Comparative analysis of antiproliferative and vasodilator effects of drugs for pulmonary hypertension: Extensive in vitro study in rats and human.

An effective pulmonary hypertension (PH) treatment should combine antiproliferative and vasodilator effects. We characterized a wide-range of drugs comparing their anti-proliferative vs vasodilator effects in human and rat pulmonary artery smooth muscle cells (PASMC). Key findings: 1) Approved PH drugs (PDE5 inhibitors, sGC stimulators and PGI2 agonists) are preferential vasodilators. 2) cGMP stimulators were more effective in cells derived from hypertensive rats. 3) Nifedipine acted equally as vasodilator and antiproliferative. 4) quercetin and imatinib were potent dual vasodilator/antiproliferative drugs. 5) Tacrolimus and levosimendan lacked antiproliferative effects. 6) Forskolin, pinacidil and hydroxyfasudil were more effective as antiproliferative in human cells.

SARS-CoV-2 Infection Triggers Auto-Immune Response in ARDS.

Acute respiratory distress syndrome (ARDS) is a severe pulmonary disease, which is one of the major complications in COVID-19 patients. Dysregulation of the immune system and imbalances in cytokine release and immune cell activation are involved in SARS-CoV-2 infection. Here, the inflammatory, antigen, and auto-immune profile of patients presenting COVID-19-associated severe ARDS has been analyzed using functional proteomics approaches. Both, innate and humoral responses have been characterized through acute-phase protein network and auto-antibody signature. Severity and sepsis by SARS-CoV-2 emerged to be correlated with auto-immune profiles of patients and define their clinical progression, which could provide novel perspectives in therapeutics development and biomarkers of COVID-19 patients. Humoral response in COVID-19 patients' profile separates with significant differences patients with or without ARDS. Furthermore, we found that this profile can be correlated with COVID-19 severity and results more common in elderly patients.

Carotid Plaque Inflammation Assessed by 18F-FDG PET/CT and Lp-PLA2 Is Higher in Symptomatic Patients.

Carotid plaque inflammation assessed by 2-deoxy-2-[18F]fluoro-D-glucose (18F-FDG) positron emission tomography/computed tomography (PET/CT) and lipoprotein-associated phospholipase A2 (Lp-PLA2) levels are higher in symptomatic patients. The aim of this study was to assess correlations between 18F-FDG uptake on PET scan of carotid artery plaques, plasma levels of Lp-PLA2, and cerebrovascular symptoms. The study included 45 consecutive patients (22 symptomatic, 23 asymptomatic) with >70% carotid stenosis. Patients were examined by hybrid PET/CT, and maximum standardized uptake values (SUVmax) were recorded. Blood samples were obtained, and plasma was stored at -80 °C for subsequent Lp-PLA2 analysis. Symptomatic and asymptomatic patients showed no significant difference in classical cardiovascular risk factors. Asymptomatic carotid stenosis patients more frequently had a history of coronary artery disease (P = .025) and peripheral artery disease (P = .012). The symptomatic group had higher 18F-FDG uptake in carotid plaques (P < .001), higher plasma Lp-PLA2 (P < .01), and higher high-sensitive C-reactive protein (P = .022). 2-Deoxy-2-[18F]fluoro-D-glucose uptake on PET/CT and plasma Lp-PLA2 show a statistically significant association with the symptomatic status of carotid plaques.

Liver-lung interactions in acute respiratory distress syndrome.

Patients with liver diseases are at high risk for the development of acute respiratory distress syndrome (ARDS). The liver is an important organ that regulates a complex network of mediators and modulates organ interactions during inflammatory disorders. Liver function is increasingly recognized as a critical determinant of the pathogenesis and resolution of ARDS, significantly influencing the prognosis of these patients. The liver plays a central role in the synthesis of proteins, metabolism of toxins and drugs, and in the modulation of immunity and host defense. However, the tools for assessing liver function are limited in the clinical setting, and patients with liver diseases are frequently excluded from clinical studies of ARDS. Therefore, the mechanisms by which the liver participates in the pathogenesis of acute lung injury are not totally understood. Several functions of the liver, including endotoxin and bacterial clearance, release and clearance of pro-inflammatory cytokines and eicosanoids, and synthesis of acute-phase proteins can modulate lung injury in the setting of sepsis and other severe inflammatory diseases. In this review, we summarized clinical and experimental support for the notion that the liver critically regulates systemic and pulmonary responses following inflammatory insults. Although promoting inflammation can be detrimental in the context of acute lung injury, the liver response to an inflammatory insult is also pro-defense and pro-survival. A better understanding of the liver-lung axis will provide valuable insights into new diagnostic targets and therapeutic strategies for clinical intervention in patients with or at risk for ARDS.

Fas activation alters tight junction proteins in acute lung injury.

Background:The acute respiratory distress syndrome (ARDS) is characterized by protein-rich oedema in the alveolar spaces, a feature in which Fas-mediated apoptosis of the alveolar epithelium has been involved. Objective:To determine whether Fas activation increases protein permeability by mechanisms involving disruption of the paracellular tight junction (TJ) proteins in the pulmonary alveoli. Methods: Protein permeability and the expression of TJ proteins were assessed in vivo in wild-type and Fas-deficient lpr mice 16 hours after the intratracheal instillation of recombinant human soluble Fas ligand (rh-sFasL), and at different time points in vitro in human pulmonary alveolar epithelial cells (HPAEpiC) exposed to rh-sFasL Results:Activation of the Fas pathway increased protein permeability in mouse lungs and altered the expression of the TJ proteins occludin and zonula occludens-1 in the alveolar-capillary membrane in vivo and in human alveolar epithelial cell monolayers in vitro. Blockade of caspase-3, but not inhibition of tyrosine kinase dependent pathways, prevented the alterations in TJ protein expression and permeability induced by the Fas/FasL system in human alveolar cell monolayers in vitro. We also observed that both the Fas-induced increase of protein permeability and disruption of TJ proteins occurred before cell death could be detected in the cell monolayers in vitro. Conclusion:Targeting caspase pathways could prevent the disruption of TJs and reduce the formation of lung oedema in the early stages of ARDS.

MicroRNAs as biomarkers of acute lung injury.

Acute respiratory distress syndrome (ARDS) is a common and complex inflammatory lung diseases affecting critically ill patients requiring mechanical ventilation. MicroRNAs (miRNAs), a novel pathway of non-coding RNA molecules that regulate gene expression at the post-transcriptional level, have emerged as a novel class of gene expression, and can play important roles in inflammation or apoptosis, which are common manifestations of ARDS and diffuse alveolar damage (DAD). In the present review, we discuss the role of miRNAs as biomarkers of ARDS and DAD, and their potential use as therapeutic targets for this condition.

Elevated levels of triglycerides and vldl-cholesterol provoke activation of nlrp1 inflammasome in endothelial cells.

BACKGROUND: Soluble stimuli present in the plasma of patients with peripheral arterial disease (PAD) are capable of directly stimulating intracellular signalling in endothelium. Oxidized-LDL (oxLDL) induces NLRP3 inflammasome activation in macrophages. However, it is not clear how lipid profile affect NLRP1 inflammasome gene expression in endothelial cells. In this study, the effect of cholesterol and TG of plasma of patients with PAD on NLRP1 inflammasome gene expression in human arterial endothelial cells (HAECS) was assessed. METHODS: We included 113 patients with symptomatic PAD. HAECs were stimulated for 2h using the plasma samples of the study participants. The NLRP1 quantification of the transcription was carried out on the 7500 real-time PCR system using the Taqman® Universal PCR Master Mix and Assays on demand. Relative quantification of the NLRP1 expression was carried out using the ΔΔCt (threshold cycle) comparative method. RESULTS: Plasma from patients with elevated VLDL-cholesterol levels (>33.6mg/dL, the median value of the sample) provoked a higher expression of NLRP1 inflammasome in HAECs (RQ=1.15±0.23 vs. 1.05±0.69; p=0.045), as well as plasma from patients with elevated TGs levels (>168mg/dL, the median value of the sample) (RQ=1.15±0.23 vs. 1.05±0.69; p=0.045). A positive correlation was found between NLRP1 inflammasome expression and VLDL-cholesterol plasma levels (r=0.4; p<0.001) as between NLRP1 inflammasome expression and TG levels (r=0.4; p<0.001). CONCLUSIONS: Plasma TG and VLDL cholesterol of patients with atherosclerosis, manifested as PAD, promote the in vitro NLRP1 inflammasome expression in HAECs.

Effect of Simvastatin Treatment on "In Vitro" NLRP1 Inflammasome Expression in Peripheral Arterial Disease.

BACKGROUND: Inflammatory stress stimuli in the plasma of patients with peripheral artery disease (PAD) are able to trigger the expression of NLRP1 inflammasome in human aortic endothelial cells (HAECs). Our objective was to elucidate the effect of simvastatin treatment on NLRP1 inflammasome expression in endothelial cells exposed to the plasma of PAD patients. METHODS: The study included 81 patients with PAD, 24 of them treated with simvastatin (20 mg/day) and 57 without statin therapy. HAECs between passages 3 and 6 were stimulated for 2 hr using the plasma samples of the study participants. NLRP1 gene transcription of HAECs exposed to the plasma of PAD patients was quantificated. RESULTS: HAECs exposed to the plasma of PAD patients with simvastatin therapy showed significantly higher expression of the NLRP1 gene compared with those exposed to the plasma of PAD patients without this treatment (relative quantitation [RQ] 1.12 ± 0.06 vs. 1.06 ± 0.07, P = 0.03). Furthermore, HAECs exposed to the plasma of patients with critical limb ischemia and treated with simvastatin responded with a higher NLRP1 expression than those exposed to the plasma of simvastatin-treated patients with claudication (RQ 1.1 ± 0.3 vs. 0.99 ± 0.14, P < 0.001). CONCLUSION: Simvastatin intake in PAD patients increases in vitro reactivity of NLRP1 inflammasome gene expression in HAECs, especially in critical limb ischemia patients.

Characteristics of microRNAs and their potential relevance for the diagnosis and therapy of the acute respiratory distress syndrome: from bench to bedside.

Acute respiratory distress syndrome (ARDS) is a complex disease associated with high morbidity and mortality. Biomarkers and specific pharmacologic treatment of the syndrome are lacking. MicroRNAs (miRNAs) are small (∼ 19-22 nucleotides) noncoding RNA molecules whose function is the regulation of gene expression. Their uncommon biochemical characteristics (eg, their resistance to degradation because of extreme temperature and pH fluctuations, freeze-thaw cycles, long storage times in frozen conditions, and RNAse digestion) and their presence in a wide range of different biological fluids and the relatively low number of individual miRNAs make these molecules good biomarkers in different clinical conditions. In addition, miRNAs are suitable therapeutic targets as their expression can be modulated by different available strategies. The aim of the present review is to offer clinicians a global perspective of miRNA, covering their structure and nomenclature, biogenesis, effects on gene expression, regulation of expression, and features as disease biomarkers and therapeutic targets, with special attention to ARDS. Because of the early stage of research on miRNAs applied to ARDS, attention has been focused on how knowledge sourced from basic and translational research could inspire future clinical studies.

Inhibition of Nitro-Oxidative Stress Attenuates Pulmonary and Systemic Injury Induced by High-Tidal Volume Mechanical Ventilation.

Mechanisms contributing to pulmonary and systemic injury induced by high tidal volume (VT) mechanical ventilation are not well known. We tested the hypothesis that increased peroxynitrite formation is involved in organ injury and dysfunction induced by mechanical ventilation. Male Sprague-Dawley rats were subject to low- (VT, 9 mL/kg; positive end-expiratory pressure, 5 cmH2O) or high- (VT, 25 mL/kg; positive end-expiratory pressure, 0 cmH2O) VT mechanical ventilation for 120 min, and received 1 of 3 treatments: 3-aminobenzamide (3-AB, 10 mg/kg, intravenous, a poly adenosine diphosphate ribose polymerase [PARP] inhibitor), or the metalloporphyrin manganese(III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP, 5 mg/kg intravenous, a peroxynitrite scavenger), or no treatment (control group), 30 min before starting the mechanical ventilation protocol (n = 8 per group, 6 treatment groups). We measured mean arterial pressure, peak inspiratory airway pressure, blood chemistry, and gas exchange. Oxidation (fluorescence for oxidized dihydroethidium), protein nitration (immunofluorescence and Western blot for 3-nitrotyrosine), PARP protein (Western blot) and gene expression of the nitric oxide (NO) synthase (NOS) isoforms (quantitative real-time reverse transcription polymerase chain reaction) were measured in lung and vascular tissue. Lung injury was quantified by light microscopy. High-VT mechanical ventilation was associated with hypotension, increased peak inspiratory airway pressure, worsened oxygenation; oxidation and protein nitration in lung and aortic tissue; increased PARP protein in lung; up-regulation of NOS isoforms in lung tissue; signs of diffuse alveolar damage at histological examination. Treatment with 3AB or MnTMPyP attenuated the high-VT mechanical ventilation-induced changes in pulmonary and cardiovascular function; down-regulated the expression of NOS1, NOS2, and NOS3; decreased oxidation and nitration in lung and aortic tissue; and attenuated histological changes. Increased peroxynitrite formation is involved in mechanical ventilation-induced pulmonary and vascular dysfunction.

Circulating anti-ß2-glycoprotein I antibodies of peripheral arterial disease patients trigger a genomic overexpression of Toll-like receptor 4 in endothelial cells.

OBJECTIVE: Circulating anti-ß2-glycoprotein I (ABGPI) antibodies are associated with peripheral arterial disease (PAD) and induce the expression of leukocyte adhesion molecules and proinflammatory cytokines by endothelial cells. Our aim is to study a transcriptional activation pathway of the innate immune system through the cellular signalling cascade triggered by receptors Toll-like receptor 2 (TLR2) and Toll-like receptor 4 (TLR4) of endothelial cells after the exposure of these cells to seropositive ABGPI human serum obtained from PAD patients. METHODS: We obtained serum samples from PAD patients and controls without PAD. ABGPI serum titer was detected using indirect immunofluorescence. Our sample was stratified into three groups: group I (PAD and ABGPI titer ≥1:100; n = 15), group II (PAD and ABGPI titer <1:100; n = 15), and control participants (no PAD; n = 15). All serum samples were incubated with human aortic endothelial cell (HAEC) culture. Genomic expression of TLR2 and TLR4 receptors and their shared intracellular signalling molecules, myeloid differentiation primary response gene 88 (MyD88), and interleukin (IL)-1 receptor-associated kinase (1IRAK1), were measured after the exposure of HAECs to each serum. RESULTS: HAEC genomic expression of TLR4 was higher after the exposure to group I serum than after the exposure to group II serum (log10×10-relative quantification [RQ]: 1.80 ± 0.42 vs 1.37 ± 0.39; P = .01) or control serum (log10×10-RQ: 1.80 ± 0.42 vs 1.09 ± 0.26; P < .01). TLR4 expression was higher in group II than in the control group (log10×10-RQ: 1.37 ± 0.39 vs 1.09 ± 0.26; P = .04). TLR4 expression correlated with MyD88 (r = 0.54; P < .01) and IRAK1 (r = 0.55; P < .01) expression. We recorded a positive correlation between MyD88 and IRAK1 genomic expression (r = 0.58; P < .01). CONCLUSIONS: Our results suggest that serum from PAD patients with elevated ABGPI antibodies induces a genomic overexpression of TLR4 and its cellular signalling molecules in endothelial cells.

Influence of mechanical ventilation and sepsis on redox balance in diaphragm, myocardium, limb muscles, and lungs.

Mechanical ventilation (MV), using high tidal volumes (V(T)), causes lung (ventilator-induced lung injury [VILI]) and distant organ injury. Additionally, sepsis is characterized by increased oxidative stress. We tested whether MV is associated with enhanced oxidative stress in sepsis, the commonest underlying condition in clinical acute lung injury. Protein carbonylation and nitration, antioxidants, and inflammation (immunoblotting) were evaluated in diaphragm, gastrocnemius, soleus, myocardium, and lungs of nonseptic and septic (cecal ligation and puncture 24 hours before MV) rats undergoing MV (n = 7 per group) for 150 minutes using 3 different strategies (low V(T) [V(T) = 9 mL/kg], moderate V(T) [V(T) = 15 mL/kg], and high V(T) [V(T) = 25 mL/kg]) and in nonventilated control animals. Compared with nonventilated control animals, in septic and nonseptic rodents (1) diaphragms, limb muscles, and myocardium of high-V(T) rats exhibited a decrease in protein oxidation and nitration levels, (2) antioxidant levels followed a specific fiber-type distribution in slow- and fast-twitch muscles, (3) tumor necrosis factor α (TNF-α) levels were higher in respiratory and limb muscles, whereas no differences were observed in myocardium, and (4) in lungs, protein oxidation was increased, antioxidants were rather decreased, and TNF-α remained unmodified. In this model of VILI, oxidative stress does not occur in distant organs or skeletal muscles of rodents after several hours of MV with moderate-to-high V(T), whereas protein oxidation levels were increased in the lungs of the animals. Inflammatory events were moderately expressed in skeletal muscles and lungs of the MV rats. Concomitant sepsis did not strongly affect the MV-induced effects on muscles, myocardium, or lungs in the rodents.

Antioxidant effect of human adult adipose-derived stromal stem cells in alveolar epithelial cells undergoing stretch.

INTRODUCTION: Alveolar epithelial cells undergo stretching during mechanical ventilation. Stretch can modify the oxidative balance in the alveolar epithelium. The aim of the present study was to evaluate the antioxidant role of human adult adipose tissue-derived stromal cells (hADSCs) when human alveolar epithelial cells were subjected to injurious cyclic overstretching. METHODS: A549 cells were subjected to biaxial stretch (0-15% change in surface area for 24h, 0.2Hz) with and without hADSCs. At the end of the experiments, oxidative stress was measured as superoxide generation using positive nuclear dihydroethidium (DHE) staining, superoxide dismutase (SOD) activity in cell lysates, 8-isoprostane concentrations in supernatant, and 3-nitrotyrosine by indirect immunofluorescence in fixed cells. RESULTS: Cyclically stretching of AECs induced a significant decrease in SOD activity, and an increase in 8-isoprostane concentrations, DHE staining and 3-nitrotyrosine staining compared with non-stretched cells. Treatment with hADSCs significantly attenuated stretch-induced changes in SOD activity, 8-isoprostane concentrations, DHE and 3-nitrotyrosine staining. CONCLUSION: These data suggest that hADSCs have an anti-oxidative effect in human alveolar epithelial cells undergoing cyclic stretch.

Genetic predisposition to acute kidney injury induced by severe sepsis.

PURPOSE: The aim of this study was to demonstrate that candidate gene polymorphisms are associated with an increased risk of acute kidney injury (AKI). MATERIALS AND METHODS: Patients admitted to the intensive care unit with the diagnosis of severe sepsis and an expected intensive care unit length of stay more than 48 hours were included. Genetic polymorphisms studied included angiotensin-converting enzyme insertion/deletion (polymerase chain reaction); tumor necrosis factor α -376, - 308, and -238; interleukin-8 -251; vascular endothelial growth factor (VEGF) +405 and +936; and pre-B-cell colony-enhancing factor -1001 (TaqMan SNP genotyping assay, Life Technologies, Grand Island, NY). Acute kidney injury was defined as the risk, injury, and failure categories, as per the RIFLE (risk, injury, failure, loss, end-stage kidney disease) classification. RESULTS: One hundred thirty-nine patients were included, 65 of whom developed AKI. In univariate analysis, the VEGF +936 CC and the pre-B-cell colony-enhancing factor -1001 GG genotypes were associated with AKI. In multivariate analysis, Simplified Acute Physiology Score II score (odds ratio [95% confidence interval], 1.06 [1.03-1.09]), chronic arterial hypertension (3.15 [1.39-7.15]), and the presence of the VEGF +936 CC genotype (3.41 [1.19-9.79]) were associated with AKI. CONCLUSION: This is the first study demonstrating an association between the VEGF +936 CC genotype and the risk to develop AKI in patients with severe sepsis.

Genetic predisposition to acute respiratory distress syndrome in patients with severe sepsis.

OBJECTIVE: The objective of this study was to analyze the association between candidate gene polymorphisms and susceptibility to acute respiratory distress syndrome (ARDS) in patients with severe sepsis. METHODS: Patients older than 18 years admitted to the intensive care unit (ICU) with the diagnosis of severe sepsis were prospectively included. A blood sample was drawn on the first day of ICU admission, and DNA was extracted. We genotyped the insertion/deletion polymorphism of the angiotensin-converting enzyme (ACE) gene (polymerase chain reaction) and the following single-nucleotide polymorphisms (TaqMan SNP genotyping assay): tumor necrosis factor α -376 G/A, -308 G/A, and -238 G/A; interleukin 8 -251 T/A; pre-B cell colony-enhancing factor -1001 G/T; and vascular endothelial growth factor +405 C/G and +936 C/T. Polymorphisms were selected based on reports on their association with ARDS. Variables associated in univariate analysis (P < 0.1) with the diagnosis of ARDS were included in a multiple logistic regression analysis. RESULTS: We studied 149 patients, of whom 35 presented ARDS. Variables included in the maximal multivariate model were male sex, chronic alcoholism, use of ACE inhibitors or angiotensin-receptor blockers, Simplified Acute Physiology Score II score, serum glucose concentration at ICU admission, and the presence of the allele D of the ACE gene. After adjustment for those variables, the presence of the allele D of the ACE gene (odds ratio, 4.75; 95% confidence interval, 1.02-22.20; P = 0.048) was significantly associated with the diagnosis of ARDS. CONCLUSION: The presence of the allele D of the ACE gene is associated with ARDS in patients with severe sepsis.

Role of peroxynitrite in sepsis-induced acute kidney injury in an experimental model of sepsis in rats.

The mechanisms involved in sepsis-induced acute kidney injury (AKI) are unknown. We investigated the role of nitrosative stress in sepsis-induced AKI by studying the effects of manganese (III) tetrakis-(1-methyl-4-pyridyl) porphyrin pentachloride (MnTMPyP), a peroxynitrite decomposition catalyst, and aminoguanidine (AG), a selective nitric oxide synthase 2 (NOS2) inhibitor and peroxynitrite scavenger, on kidney function of rats subjected to cecal ligation and puncture (CLP). Sprague-Dawley rats (weighing 350 [SD, 50] g) were treated with MnTMPyP (6 mg/kg i.p.) or AG (50 mg/kg i.p.) at t = 12 and 24 h after CLP or sham procedure. At t = 36 h, mean arterial pressure and aortic blood flow were measured, and blood and urine samples were obtained for biochemical determinations, including creatinine clearance, fractional excretion of sodium, and neutrophil gelatinase-associated lipocalin concentration in the urine. Kidney tissue samples were obtained for (i) light microscopy, (ii) immunofluorescence and Western blot for 3-nitrotyrosine and NOS2, (iii) gene expression (quantitative real-time polymerase chain reaction) studies (NOS1, NOS2, NOS3, and superoxide dismutase 1), and (iv) matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Mean arterial pressure was unchanged and aortic blood flow decreased 25% in CLP animals. The sepsis-induced (i) decreased urine output and creatinine clearance and increased fractional excretion of sodium and urinary neutrophil gelatinase-associated lipocalin concentration, (ii) increased protein nitration and NOS2 protein, and (iii) NOS1 and NOS2 upregulation were all significantly attenuated by treatment with MnTMPyP or AG. Nitrated proteins in renal tissue from CLP animals (matrix-assisted laser desorption ionization time-of-flight mass spectrometry) were glutamate dehydrogenase, methylmalonate-semialdehyde dehydrogenase, and aldehyde dehydrogenase, mitochondrial proteins involved in energy metabolism or antioxidant defense. Nitro-oxidative stress is involved in sepsis-induced AKI, and protein nitration seems to be one mechanism involved.

Vascular endothelial growth factor polymorphisms are involved in the late vascular complications in Type II diabetic patients.

AIM: To determine the potential genotype differences in the vascular endothelial growth factor (VEGF) gene in diabetic patients, which might explain the difference in terms of the development of clinical vascular complications: great vessels atherosclerosis vs. retinopathy. METHODS: Genotyping of the VEFG gene insertion/deletion -2549, the C-2578A and the G+405C polymorphisms was done in 40 diabetic patients (26 with peripheral artery disease (PAD) and 14 with diabetic retinopathy (DR)). RESULTS: There was a significant increase in the frequency of the VEGF -2549 DD genotype in PAD patients compared with the DR group (34.6 vs. 0; p = 0.016), as well as in the distribution of the VEGF -2549 ID genotype in DR compared with PAD patients (85.7 vs. 38.4; p = 0.005). There was a significant increase in the frequency of the VEGF -2578 CC genotype in the PAD group compared with DR (34.6 vs. 0; p = 0.016), as well as in the VEGF -2578 CA genotype in DR patients compared with PAD (85.7 vs. 34.6; p = 0.002). The VEGF +405 genotype was not associated with diabetic vascular complications. CONCLUSION: This study provides preliminary evidence that VEGF polymorphisms are associated with a differential presentation of diabetic vascular complications.

Vascular dysfunction in sepsis: effects of the peroxynitrite decomposition catalyst MnTMPyP.

The mechanisms contributing to sepsis vascular dysfunction are not well known. We tested the hypothesis that peroxynitrite scavenging ameliorates sepsis-induced macrovascular and microvascular dysfunction. Male Sprague-Dawley rats were killed 48 h after cecal ligation (n = 15) and puncture or sham procedure (n = 15). Their aortas and mesenteric vessels were mounted in organ baths for isometric tension recording. We studied contraction in resting vessels (norepinephrine 1 nM-10 µM and 10 nM-10 µM) and endothelium-dependent relaxation (acetylcholine, 10 nM-10 µM and 1 nM-10 µM) for aortas and microvessels, respectively. Vascular rings were preincubated for 30 min with the superoxide scavenger Cu-Zn-superoxide dismutase (SOD) (100 U/mL), the SOD mimetic and peroxynitrite scavenger tempol (10 M), the NO synthase inhibitor N-nitro-l-arginine methyl ester (10 M), or the peroxynitrite decomposition catalyst manganese tetrakis(4-N-methylpyridyl)porphyrin (MnTMPyP) (10 M). Fluorescence to 3-nitrotyrosine, oxidized dihydroethidium, and NOS2 was assessed in vascular tissue. Vascular NOS2, endothelial nitric oxide synthase (NOS1), NADPH-oxidase-1 (NOX-1), and SOD expression was analyzed by reverse transcription-polymerase chain reaction. Sepsis induced (i) in macrovessels, impairment of norepinephrine-induced contractions; (ii) in microvessels, impairment in norepinephrine-induced contractions and acetylcholine-induced relaxations; (iii) aortic and microvascular tissue increased reactivity to 3-nitrotyrosine, oxidized dihydroethidium, NOS2, and increased expression of NOS2, as well as increased expression of NOX-1 in microvascular tissue. Contractile responses in aortic and microvascular rings improved by ex vivo treatment with MnTMPyP and tempol, whereas vascular relaxation in microvessels improved only with MnTMPyP. Peroxynitrite scavenging protects from vascular dysfunction in sepsis.