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The placenta plays an essential role in pregnancy, leading to proper fetal development and growth. As an organ with multiple physiological functions for both mother and fetus, it is a highly energetic and metabolically demanding tissue. Mitochondrial physiology plays a crucial role in the metabolism of this organ and thus any alteration leading to mitochondrial dysfunction has a severe outcome in the development of the fetus. Pregnancy-related pathological states with a mitochondrial dysfunction outcome include preeclampsia and gestational diabetes mellitus. In this review, we address the role of mitochondrial morphology, metabolism and physiology of the placenta during pregnancy, highlighting the roles of the cytotrophoblast and syncytiotrophoblast. We also describe the relationship between preeclampsia, gestational diabetes, gestational diabesity and pre-pregnancy maternal obesity with mitochondrial dysfunction.
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Maternal obesity (MO) is a significant cause of increased cardiometabolic risk in offspring, who present endothelial dysfunction at birth. Alterations in physiologic and cellular redox status are strongly associated with altered gene regulation in arterial endothelium. However, specific mechanisms by which the pro-oxidant fetal environment in MO could modulate the vascular gene expression and function during the offspring's postnatal life are elusive. We tested if oxidative stress could reprogram the antioxidant-coding gene's response to a pro-oxidant challenge through an epigenetic transcriptional memory (ETM) mechanism. A pro-oxidant double-hit protocol was applied to human umbilical artery endothelial cells (HUAECs) and EA.hy 926 endothelial cell lines. The ETM acquisition in the HMOX1 gene was analyzed by RT-qPCR. HMOX1 mRNA decay was evaluated by Actinomycin-D treatment and RT-qPCR. To assess the chromatin accessibility and the enrichment of NRF2, RNAP2, and phosphorylation at serin-5 of RNAP2, at HMOX1 gene regulatory regions, were used DNase HS-qPCR and ChIP-qPCR assays, respectively. The CpG methylation pattern at the HMOX1 core promoter was analyzed by DNA bisulfite conversion and Sanger sequencing. Data were analyzed using two-way ANOVA, and p < 0.05 was statistically significant. Using a pro-oxidant double-hit protocol, we found that the Heme Oxygenase gene (HMOX1) presents an ETM response associated with changes in the chromatin structure at the promoter and gene regulatory regions. The ETM response was characterized by a paused-RNA Polymerase 2 and NRF2 enrichment at the transcription start site and Enhancer 2 of the HMOX1 gene, respectively. Changes in DNA methylation pattern at the HMOX1 promoter were not a hallmark of this oxidative stress-induced ETM. These data suggest that a pro-oxidant milieu could trigger an ETM at the vascular level, indicating a potential epigenetic mechanism involved in the increased cardiovascular risk in the offspring of women with obesity.
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BACKGROUND: Ex-vivo myography enables the assessment of muscle electrical activity response. This study explored the viability of determining the physiological responses in muscles without tendon, as rectus abdominis muscle (RAM), through ex-vivo myography to assess its potential as a diagnostic tool. RESULTS: All tested RAM samples (five different samples) show patterns of electrical activity. A positive response was observed in 100% of the programmed stimulation. RAM 3 showed greater weight (0.47 g), length (1.66 cm), and width (0.77 cm) compared to RAM 1, RAM 2, RAM 4 and RAM 5 with more sustained electrical activity over time, a higher percentage of fatigue was analyzed at half the time of the electrical activity. The order of electrical activity (Mn) was RAM 3 > RAM 5 > RAM 1 > RAM 4 > RAM 2. No electrical activity was recorded in the Sham group. CONCLUSIONS: This study shows that it is feasible to assess the physiological responses of striated muscle without tendon as RAM, obtained at C-section, under ex vivo myography. These results could be recorded, properly analyzed, and demonstrated its potential as a diagnostic tool for rectus abdominis muscle electrical activity.
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Cesárea , Reto do Abdome , Estudos de Coortes , Feminino , Humanos , Miografia , GravidezRESUMO
Gestational diabetes mellitus (GDM) is recognized as a "window of opportunity" for the future prediction of such complications as type 2 diabetes mellitus and pelvic floor muscle disorders, including urinary incontinence and genitourinary dysfunction. Translational studies have reported that pelvic floor muscle disorders are due to a GDM-induced-myopathy (GDiM) of the pelvic floor muscle and rectus abdominis muscle (RAM). We now describe the transcriptome profiling of the RAM obtained by Cesarean section from GDM and non-GDM women with and without pregnancy-specific urinary incontinence (PSUI). We identified 650 genes in total, and the differentially expressed genes were defined by comparing three control groups to the GDM with PSUI group (GDiM). Enrichment analysis showed that GDM with PSUI was associated with decreased gene expression related to muscle structure and muscle protein synthesis, the reduced ability of muscle fibers to ameliorate muscle damage, and the altered the maintenance and generation of energy through glycogenesis. Potential genetic muscle biomarkers were validated by RT-PCR, and their relationship to the pathophysiology of the disease was verified. These findings help elucidate the molecular mechanisms of GDiM and will promote the development of innovative interventions to prevent and treat complications such as post-GDM urinary incontinence.
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Diabetes Mellitus Tipo 2 , Diabetes Gestacional , Doenças Musculares , Incontinência Urinária , Gravidez , Humanos , Feminino , Diabetes Gestacional/metabolismo , Reto do Abdome/metabolismo , Cesárea/efeitos adversos , Diabetes Mellitus Tipo 2/complicações , Transcriptoma , Incontinência Urinária/genética , Biomarcadores , Perfilação da Expressão GênicaRESUMO
Insulin regulates the l-arginine/nitric oxide (NO) pathway in human umbilical vein endothelial cells (HUVECs), increasing the plasma membrane expression of the l-arginine transporter hCAT-1 and inducing vasodilation in umbilical and placental veins. Placental vascular relaxation induced by insulin is dependent of large conductance calcium-activated potassium channels (BKCa), but the role of KCa channels on l-arginine transport and NO synthesis is still unknown. The aim of this study was to determine the contribution of KCa channels in both insulin-induced l-arginine transport and NO synthesis, and its relationship with placental vascular relaxation. HUVECs, human placental vein endothelial cells (HPVECs) and placental veins were freshly isolated from umbilical cords and placenta from normal pregnancies. Cells or tissue were incubated in absence or presence of insulin and/or tetraethylammonium, 1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole, iberiotoxin or NG-nitro-l-arginine methyl ester. l-Arginine uptake, plasma membrane polarity, NO levels, hCAT-1 expression and placenta vascular reactivity were analyzed. The inhibition of intermediate-conductance KCa (IKCa) and BKCa increases l-arginine uptake, which was related with protein abundance of hCAT-1 in HUVECs. IKCa and BKCa activities contribute to NO-synthesis induced by insulin but are not directly involved in insulin-stimulated l-arginine uptake. Long term incubation (8 h) with insulin increases the plasma membrane hyperpolarization and hCAT-1 expression in HUVECs and HPVECs. Insulin-induced relaxation in placental vasculature was reversed by KCa inhibition. The results show that the activity of IKCa and BKCa channels are relevant for both physiological regulations of NO synthesis and vascular tone regulation in the human placenta, acting as a part of negative feedback mechanism for autoregulation of l-arginine transport in HUVECs.
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Canais de Potássio Ativados por Cálcio de Condutância Intermediária/metabolismo , Canais de Potássio Ativados por Cálcio de Condutância Alta/metabolismo , Óxido Nítrico/metabolismo , Veias Umbilicais/metabolismo , Adulto , Arginina/metabolismo , Transportador 1 de Aminoácidos Catiônicos/metabolismo , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Feminino , Células Endoteliais da Veia Umbilical Humana , Humanos , Insulina/farmacologia , Canais de Potássio Ativados por Cálcio de Condutância Intermediária/antagonistas & inibidores , Canais de Potássio Ativados por Cálcio de Condutância Alta/antagonistas & inibidores , Peptídeos/farmacologia , Placenta/efeitos dos fármacos , Placenta/metabolismo , Bloqueadores dos Canais de Potássio/farmacologia , Gravidez , Pirazóis/farmacologia , Veias Umbilicais/efeitos dos fármacos , Adulto JovemRESUMO
Gestational diabetes mellitus (GDM) characterizes by foetoplacental endothelial dysfunction. Human umbilical vein endothelial cells (HUVECs) from women with GDM show increased L-arginine transport via the human cationic amino acid transporter 1 (hCAT-1). Moreover, expression of endothelial nitric oxide synthase (eNOS) and nitric oxide synthesis are increased. Exosomes are increased in maternal plasma from GDM. We evaluated the role of foetoplacental endothelial exosomes on endothelial dysfunction in GDM. Exosomes were isolated from HUVECs from normal (ExN) and GDM (ExGDM) pregnancies. HUVECs were exposed (8h) to ExN or ExGDM and used for wound recovery assay (up to 8h), L-arginine transport, hCAT-1 and eNOS expression and activity, reactive oxygen species (ROS) generation, and 44 and 42kDa mitogen activated protein kinases (p44/42mapk) and protein kinase B/Akt (Akt) activation. Wound recovery was slower in GDM compared with normal pregnancies and was recovered by ExN. However, ExGDM delayed wound recovery in cells from normal pregnancies. GDM-increased L-arginine transport, hCAT-1 and eNOS expression and activity, and p44/42mapk activation were blocked by ExN, but ExGDM increased these parameters and ROS generation, and reduced eNOS phosphorylation at threonine495 in cells from normal pregnancies. Inhibition of p44/42mapk, but not Akt reversed GDM-increased L-arginine uptake. In conclusion foetoplacental endothelial-released exosomes play a role in the maintenance of a GDM phenotype in HUVECs. It is suggested that ExN and ExGDM cargo are different with differential effects in cells from normal or GDM pregnancies. This phenomenon could contribute to the understanding of mechanisms behind foetoplacental endothelial dysfunction in GDM pregnancies.
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Diabetes Gestacional/metabolismo , Endotélio Vascular/metabolismo , Exossomos/metabolismo , Placenta/metabolismo , Adulto , Arginina/química , Transporte Biológico , Transportador 1 de Aminoácidos Catiônicos/metabolismo , Feminino , Teste de Tolerância a Glucose , Células Endoteliais da Veia Umbilical Humana , Humanos , Recém-Nascido , Insulina/metabolismo , Masculino , Óxido Nítrico/química , Óxido Nítrico Sintase Tipo III , Fosforilação , Gravidez , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Veias Umbilicais/metabolismoRESUMO
Gestational diabetes mellitus (GDM) is a disease characterised by glucose intolerance and first diagnosed in pregnancy. This condition relates to an anomalous placental environment and aberrant placental vascular function. GDM-associated hyperglycaemia changes the placenta structure leading to abnormal development and functionality of this vital organ. Aiming to avoid the GDM-hyperglycaemia and its deleterious consequences in the mother, the foetus and newborn, women with GDM are firstly treated with a controlled diet therapy; however, some of the women fail to reach the recommended glycaemia values and therefore they are passed to the second line of treatment, i.e., insulin therapy. The several protocols available in the literature regarding insulin therapy are variable and not a clear consensus is yet reached. Insulin therapy restores maternal glycaemia, but this beneficial effect is not reflected in the foetus and newborn metabolism, suggesting that other factors than d-glucose may be involved in the pathophysiology of GDM. Worryingly, insulin therapy may cause alterations in the placenta and umbilical vessels as well as the foetus and newborn additional to those seen in pregnant women with GDM treated with diet. In this review, we summarised the variable information regarding indications and protocols for administration of the insulin therapy and the possible outcomes on the function and structure of the foetoplacental unit and the neonate parameters from women with GDM.
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Glicemia/efeitos dos fármacos , Diabetes Gestacional/tratamento farmacológico , Macrossomia Fetal/prevenção & controle , Insulina/uso terapêutico , Placenta/efeitos dos fármacos , Diabetes Gestacional/sangue , Diabetes Gestacional/dietoterapia , Feminino , Macrossomia Fetal/epidemiologia , Macrossomia Fetal/etiologia , Teste de Tolerância a Glucose , Humanos , Incidência , Recém-Nascido , GravidezRESUMO
l-Arginine is taken up via the cationic amino acid transporters (system y+/CATs) and system y+L in human umbilical vein endothelial cells (HUVECs). l-Arginine is the substrate for endothelial NO synthase (eNOS) which is activated by intracellular alkalization, but nothing is known regarding modulation of system y+/CATs and system y+L activity, and eNOS activity by the pHi in HUVECs. We studied whether an acidic pHi modulates l-arginine transport and eNOS activity in HUVECs. Cells loaded with a pH-sensitive probe were subjected to 0.1-20â¯mmol/L NH4Cl pulse assay to generate pHi 7.13-6.55. Before pHi started to recover, l-arginine transport (0-20 or 0-1000⯵mol/L, 10â¯s, 37⯰C) in the absence or presence of 200⯵mol/L N-ethylmaleimide (NEM) (system y+/CATs inhibitor) or 2â¯mmol/L l-leucine (systemy+L substrate) was measured. Protein abundance for eNOS and serine1177 or threonine495 phosphorylated eNOS was determined. The results show that intracellular acidification reduced system y+L but not system y+/CATs mediated l-arginine maximal transport capacity due to reduced maximal velocity. Acidic pHi reduced NO synthesis and eNOS serine1177 phosphorylation. Thus, system y+L activity is downregulated by an acidic pHi, a phenomenon that may result in reduced NO synthesis in HUVECs.
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Sistema y+L de Transporte de Aminoácidos/metabolismo , Sistema y+ de Transporte de Aminoácidos/metabolismo , Arginina/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Óxido Nítrico/biossíntese , Sistema y+ de Transporte de Aminoácidos/antagonistas & inibidores , Transporte Biológico Ativo/efeitos dos fármacos , Transporte Biológico Ativo/fisiologia , Etilmaleimida/farmacologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Concentração de Íons de HidrogênioRESUMO
Maternal physiological hypercholesterolemia (MPH) allows a proper foetal development; however, maternal supraphysiological hypercholesterolemia (MSPH) associates with foetal endothelial dysfunction and early development of atherosclerosis. MSPH courses with reduced endothelium-dependent dilation of the human umbilical vein due to reduced endothelial nitric oxide synthase activity compared with MPH. Whether MSPH modifies the availability of the nitric oxide synthase cofactor tetrahydrobiopterin is unknown. We investigated whether MSPH-associated lower umbilical vein vascular reactivity results from reduced bioavailability of tetrahydrobiopterin. Total cholesterol <7.2mmol/L was considered as maternal physiological hypercholesterolemia (n=72 women) and ≥7.2mmol/L as MSPH (n=35 women). Umbilical veins rings were used for vascular reactivity assays (wire myography), and primary cultures of human umbilical vein endothelial cells (HUVECs) to measure nitric oxide synthase, GTP cyclohydrolase 1, and dihydrofolate reductase expression and activity, as well as tetrahydrobiopterin content. MSPH reduced the umbilical vein rings relaxation caused by calcitonine gene-related peptide, a phenomenon partially improved by incubation with sepiapterin. HUVECs from MSPH showed lower nitric oxide synthase activity (l-citrulline synthesis from l-arginine) without changes in its protein abundance, as well as reduced tetrahydrobiopterin level compared with MPH, a phenomenon reversed by incubation with sepiapterin. Expression and activity of GTP cyclohydrolase 1 was lower in MSPH, without changes in dihydrofolate reductase expression. MSPH is a pathophysiological condition reducing human umbilical vein reactivity due to lower bioavailability of tetrahydrobiopterin leading to lower NOS activity in the human umbilical vein endothelium.
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Células Endoteliais da Veia Umbilical Humana/metabolismo , Hipercolesterolemia/metabolismo , Complicações na Gravidez/metabolismo , Pterinas/farmacologia , Veias Umbilicais/metabolismo , Adolescente , Adulto , Feminino , GTP Cicloidrolase/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Hipercolesterolemia/patologia , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase/metabolismo , Gravidez , Complicações na Gravidez/patologia , Veias Umbilicais/patologiaRESUMO
Cancer cells generate protons (H+) that are extruded to the extracellular medium mainly via the Na+/H+ exchanger 1 (NHE1), which regulates intracellular pH (pHi) and cell proliferation. In primary cultures of human ascites-derived ovarian cancer cells (haOC) we assayed whether NHE1 was required for pHi modulation and cell proliferation. Human ovary expresses NHE1, which is higher in haOC and A2780 (ovarian cancer cells) compared with HOSE cells (normal ovarian cells). Basal pHi and pHi recovery (following a NH4Cl pulse) was higher in haOC and A2780, compared with HOSE cells. Zoniporide (NHE1 inhibitor) caused intracellular acidification and pHi recovery was independent of intracellular buffer capacity, but reduced in NHE1 knockdown A2780 cells. Zoniporide reduced the maximal proliferation capacity, cell number, thymidine incorporation, and ki67 (marker of proliferation) fluorescence in haOC cells. SLC9A1 (for NHE1) amplification associated with lower overall patient survival. In conclusion, NHE1 is expressed in human ovarian cancer where it has a pro-proliferative role. Increased NHE1 expression and activity constitute an unfavourable prognostic factor in these patients.
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Neoplasias Ovarianas/metabolismo , Trocador 1 de Sódio-Hidrogênio/metabolismo , Proliferação de Células , Células Cultivadas , Feminino , Humanos , Concentração de Íons de Hidrogênio , Neoplasias Ovarianas/patologia , Ovário/metabolismo , Ovário/patologia , Trocador 1 de Sódio-Hidrogênio/análiseRESUMO
Pregnant women diagnosed with gestational diabetes mellitus subjected to diet (GDMd) that do not reach normal glycaemia are passed to insulin therapy (GDMi). GDMd associates with increased human cationic amino acid transporter 1 (hCAT-1)-mediated transport of L-arginine and nitric oxide synthase (NOS) activity in foetoplacental vasculature, a phenomenon reversed by exogenous insulin. Whether insulin therapy results in reversal of the GDMd effect on the foetoplacental vasculature is unknown. We assayed whether insulin therapy normalizes GDMd-associated foetoplacental endothelial dysfunction. Primary cultures of human umbilical vein endothelial cells (HUVECs) from GDMi pregnancies were used to assay L-arginine transport kinetics, NOS activity, p44/42mapk and protein kinase B/Akt activation, and umbilical vein rings reactivity. HUVECs from GDMi or GDMd show increased hCAT-1 expression and maximal transport capacity, NOS activity, and eNOS, and p44/42mapk, but not Akt activator phosphorylation. Dilation in response to insulin or calcitonin-gene related peptide was impaired in umbilical vein rings from GDMi and GDMd pregnancies. Incubation of HUVECs in vitro with insulin (1 nmol/L) restored hCAT-1 and eNOS expression and activity, and eNOS and p44/42mapk activator phosphorylation. Thus, maternal insulin therapy does not seem to reverse GDMd-associated alterations in human foetoplacental vasculature.
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Diabetes Gestacional , Endotélio Vascular/metabolismo , Insulina/administração & dosagem , Placenta/metabolismo , Adulto , Transportador 1 de Aminoácidos Catiônicos/metabolismo , Diabetes Gestacional/dietoterapia , Diabetes Gestacional/tratamento farmacológico , Diabetes Gestacional/metabolismo , Diabetes Gestacional/patologia , Endotélio Vascular/patologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Proteína Quinase 3 Ativada por Mitógeno/biossíntese , Óxido Nítrico Sintase Tipo III/biossíntese , Fosforilação/efeitos dos fármacos , Placenta/patologia , Gravidez , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Nitric oxide plays several roles in cellular physiology, including control of the vascular tone and defence against pathogen infection. Neuronal, inducible and endothelial nitric oxide synthase (NOS) isoforms synthesize nitric oxide. Cells generate acid and base equivalents, whose physiological intracellular concentrations are kept due to membrane transport systems, including Na+ /H+ exchangers and Na+ /HCO3- transporters, thus maintaining a physiological pH at the intracellular (~7.0) and extracellular (~7.4) medium. In several pathologies, including cancer, cells are exposed to an extracellular acidic microenvironment, and the role for these membrane transport mechanisms in this phenomenon is likely. As altered NOS expression and activity is seen in cancer cells and because this gas promotes a glycolytic phenotype leading to extracellular acidosis in gynaecological cancer cells, a pro-inflammatory microenvironment increasing inducible NOS expression in this cell type is feasible. However, whether abnormal control of intracellular and extracellular pH by cancer cells regards with their ability to synthesize or respond to nitric oxide is unknown. We, here, discuss a potential link between pH alterations, pH controlling membrane transport systems and NOS function. We propose a potential association between inducible NOS induction and Na+ /H+ exchanger expression and activity in human ovary cancer. A potentiation between nitric oxide generation and the maintenance of a low extracellular pH (i.e. acidic) is proposed to establish a sequence of events in ovarian cancer cells, thus preserving a pro-proliferative acidic tumour extracellular microenvironment. We suggest that pharmacological therapeutic targeting of Na+ /H+ exchangers and inducible NOS may have benefits in human epithelial ovarian cancer.
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Neoplasias dos Genitais Femininos/metabolismo , Óxido Nítrico/metabolismo , Animais , Membrana Celular/metabolismo , Feminino , Humanos , Concentração de Íons de Hidrogênio , Modelos BiológicosRESUMO
Reduced adenosine uptake via human equilibrative nucleoside transporter 1 (hENT1) in human umbilical vein endothelial cells (HUVECs) from gestational diabetes mellitus (GDM) is reversed by insulin by restoring hENT1 expression. Insulin receptors A (IR-A) and B (IR-B) are expressed in HUVECs, and GDM results in higher IR-A mRNA expression vs. cells from normal pregnancies. We studied whether the reversal of GDM effects on transport by insulin depends on restoration of IR-A expression. We specifically measured hENT1 expression [mRNA, protein abundance, SLC29A1 (for hENT1) promoter activity] and activity (adenosine transport kinetics) and the role of IR-A/IR-B expression and signaling [total and phosphorylated 42 and 44 kDa mitogen-activated protein kinases (p44/42(mapk)) and Akt] in IR-A, IR-B, and IR-A/B knockdown HUVECs from normal (n = 33) or GDM (n = 33) pregnancies. GDM increases IR-A/IR-B mRNA expression (1.8-fold) and p44/42(mapk):Akt activity (2.7-fold) ratios. Insulin reversed GDM-reduced hENT1 expression and maximal transport capacity (V(max)/K(m)), and GDM-increased IR-A/IR-B mRNA expression and p44/42(mapk):Akt activity ratios to values in normal pregnancies. Insulin's effect was abolished in IR-A or IR-A/B knockdown cells. Thus, insulin requires normal IR-A expression and p44/42(mapk)/Akt signaling to restore GDM-reduced hENT1 expression and activity in HUVECs. This could be a protective mechanism for the placental macrovascular endothelial dysfunction seen in GDM.
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Adenosina/metabolismo , Antígenos CD/metabolismo , Diabetes Gestacional/metabolismo , Insulina/metabolismo , Receptor de Insulina/metabolismo , Adolescente , Adulto , Antígenos CD/genética , Transporte Biológico Ativo , Estudos de Casos e Controles , Diabetes Gestacional/genética , Transportador Equilibrativo 1 de Nucleosídeo/genética , Transportador Equilibrativo 1 de Nucleosídeo/metabolismo , Feminino , Expressão Gênica , Técnicas de Silenciamento de Genes , Células Endoteliais da Veia Umbilical Humana , Humanos , Recém-Nascido , Cinética , Sistema de Sinalização das MAP Quinases , Masculino , Gravidez , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor de Insulina/antagonistas & inibidores , Receptor de Insulina/genética , Transdução de Sinais , Veias Umbilicais/metabolismo , Adulto JovemRESUMO
The alpha2-adrenergic receptor agonist Dexmedetomidine (Dex) is a sedative medication used by anesthesiologists. Dex protects the heart against ischemia-reperfusion (IR) and can also act as a preconditioning mimetic. The mechanisms involved in Dex-dependent cardiac preconditioning, and whether this action occurs directly or indirectly on cardiomyocytes, still remain unclear. The endothelial nitric oxide synthase (eNOS)/nitric oxide (NO) signaling pathway and endothelial cells are known to play key roles in cardioprotection against IR injury. Therefore, the aims of this work were to evaluate whether the eNOS/NO pathway mediates the pharmacological cardiac effect of Dex, and whether endothelial cells are required in this cardioprotective action. Isolated adult rat hearts were treated with Dex (10nM) for 25min and the dimerization of eNOS and production of NO were measured. Hearts were then subjected to global IR (30/120min) and the role of the eNOS/NO pathway was evaluated. Dex promoted the activation of eNOS and production of NO. Dex reduced the infarct size and improved the left ventricle function recovery, but this effect was reversed when Dex was co-administered with inhibitors of the eNOS/NO/PKG pathway. In addition, Dex was unable to reduce cell death in isolated adult rat cardiomyocytes subjected to simulated IR. Cardiomyocyte death was attenuated by co-culturing them with endothelial cells pre-treated with Dex. In summary, our results show that Dex triggers cardiac protection by activating the eNOS/NO signaling pathway. This pharmacological effect of Dex requires its interaction with the endothelium.
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Agonistas de Receptores Adrenérgicos alfa 2/farmacologia , Cardiotônicos/farmacologia , Dexmedetomidina/farmacologia , Traumatismo por Reperfusão Miocárdica/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Óxido Nítrico/metabolismo , Agonistas de Receptores Adrenérgicos alfa 2/uso terapêutico , Animais , Cardiotônicos/uso terapêutico , Células Cultivadas , Técnicas de Cocultura , Dexmedetomidina/uso terapêutico , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Coração/efeitos dos fármacos , Coração/fisiopatologia , Humanos , Masculino , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Miocárdio/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Ratos Sprague-DawleyRESUMO
Gestational diabetes mellitus (GDM) associates with increased L-arginine transport and extracellular concentration of adenosine in human umbilical vein endothelial cells (HUVECs). In this study we aim to determine whether insulin reverses GDM-increased L-arginine transport requiring adenosine receptors expression in HUVECs. Primary cultured HUVECs from full-term normal (n = 38) and diet-treated GDM (n = 38) pregnancies were used. Insulin effect was assayed on human cationic amino acid transporter 1 (hCAT1) expression (protein, mRNA, SLC7A1 promoter activity) and activity (initial rates of L-arginine transport) in the absence or presence of adenosine receptors agonists or antagonists. A1 adenosine receptors (A1AR) and A2AAR expression (Western blot, quantitative PCR) was determined. Experiments were done in cells expressing or siRNA-suppressed expression of A1AR or A2AAR. HUVECs from GDM exhibit higher maximal transport capacity (maximal velocity (V max)/apparent Michaelis Menten constant (K m), V max/K m), which is blocked by insulin by reducing the V max to values in cells from normal pregnancies. Insulin also reversed the GDM-associated increase in hCAT-1 protein abundance and mRNA expression, and SLC7A1 promoter activity for the fragment -606 bp from the transcription start point. Insulin effects required A1AR, but not A2AAR expression and activity in this cell type. In the absence of insulin, GDM-increased hCAT-1 expression and activity required A2AAR expression and activity. HUVECs from GDM pregnancies exhibit a differential requirement of A1AR or A2AAR depending on the level of insulin, a phenomenon that represent a condition where adenosine or analogues of this nucleoside could be acting as helpers of insulin biological effects in GDM.
Assuntos
Arginina/metabolismo , Diabetes Gestacional/tratamento farmacológico , Diabetes Gestacional/metabolismo , Endotélio Vascular/metabolismo , Hipoglicemiantes/farmacologia , Insulina/farmacologia , Receptor A1 de Adenosina/metabolismo , Veias Umbilicais/metabolismo , Adolescente , Adulto , Transportador 1 de Aminoácidos Catiônicos/biossíntese , Transportador 1 de Aminoácidos Catiônicos/genética , Diabetes Gestacional/dietoterapia , Endotélio Vascular/efeitos dos fármacos , Transportador Equilibrativo 1 de Nucleosídeo/metabolismo , Feminino , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Recém-Nascido , Masculino , Gravidez , Cultura Primária de Células , Receptor A1 de Adenosina/efeitos dos fármacos , Veias Umbilicais/efeitos dos fármacos , Adulto JovemRESUMO
NEW FINDINGS: What is the topic of this review? This review focuses on the effects of insulin therapy on fetoplacental vasculature in gestational diabetes mellitus and the potentiating effects of adenosine on this therapy. What advances does it highlight? This review highlights recent studies exploring a potential functional link between insulin receptors and their dependence on adenosine receptor activation (insulin-adenosine axis) to restore placental endothelial function in gestational diabetes mellitus. Gestational diabetes mellitus (GDM) is a disease that occurs during pregnancy and is associated with maternal and fetal hyperglycaemia. Women with GDM are treated via diet to control their glycaemia; however, a proportion of these patients do not achieve the recommended values of glycaemia and are subjected to insulin therapy until delivery. Even if a diet-treated GDM pregnancy leads to normal maternal and newborn glucose levels, fetoplacental vascular dysfunction remains evident. Thus, control of glycaemia via diet does not prevent GDM-associated fetoplacental vascular and metabolic alterations. We review the available information regarding insulin therapy in the context of its potential consequences for fetoplacental vascular function in GDM. We propose the possibility that insulin therapy to produce normoglycaemia in the mother and newborn may require additional therapeutic measures to restore the normal metabolic condition of the vascular network in GDM. A role for A1 and A2A adenosine receptors and insulin receptors A and B as well as a potential functional link in the cell signalling associated with the activation of these receptors is proposed. This possibility could be helpful for the planning of strategies, including adenosine receptor-improved insulin therapy, for the treatment of GDM patients, thereby promoting the wellbeing of the growing fetus, newborn and mother.
Assuntos
Diabetes Gestacional/tratamento farmacológico , Diabetes Gestacional/fisiopatologia , Insulina/uso terapêutico , Circulação Placentária/fisiologia , Feminino , Humanos , Recém-Nascido , Testes de Função Placentária/métodos , GravidezRESUMO
Microvascular and macrovascular endothelial function maintains vascular reactivity. Several diseases alter endothelial function, including hypertension, obesity, and diabetes mellitus. In addition, micro- and macrovascular endothelial dysfunction is documented in GDM with serious consequences for the growing fetus. Increased l-arginine uptake via hCAT-1 and NO synthesis by eNOS is associated with GDM. These alterations are paralleled by activation of purinergic receptors and increased umbilical vein, but not arteries blood adenosine accumulation. GDM associates with NO-reduced adenosine uptake in placental endothelium, suggested to maintain and/or facilitate insulin vasodilation likely increasing hCAT-1 and eNOS expression and activity. It is proposed that increased umbilical vein blood adenosine concentration in GDM reflects a defective metabolic state of human placenta. In addition, insulin recovers GDM-alterations in hCAT-1 and eNOS in human micro- and macrovascular endothelium, and its biological actions depend on preferential activation of insulin receptors A and B restoring a normal-like from a GDM-like phenotype. We summarized existing evidence for a potential role of insulin/adenosine/micro- and macrovascular endothelial dysfunction in GDM. These mechanisms could be crucial for a better management of the mother, fetus and newborn in GDM pregnancies.
Assuntos
Adenosina/metabolismo , Diabetes Gestacional , Células Endoteliais , Insulina/metabolismo , Placenta , Arginina/metabolismo , Diabetes Gestacional/metabolismo , Diabetes Gestacional/patologia , Diabetes Gestacional/fisiopatologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Feminino , Humanos , Óxido Nítrico/biossíntese , Óxido Nítrico Sintase Tipo III/biossíntese , Placenta/irrigação sanguínea , Placenta/metabolismo , Placenta/patologia , Placenta/fisiopatologia , GravidezRESUMO
OBJECTIVE: Human pregnancy that courses with maternal supraphysiological hypercholesterolemia (MSPH) correlates with atherosclerotic lesions in fetal arteries. It is known that hypercholesterolemia associates with endothelial dysfunction in adults, a phenomenon where nitric oxide (NO) and arginase are involved. However, nothing is reported on potential alterations in the fetoplacental endothelial function in MSPH. The aim of this study was to determine whether MSPH alters fetal vascular reactivity via endothelial arginase/urea and L-arginine transport/NO signaling pathways. APPROACH AND RESULTS: Total cholesterol <280 mg/dL was considered as maternal physiological hypercholesterolemia (n=46 women) and ≥ 280 mg/dL as MSPH (n=28 women). Maternal but not fetal total cholesterol and low-density lipoprotein-cholesterol levels were elevated in MSPH. Umbilical veins were used for vascular reactivity assays (wire myography), and primary cultures of umbilical vein endothelial cells to determine arginase, endothelial NO synthase (eNOS), and human cationic amino acid transporter 1 and human cationic amino acid transporter 2A/B expression and activity. MSPH reduced calcitonine gene-related peptide-umbilical vein relaxation and increased intima/media ratio (histochemistry), as well as reduced eNOS activity (L-citrulline synthesis from L-arginine, eNOS phosphorylation/dephosphorylation), but increased arginase activity and arginase II protein abundance. Arginase inhibition increased eNOS activity and L-arginine transport capacity without altering human cationic amino acid transporter 1 or human cationic amino acid transporter 2A/B protein abundance in maternal physiological hypercholesterolemia and MSPH. CONCLUSIONS: MSPH is a pathophysiological condition altering umbilical vein reactivity because of fetal endothelial dysfunction associated with arginase and eNOS signaling imbalance. We speculate that elevated maternal circulating cholesterol is a factor leading to fetal endothelial dysfunction, which could have serious consequences to the growing fetus.
Assuntos
Arginase/metabolismo , Células Endoteliais da Veia Umbilical Humana/enzimologia , Hipercolesterolemia/enzimologia , Óxido Nítrico Sintase Tipo III/metabolismo , Complicações na Gravidez/enzimologia , Veias Umbilicais/enzimologia , Adulto , Transportador 1 de Aminoácidos Catiônicos/metabolismo , Transportador 2 de Aminoácidos Catiônicos/metabolismo , Células Cultivadas , Feminino , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Hipercolesterolemia/sangue , Hipercolesterolemia/patologia , Hipercolesterolemia/fisiopatologia , Cinética , Lipídeos/sangue , Óxido Nítrico/metabolismo , Gravidez , Complicações na Gravidez/sangue , Complicações na Gravidez/patologia , Complicações na Gravidez/fisiopatologia , Trimestres da Gravidez/metabolismo , Transdução de Sinais , Veias Umbilicais/patologia , Veias Umbilicais/fisiopatologia , Ureia/metabolismo , Adulto JovemRESUMO
OBJECTIVES: The aim of this study was to assess maternal dietary food intake patterns, anthropometric measures, and multiple biochemical markers in women with gestational diabetes mellitus and pregnancy-specific urinary incontinence and to explore whether antedating gestational diabetes mellitus environment affects the pregnancy-specific urinary incontinence development in a cohort of pregnant women with gestational diabetes mellitus and pregnancy-specific urinary incontinence. METHODS: Maternal dietary information and anthropometric measurements were collected. At 24 wk of gestation, with a fasting venipuncture sample, current blood samples for biochemical markers of hormones, vitamins, and minerals were analyzed. The groups were compared in terms of numerical variables using analysis of variance for independent samples followed by multiple comparisons. RESULTS: Of the 900 pregnant women with complete data, pregnant women in the gestational diabetes mellitus pregnancy-specific urinary incontinence group had higher body mass index during pregnancy, arm circumference, and triceps skinfold than the non-gestational diabetes mellitus continent and non-gestational diabetes mellitus pregnancy-specific urinary incontinence groups, characterizing an obesogenic maternal environment. Regarding dietary food intake, significant increases in aromatic amino acids, branched-chain amino acids, dietary fiber, magnesium, zinc, and water were observed in pregnancy-specific urinary incontinence group compared with the non-gestational diabetes mellitus continent group. Serum vitamin C was reduced in the gestational diabetes mellitus pregnancy-specific urinary incontinence group compared with the non-gestational diabetes mellitus pregnancy-specific urinary incontinence group. CONCLUSIONS: This study emphasizes the necessity for a comprehensive strategy for gestational diabetes mellitus women with pregnancy-specific urinary incontinence in terms of deviation in maternal adaptation trending toward obesity and maternal micronutrients deficiencies.