was reversed by Ang-(1-7). In diabetic BMPCs, SDF and angiopoietin-1 were upregulated by Ang-(1-7), and in cavernosum, VEGFR1, Tie-2, and SDF were upregulated and angiopoietin-2 was down-regulated. CONCLUSIONS: Ang-(1-7) stimulates angiogenic function of cavernosum in diabetes via its stimulating effects on both cavernosal microvasculature and BMPCs.


Assuntos
Angiotensina I/farmacologia , Células da Medula Óssea/efeitos dos fármacos , Diabetes Mellitus Experimental/fisiopatologia , Microvasos/efeitos dos fármacos , Neovascularização Patológica/prevenção & controle , Pênis/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Animais , Diabetes Mellitus Tipo 1/fisiopatologia , Citometria de Fluxo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pênis/irrigação sanguínea , Pênis/metabolismo , Proto-Oncogene Mas , Reação em Cadeia da Polimerase em Tempo Real , Fator A de Crescimento do Endotélio Vascular/farmacologia
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