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Background Coronavirus disease 2019 (COVID‐19) ranges from asymptomatic disease to respiratory failure and requires invasive mechanical ventilation (IMV). Data about the sequelae after infection are scarce. The study aims to describe the prevalence of symptoms, pulmonary function tests (PFTs), and radiological changes after four months of follow‐up. Methods A prospective, cross‐sectional, multicentre study was performed. Patients with different illness severities were consecutively included (mild; moderate: hospitalized without IMV; severe: hospitalized with IMV). Clinical variables, health‐related quality of life (HRQoL), PFT (spirometry, diffusing capacity of the lungs for carbon monoxide (DLCO)), and (CT) scans of the chest were obtained. The association between the risk of sequelae (DLCO

BackgroundRobust biomarkers that predict disease outcomes amongst COVID-19 patients are necessary for both patient triage and resource prioritisation. Numerous candidate biomarkers have been proposed for COVID-19. However, at present, there is no consensus on the best diagnostic approach to predict outcomes in infected patients. Moreover, it is not clear whether such tools would apply to other potentially pandemic pathogens and therefore of use as stockpile for future pandemic preparedness.MethodsWe conducted a multi-cohort observational study to investigate the biology and the prognostic role of interferon alpha-inducible protein 27 (IFI27) in COVID-19 patients.FindingsWe show that IFI27 is expressed in the respiratory tract of COVID-19 patients and elevated IFI27 expression is associated with the presence of a high viral load. We further demonstrate that systemic host response, as measured by blood IFI27 expression, is associated with COVID-19 severity. For clinical outcome prediction (e.g. respiratory failure), IFI27 expression displays a high positive (0.83) and negative (0.95) predictive value, outperforming all other known predictors of COVID-19 severity. Furthermore, IFI27 is upregulated in the blood of infected patients in response to other respiratory viruses. For example, in the pandemic H1N1/09 swine influenza virus infection, IFI27-like genes were highly upregulated in the blood samples of severely infected patients.InterpretationThese data suggest that prognostic biomarkers targeting the family of IFI27 genes could potentially supplement conventional diagnostic tools in future virus pandemics, independent of whether such pandemics are caused by a coronavirus, an influenza virus or another as yet-to-be discovered respiratory virus.Research in contextEvidence before this studyWe searched the scientific literature using PubMed to identify studies that used the IFI27 biomarker to predict outcomes in COVID-19 patients. We used the search terms “IFI27”, “COVID-19, “gene expression” and “outcome prediction”. We did not identify any study that investigated the role of IFI27 biomarker in outcome prediction. Although ten studies were identified using the general terms of “gene expression” and “COVID-19”, IFI27 was only mentioned in passing as one of the identified genes. All these studies addressed the broader question of the host response to COVID-19; none focused solely on using IFI27 to improve the risk stratification of infected patients in a pandemic.Added value of this studyHere, we present the findings of a multi-cohort study of the IFI27 biomarker in COVID-19 patients. Our findings show that the host response, as reflected by blood IFI27 gene expression, accurately predicts COVID-19 disease progression (positive and negative predictive values; 0.83 and 0.95, respectively), outperforming age, comorbidity, C-reactive protein and all other known risk factors. The strong association of IFI27 with disease severity occurs not only in SARS-CoV-2 infection, but also in other respiratory viruses with pandemic potential, such as the influenza virus. These findings suggest that host response biomarkers, such as IFI27, could help identify high-risk COVID-19 patients - those who are more likely to develop infection complications - and therefore may help improve patient triage in a pandemic.Implications of all the available evidenceThis is the first systemic study of the clinical role of IFI27 in the current COVID-19 pandemic and its possible future application in other respiratory virus pandemics. The findings not only could help improve the current management of COVID-19 patients but may also improve future pandemic preparedness.

Background: The maternal environment influences embryonic and fetal life. Nutritional deficits or excesses alter the trajectory of fetus/offspring’s development. The concept of “developmental programming” and “developmental origins of health and disease” consists of the idea that maternal diet may remodel the genome and lead to epigenetic changes. These changes are induced during early life, permanently altering the phenotype in the posterior adult stage, favoring the development of metabolic diseases such as obesity, dyslipidemia, hypertension, hyperinsulinemia, and metabolic syndrome. In this review, it is aimed to overview epigenetics, maternal diet and metabolic programming factors and determine which of these might affect future generations. Scope and Approach: Nutrients interfere with the epigenome by influencing the supply and use of methyl groups through DNA transmethylation and demethylation mechanisms. They also influence the remodeling of chromatin and arginine or lysine residues at the N-terminal tails of histone, thus altering miRNA expression. Fats, proteins, B vitamins and folates act as important cofactors in methylation processes. The metabolism of carbon in the methyl groups of choline, folic acid and methionine to S-Adenosyl Methionine (SAM), acts as methyl donors to methyl DNA, RNA, and proteins. B-complex vitamins are important since they act as coenzymes during this process. Key Findings and Conclusion: Nutrients, during pregnancy, potentially influence susceptibility to diseases in adulthood. Additionally, the deficit or excess of nutrients alter the epigenetic machinery, affecting genes and influencing the genome of the offspring and therefore, predisposing the development of chronic diseases in adults.

Dietary supplements have become the key to complement deficiencies in the occidental diet and therefore to reduce the incidence of oxidative stress related diseases. A bench-scale extraction procedure was studied to obtain a valuable product rich in phenolic compounds and antioxidant capacity from Pinot Noir grape cane enhanced by storage. Extraction solvent, cane-size, solid:liquid ratio, temperature, and extraction time, were systematically evaluated in order to obtain a natural functional product. Complete chemical characterization of a Pinot Noir grape cane extract produced under bench scale process is presented for the first time. Phenolic profiles of the extracts were characterized by HPLC-PDA-MS/MS and minerals by ICP-OES. Proteins, carbohydrates and lignins were also evaluated. The main phenolic compounds in the final product were stilbenoids, flavan-3-ols, procyanidins, and flavonols, with 6.53%, 4.84%, 2.11%, and 0.25%, respectively on a dry matter basis. Other chemical constituents were carbohydrates (27%), minerals (1%) and lignins (38.7%). The antioxidant capacity of the product was demonstrated using chemical assays (TEACABTS/CUPRAC and ORAC-FL) and endothelial cells model. The extract produced under the described bench scale process using grape cane enhanced by storage have a chemical composition and protecting capacities to be used in functional foods industry.

Background: In January 2021, the city of Concepcion in Chile suffered a second wave of COVID-19, while in early April 2021, all of Chile was facing the same situation. This generated the need to modify and validate a methodology for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in saliva, thereby expanding the capacity and versatility of testing. Methods: People who came to the health center in Concepcion city to perform a test of real-time reverse transcription polymerase chain reaction (RT-PCR) from a nasopharyngeal swab (NPS) specimen were invited to participate in this study. A total of 131 participants agreed to sign an informed consent and provide saliva and NPS specimens to validate a methodology in terms of sensitivity, specificity, and statistical analysis of the Ct values from RT-PCR. Findings: Calculations pertaining to the 127 participants who were ultimately included in the analysis were the following: sensitivity at 94·34% (95% CI: 84·34%-98·82%) and specificity at 98·65% (95% CI: 92·70%-99·97%). The saliva specimen showed a very similar performance to NPS as demonstrated with the diagnostic parameters. Interpretations: This RT-PCR methodology from the saliva specimen is a highly sensitive and specific alternative as compared to the reference methodology, which uses an NPS specimen. This modified and validated methodology is intended for use in the in vitro diagnosis of SARS-CoV-2, which provides health authorities in Chile and local laboratories with a real alternative for RT-PCR from NPS. Funding Information: Health Public Institute of Chile. Declaration of Interests: All authors declare no competing interests. Ethics Approval Statement: The study had the authorization of the Scientific Ethics Committee of the Health Service of Concepcion, Chile Number 20-01-02. Parents or legal guardians for volunteers under the age of 18 signed the informed consent.

Introduction Patients with severe COVID-19 develops an acute respiratory distress syndrome (ARDS), requiring admission to the intensive care unit. COVID-19 also reports an increased prevalence of comorbidities, similar to patients with Sleep disorder breathing (SDB). Objectives To evaluate the association between undiagnosed SDB and the risk of ARDS and pulmonary abnormalities in a cohort of patients’ survivors of COVID-19 between 3 and 6 months after diagnosis. Methods Prospective cohort study of patients who developed ARDS during hospitalization due to COVID-19 compared with a control group of patients who had COVID-19 with mild to moderate symptoms. All patients were evaluated between the 12th and 24th week after SARS-CoV-2 infection. The evaluation includes persistent symptoms, lung diffusing capacity of carbon monoxide (DLCO), chest CT scan and home sleep apnea test. SDB was diagnosed by the respiratory disturbance index ≥5 ev/h. The association between SDB and ARDS, the hazards of lung impairment and the hazard ratios (HR) were analyzed. Results A total of 60 patients were included (ARDS: 34 patients, Control: 26 patients). The mean follow-up was 16 weeks (range 12–24). ARDS reported a high prevalence of SDB (79% vs. 38% in control group). A total of 35% reported DLCO impairment, and 67.6% abnormal chest CT. SDB was independently associated to ARDS, OR 6.72 (CI, 1.56–28.93), p, Graphical abstract Image 1

Obstructive sleep apnea syndrome (OSAS) is a prevalent condition caused by dynamic upper airway collapse during sleep. The pathological impact and consequences are due to chronic intermittent hypoxia (CIH). Hypoxia increases the expression of several inflammatory stress markers and endothelial dysfunction. Recent studies suggest that patients with a similar AHI but with severe nocturnal hypoxia using oximetric parameters, such as the lowest saturation of oxygen during the night (min SaO2), percentage of total sleep time with oxygen saturation < 90% (T90) or the oxygen desaturation index (ODI-3%), commonly reported during the sleep study, are indicative of the increased expression of inflammatory markers due to severe nocturnal hypoxia and CIH during the night compared to subjects with moderate-severe OSAS without severe nocturnal hypoxia. The aim of this review is to describe physiological pathways involved in OSAS and their clinical consequences, focused in CIH and oximetric parameters showed in sleep study and their potential utility as inflammatory markers.

Rheumatoid arthritis (RA) has been associated with a higher risk of developing cardiovascular (CV) diseases. It has been proposed that systemic inflammation plays a key role in premature atherosclerosis development, and is therefore crucial to determine whether systemic components from RA patients promotes endothelial cell-oxidative stress by affecting reactive oxygen species (ROS) and nitric-oxide (NO) production. The aim of this study was to evaluate whether plasma from RA patients impair NO synthesis and ROS production by using the cell-line ECV-304 as a biosensor. NO synthesis and ROS production were measured in cells incubated with plasma from 73 RA patients and 52 healthy volunteers by fluorimetry. In addition, traditional CV risk factors, inflammatory molecules and disease activity parameters were measured. Cells incubated with plasma from RA patients exhibited reduced NO synthesis and increased ROS production compared to healthy volunteers. Furthermore, the imbalance between NO synthesis and ROS generation in RA patients was not associated with traditional CV risk factors. Our data suggest that ECV-304 cells can be used as a biosensor of systemic inflammation-induced endothelial cell-oxidative stress. We propose that both NO and ROS production are potential biomarkers aimed at improving the current assessment of CV risk in RA.

A new method for the analysis of phospholipids (Lysophosphatidylethanolamine, Phosphatidylethanolamine, Phosphatidylcholine, Sphingomyelin, and Lysophosphatidylcholine) by NP/HILIC-ELSD is described using a silica column. Addition of formic acid and triethylamine to a mobile phase based on acetonitrile/methanol/water delivered isocratically promoted a good and rapid separation of 5 phospholipid classes (15 min run). The use of an evaporative light scattering detector allowed the direct analysis of a mixture of phospholipids without complex sample pretreatment. Both power and linear regression showed satisfactory concordance with the relationship between response and concentration. The LOD and LOQ obtained were below 10 ng and 30 ng of injected mass for all cases, respectively. The method was validated with a standard mixture of phospholipids. This method has been applied to primarily characterize the phospholipid fraction of both native and oxidized low density lipoproteins.

Nitric oxide (NO) is an endogenous vasodilator molecule synthetized from L-arginine by a family of nitric oxide synthases. In differentiated human endothelial cells, it is well known that L-arginine uptake via cationic amino acid transporters (y(+)/CAT) or system y(+)L is required for the NO synthesis via endothelial nitric oxide synthase, but there are no reports in human endothelial progenitor cell (hEPC). Therefore, we isolated hEPCs from peripheral blood of healthy donors and cultured them for either 3 (hEPC-3d) or 14 days (hEPC-14d) to characterize the L-arginine transport and NO synthesis in those cells. L-arginine transport and NO synthesis were analyzed in the presence or absence of N-ethylmaleimide or L-nitroarginine methyl ester, as inhibitors of y(+)/CAT system and nitric oxide synthases, respectively. The results showed that L-arginine uptake is higher in hEPC-14d than in hEPC-3d. Kinetic parameters for L-arginine transport showed the existence of at least 2 transporter systems in hEPC: a high affinity transporter system (K(m)= 4.8 ± 1.1 μM for hEPC-3d and 6.1 ± 2.4 μM for hEPC-14d) and a medium affinity transporter system (K(m) = 85.1 ± 4.0 μM for hEPC-3d and 95.1 ± 8 μM for hEPC-14d). Accordingly, hEPC expressed mRNA and protein for CAT-1 (ie, system y(+)) and mRNA for 2 subunits of y(+)L system, yLAT1, and 4F2hc. Higher L-citruline production and NO bioavailability (4-fold), and endothelial nitric oxide synthase expression (both mRNA and protein) were observed in hEPC-14d compared with hEPC-3d. Finally, the high L-citruline formation observed in hEPC-14d was blocked by N-ethylmaleimide. In conclusion, this study allowed to identity a functional L-arginine/NO pathway in two hEPC differentiation stages, which improves the understanding of the physiology of these precursor cells.

Human endothelial progenitor cells (hEPC) are recruited to sites of neovascularization where they differentiate into endothelial cells. The signals/factors responsible for hEPC migration and adhesion to sites of injury are not well understood. Elevated levels of adenosine are known to incre

Antecedentes y Objetivos: Las mujeres diabéticas tienen mayor morbi-mortalidad asociada a eventos coronarios isquémicos que los hombres. Este estudio describe la extensión, magnitud y severidad de la ateros-clerosis coronaria entre hombres y mujeres con diabetes mellitus tipo 2. Métodos: Estudio de tipo descriptivo que incluyó a 162 pacientes diabéticos consecutivos con sospecha de aterosclerosis coronaria y estudiados con coronariogra-fía. La magnitud de la aterosclerosis fue cuantificada utilizando el Score de Gensini. Resultados: La edad promedio fue 64.8 años. Noventa y cuatro eran mujeres (58.0%). El tiempo de evolución de la diabetes fue mayor para mujeres que para hombres (152.1 ± 90.0 vs 120.2 ± 99.4 meses) respectivamente (p

An elevated extracellular concentration of D-glucose (i.e. hyperglycaemia) inhibits cell proliferation and incorporation of the endogenous nucleoside thymidine into DNA in human umbilical vein endothelial cells (HUVECs). Cells in their log-phase of growth (3.7 ± 0.3 days, n = 27) incubated for 30 min with 25 mM D-glucose, but not with equimolar concentrations of L-glucose or D-mannitol, exhibited reduced [3H]thymidine incorporation and cell growth rate, with no change in cell viability (> 98 %), total DNA, protein content or cell volume. Incubation with D-glucose activated protein kinase C (PKC), endothelial NO synthase (eNOS), p42 and p44 mitogen-activated protein kinases (p42/44mapk), but inhibited superoxide dismutase (SOD). Incubation with D-glucose also increased cGMP and cAMP levels. The effect of D-glucose was blocked by the PKC inhibitor calphostin C, the MAP kinase kinase 1/2 (MEK1/2) inhibitor PD-98059, the eNOS inhibitor L-NAME, the protein kinase G (PKG) inhibitor KT-5823 and the protein kinase A (PKA) inhibitor KT-5720. In the presence of 5 mM D-glucose, [3H]thymidine incorporation and cell growth were reduced by the PKC activator phorbol 12-myristate 13-acetate (PMA), the NO donor S-nitroso-N-acetyl-L,D-penicillamine (SNAP), dibutyryl cGMP, dibutyryl cAMP and the Ca2+ ionophore A-23187. The effect of A-23187 was blocked by calphostin C and PD-98059. D-Glucose-dependent inhibition of thymidine incorporation and cell proliferation is associated with increased PKC, eNOS, and MEK1/2, but decreased SOD activity, and higher intracellular levels of cGMP, cAMP and Ca2+ in HUVECs. These are cellular mechanisms which may reduce endothelial cell growth in pathological conditions such as in diabetes mellitus or hyperglycaemia. Experimental Physiology (2003) 88.2, 209-219.

Altered expression and function of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) has been associated with several diseases such as endothelial dysfunction, atherosclerosis and obesity. In these pathologies, oxLDL/LOX-1 activates signaling pathways that promote cell proliferation, cell motility and angiogenesis. Recent studies have indicated that olr1 mRNA is over-expressed in stage III and IV of human prostatic adenocarcinomas. However, the function of LOX-1 in prostate cancer angiogenesis remains to be determined. Our aim was to analyze the contribution of oxLDL and LOX-1 to tumor angiogenesis using C4-2 prostate cancer cells. We analyzed the expression of pro-angiogenic molecules and angiogenesis on prostate cancer tumor xenografts, using prostate cancer cell models with overexpression or knockdown of LOX-1 receptor. Our results demonstrate that the activation of LOX-1 using oxLDL increases cell proliferation, and the expression of the pro-angiogenic molecules VEGF, MMP-2, and MMP-9 in a dose-dependent manner. Noticeably, these effects were prevented in the C4-2 prostate cancer model when LOX-1 expression was knocked down. The angiogenic effect of LOX-1 activated with oxLDL was further demonstrated using the aortic ring assay and the xenograft model of tumor growth on chorioallantoic membrane of chicken embryos. Consequently, we propose that LOX-1 activation by oxLDL is an important event that enhances tumor angiogenesis in human prostate cancer cells.

The bioavailability of nitric oxide (NO) represents a key marker in vascular health. A decrease in NO induces a pathological condition denominated endothelial dysfunction, syndrome observed in different pathologies, such as obesity, diabetes, kidney disease, cardiovascular disease, and preeclampsia (PE). PE is one of the major risks for maternal death and fetal loss. Recent studies suggest that the placenta of pregnant women with PE express high levels of lectin-like oxidized LDL receptor-1 (LOX-1), which induces endothelial dysfunction by increasing reactive oxygen species (ROS) and decreasing intracellular NO. Besides LOX-1 activation induces changes in migration and apoptosis of syncytiotrophoblast cells. However, the role of this receptor in placental tissue is still unknown. In this review we will describes the physiological roles of LOX-1 in normal placenta development and the potential involvement of this receptor in the pathophysiology of PE.

Background: Mesenchymal stem cells have a high capacity for trans-differentiation toward many adult cell types, including endothelial cells. Feto-placental tissue, such as Wharton’s jelly is a potential source of mesenchymal stem cells with low immunogenic capacity; make them an excellent source of progenitor cells with a potential use for tissue repair. We evaluated whether administration of endothelial cells derived from mesenchymal stem cells isolated from Wharton’s jelly (hWMSCs) can accelerate tissue repair in vivo. Methods: Mesenchymal stem cells were isolated from human Wharton’s jelly by digestion with collagenase type I. Endothelial trans-differentiation was induced for 14 (hWMSC-End14d) and 30 (hWMSC-End30d) days. Cell phenotyping was performed using mesenchymal (CD90, CD73, CD105) and endothelial (Tie-2, KDR, eNOS, ICAM-1) markers. Endothelial transdifferentiation was demonstrated by the expression of endothelial markers and their ability to synthesize nitric oxide (NO). Results: hWMSCs can be differentiated into adipocytes, osteocytes, chondrocytes and endothelial cells. Moreover, these cells show high expression of CD73, CD90 and CD105 but low expression of endothelial markers prior to differentiation. hWMSCs-End express high levels of endothelial markers at 14 and 30 days of culture, and also they can synthesize NO. Injection of hWMSC-End30d in a mouse model of skin injury significantly accelerated wound healing compared with animals injected with undifferentiated hWMSC or injected with vehicle alone. These effects were also observed in animals that received conditioned media from hWMSC-End30d cultures. Conclusion: These results demonstrate that mesenchymal stem cells isolated from Wharton’s jelly can be cultured in vitro and trans-differentiated into endothelial cells. Differentiated hWMSC-End may promote neovascularization and tissue repair in vivo through the secretion of soluble pro-angiogenic factors.

The aromatic herb Cymbopogon citratus Stapf is widely used in tropical and subtropical countries in cooking, as a herbal tea, and in traditional medicine for hypertension and diabetes. Some of its properties have been associated with the in vitro antioxidant effect of polyphenols isolated from their aerial parts. However, little is known about C. citratus effects on endothelial cells oxidative injury. Using chromatographic procedures, a polyphenol-rich fraction was obtained from C. citratus (CCF) and their antioxidant properties were assessed by cooper-induced LDL oxidation assay. The main constituents of the active CCF, identified by high-performance liquid chromatography with diode-array detection and mass spectrometry (HPLC-DAD-MS), were chlorogenic acid, isoorientin and swertiajaponin. CCF 10 and 100 μg/ml diminishes reactive oxidative species (ROS) production in human umbilical vein endothelial cell (HUVECs), challenged with high D-glucose (60% inhibition), hydrogen peroxide (80% inhibition) or oxidised low-density lipoprotein (55% inhibition). CCF 10 or 100 μg/ml did not change nitric oxide (NO) production. However, CCF was able to inhibit vasoconstriction induced by the thromboxane A2 receptor agonist U46619, which suggest a NO-independent vasodilatador effect on blood vessels. Our results suggest that lemon grass antioxidant properties might prevent endothelial dysfunction associated to an oxidative imbalance promoted by different oxidative stimuli.

Human endothelial progenitor cells (hEPC) are recruited to sites of neovascularization where they differentiate into endothelial cells. The signals/factors responsible for hEPC migration and adhesion to sites of injury are not well understood. Elevated levels of adenosine are known to increase mature endothelial cell migration in response to tissue injury. However, the understanding of the role of adenosine in the physiology of hEPC is very limited. Using quantitative polymerase chain reaction and western blot analyses, we detected the expression of the adenosine receptors A₂A, A₂B, and A₃ in hEPC. Stimulation of adenosine receptors using adenosine or the nonselective agonist adenosine-5'-N-ethylcarboxamide (NECA) increased hEPC migration in 1.4-fold and 2.1-fold (P0.01), respectively. Stimulation of hEPC using the A₂A-specific agonist CGS-21680 resembled the effect observed in migration when using adenosine or NECA. Consequently, NECA and CGS-21680-stimulated migration of hEPC were reverted using the A₂A receptor antagonist ZM-241385. NECA-stimulated migration was inhibited in dose-dependent manner using MRS-1523 (Ki of 147 ± 0.016 nM), MRS-1754 (Ki of 1900 ± 0.02 nM), or ZM-241385 (Ki of 0.2 ± 0.01 nM). In conclusion, adenosine stimulates hEPC migration by activating A₂A and A₃ but not A₂B receptors and provides evidence to support a role of adenosine in modulating angiogenic capacity of hEPC.

Objective: To evaluate in vitro the effects of serum and LDL fractions isolated from hypercholesterolemic patients on nitric oxide (NO) synthesis and reactive oxygen species (ROS) production by human umbilical vein endothelial cells (HUVECs). Design and methods: Serum and LDL isolated from subjects with high (n=18) and normal (n=21) LDLcholesterol levels were analyzed on NO synthesis and ROS production in vitro models of HUVECs. LDL was furthers characterized in their chemical composition and activities of lipoprotein-associated phospholipase A2 (Lp-PLA2), cholesteryl ester transfer protein (CETP) and paraoxonase. Results: NO bioavailability was significantly lower and ROS production higher in HUVECs incubated withserum samples from patients with high LDL-cholesterol levels in comparison to control subjects. Moreover, hypercholesterolemic patients presented higher CETP and Lp-PLA2 activities than control subjects. LDL fractions isolated from patients and controls were not different in their chemical composition, Lp-PLA2 activity, and their capacity to reduce NO synthesis and increase ROS production. Conclusion: Alterations of serum from hypercholesterolemic patients could be due to the increment in LDL concentration, main Lp-PLA2 carrier, and not to LDL composition or intrinsic Lp-PLA2 activity. Fil: Searle, Andrea. Universidad de Concepción; Chile Fil: Gomez Rosso, Leonardo Adrián. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Bioquímica Clínica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina Fil: Meroño, Tomás. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Bioquímica Clínica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina Fil: Salomon, Carlos. Universidad de Concepción; Chile Fil: Durán Sandoval, Daniel. Universidad de Concepción; Chile Fil: Giunta, Gustavo Ariel. Fundacion Favaloro; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Grant, Carlos. Universidad de Concepción; Chile Fil: Calvo, Carlos. Universidad de Concepción; Chile Fil: Lamperti, Liliana. Universidad de Concepción; Chile Fil: Brites, Fernando Daniel. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Bioquímica Clínica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; Argentina Fil: Aguayo, Claudio. Universidad de Concepción; Chile

To evaluate the association between endothelial activation markers in the maternal circulation with nitric oxide (NO) synthesis in human umbilical endothelial cells.This is a case-control study of normal and pre-eclamptic pregnancies. The levels of sE-selectin, soluble vascular cell adhesion molecule 1 (sVCAM-1), and soluble fms-like tyrosine kinase 1 (sFlt-1) were measured by enzyme-linked immunosorbent assay, and histamine-induced NO synthesis was detected by fluorometric examination of the human umbilical vein endothelial cells (HUVECs) isolated from normal and pathological pregnancies.Mothers with severe pre-eclamptic pregnancies have premature and smaller babies than mothers with normal pregnancies (P0.05); they also have high maternal plasma levels of sVCAM-1 (∼2-fold), sFlt-1 (∼2.5-fold), and lower (∼70%) histamine-stimulated NO synthesis in HUVECs. A positive relationship between systolic blood pressure (SBP) and plasma levels of sE-selectin, sVCAM-1, and sFlt-1 was demonstrated. Moreover, levels of sE-selectin, sVCAM-1, and sFlt-1 were negatively associated with newborn weight (NBW), gestational age at delivery, and NO synthesis. Women with high E-selectin (63 ng/ml), VCAM-1 (752 ng/ml), and sFlt-1 (15204 pg/ml) showed high risk (∼2-fold) for preterm delivery and very preterm delivery, or fetal weight1500 g (∼1.5-fold) compared with women with low levels.High circulating levels of maternal endothelial dysfunction markers present in pre-eclampsia are associated with decreased NO synthesis in fetal endothelium.

Extracellular adenosine removal is via human equilibrative nucleoside transporters 1 (hENT1) and 2 (hENT2) in the endothelium, thus regulating adenosine-induced revascularization and angiogenesis. Since human endothelial progenitor cells (hEPCs) promote revascularization, we hypothesize differential expression of nucleoside transporters in hEPCs. hEPCs were cultured 3 (hEPC-3d) or 14 (hEPC-14d) days. RT-PCR for prominin 1, CD34, octamer-4, kinase insert domain receptor, oxidized low-density lipoprotein (lectin-like) receptor 1 and tyrosine endothelial kinase was used to evaluate phenotypic differentiation. Flow cytometry was used to estimate CD34(+)/KDR(-) (non-differentiated), CD34(-)/KDR(+) (differentiated) or CD34(+)/KDR(+) (mixed) cell populations. Adenosine transport was measured in absence or presence of sodium, S-(4-nitrobenzyl)-6-thio-inosine (NBTI, 1-10 μM), inosine, hypoxanthine or guanine (0.1-5 mM), hENTs protein abundance by western blot, and hENTs, hCNT1, hCNT2 and hCNT3 mRNA expression by real time RT-PCR. hEPC-3d cells were CD34(+)/KDR(-) compared with hEPC-14d cells that were CD34(-)/KDR(+). hEPC-3d cells exhibit hENT1-like adenosine transport (NBTI-sensitive, Na(+)-independent), which is absent in hEPC-14d cells. hEPC-14d cells exhibit two transport components: component 1 (NBTI insensitive, Na(+)-independent) and component 2 (NBTI insensitive, Na(+)-dependent, Hill coefficient ∼1.8), the latter resembling CNT3-like transport. hEPC-3d cells express hENT1 protein and mRNA, which is reduced (∼90%) in hEPC-14d cells, but instead only hCNT3 mRNA is expressed in this cell type. hENT2, hCNT1 and hCNT2 were undetectable in hEPCs. Thus, hEPCs exhibit a differential expression of hENT1 and hCNT3 functional nucleoside transporters, which could be related with its differentiation stage.