Autoimmunity in CD73/Ecto-5′-Nucleotidase Deficient Mice Induces Renal Injury

Extracellular adenosine formed by 5′-ectonucleotidase (CD73) is involved in tubulo-glomerular feedback in the kidney but is also known to be an important immune modulator. Since CD73−/−mutant mice exhibit a vascular proinflammatory phenotype, we asked whether long term lack of CD73 causes inflammation related kidney pathologies. CD73−/−mice (13 weeks old) showed significantly increased low molecule proteinuria compared to C57BL6 wild type controls (4.8≥0.52 vs. 2.9±0.54 mg/24 h, p<0.03). Total proteinuria increased to 5.97±0.78 vs. 2.55±0.35 mg/24 h at 30 weeks (p<0.01) whereas creatinine clearance decreased (0.161±0.02 vs. 0.224±0.02 ml/min). We observed autoimmune inflammation in CD73−/−mice with glomerulitis and peritubular capillaritis, showing glomerular deposition of IgG and C3 and enhanced presence of CD11b, CD8, CD25 as well as GR-1-positive cells in the interstitium. Vascular inflammation was associated with enhanced serum levels of the cytokines IL-18 and TNF-α as well as VEGF and the chemokine MIP-2 (CXCL-2) in CD73−/−mice, whereas chemokines and cytokines in the kidney tissue were unaltered or reduced. In CD73−/−mice glomeruli, we found a reduced number of podocytes and endothelial fenestrations, increased capillaries per glomeruli, endotheliosis and enhanced tubular fibrosis. Our results show that adult CD73−/−mice exhibit spontaneous proteinuria and renal functional deterioration even without exogenous stress factors. We have identified an autoimmune inflammatory phenotype comprising the glomerular endothelium, leading to glomeruli inflammation and injury and to a cellular infiltrate of the renal interstitium. Thus, long term lack of CD73 reduced renal function and is associated with autoimmune inflammation

Nitric oxide synthetic pathway and cGMP levels are altered in red blood cells from end-stage renal disease patients

Red blood cells (RBCs) enzymatically produce nitric oxide (NO) by a functional RBC-nitric oxide synthase (RBC-NOS). NO is a vascular key regulatory molecule. In RBCs its generation is complex and influenced by several factors, including insulin, acetylcholine, and calcium. NO availability is reduced in end-stage renal disease (ESRD) and associated with endothelial dysfunction. We previously demonstrated that, through increased phosphatidylserine membrane exposure, ESRD-RBCs augmented their adhesion to human cultured endothelium, in which NO bioavailability decreased. Since RBC-NOS-dependent NO production in ESRD is unknown, this study aimed to investigate RBC-NOS levels/activation, NO production/bioavailability in RBCs from healthy control subjects (C, N = 18) and ESRD patients (N = 27). Although RBC-NOS expression was lower in ESRD-RBCs, NO, cyclic guanosine monophosphate (cGMP), RBC-NOS Serine1177 phosphorylation level and eNOS/Calmodulin (CaM)/Heat Shock Protein-90 (HSP90) interaction levels were higher in ESRD-RBCs, indicating increased enzyme activation. Conversely, following RBCs stimulation with insulin or ionomycin, NO and cGMP levels were significantly lower in ESRD- than in C-RBCs, suggesting that uremia might reduce the RBC-NOS response to further stimuli. Additionally, the activity of multidrug-resistance-associated protein-4 (MRP4; cGMP-membrane transporter) was significantly lower in ESRD-RBCs, suggesting a possible compromised efflux of cGMP across the ESRD-RBCs membrane. This study for the first time showed highest basal RBC-NOS activation in ESRD-RBCs, possibly to reduce the negative impact of decreased NOS expression. It is further conceivable that high NO production only partially affects cell function of ESRD-RBCs maybe because in vivo they are unable to respond to physiologic stimuli, such as calcium and/or insulin

TRPC3-mediated Ca2+signals as a promising strategy to boost therapeutic angiogenesis in failing hearts: The role of autologous endothelial colony forming cells

Endothelial progenitor cells (EPCs) are a sub-population of bone marrow-derived mononuclear cells that are released in circulation to restore damaged endothelium during its physiological turnover or rescue blood perfusion after an ischemic insult. Additionally, they may be mobilized from perivascular niches located within larger arteries’ wall in response to hypoxic conditions. For this reason, EPCs have been regarded as an effective tool to promote revascularization and functional recovery of ischemic hearts, but clinical application failed to exploit the full potential of patients-derived cells. Indeed, the frequency and biological activity of EPCs are compromised in aging individuals or in subjects suffering from severe cardiovascular risk factors. Rejuvenating the reparative phenotype of autologous EPCs through a gene transfer approach has, therefore, been put forward as an alternative approach to enhance their therapeutic potential in cardiovascular patients. An increase in intracellular Ca 2+ concentration constitutes a pivotal signal for the activation of the so-called endothelial colony forming cells (ECFCs), the only known truly endothelial EPC subset. Studies from our group showed that the Ca 2+ toolkit differs between peripheral blood- and umbilical cord blood (UCB)-derived ECFCs. In the present article, we first discuss how VEGF uses repetitive Ca 2+ spikes to regulate angiogenesis in ECFCs and outline how VEGF-induced intracellular Ca 2+ oscillations differ between the two ECFC subtypes. We then hypothesize about the possibility to rejuvenate the biological activity of autologous ECFCs by transfecting the cell with the Ca 2+ -permeable channel Transient Receptor Potential Canonical 3, which selectively drives the Ca 2+ response to VEGF in UCB-derived ECFCs

Haematopoietic stem cells and endothelial progenitor cells in healthy men: effect of aging and training.

Contains fulltext : 50102.pdf (publisher's version ) (Open Access)The number of hematopoietic stem cells (HSC) and endothelial progenitor cells (EPC) is thought to be a marker for neovascularization and vascular repair. Because physical inactivity and aging are risk factors for cardiovascular diseases, these factors may influence the numbers of HSCs and EPCs. Therefore, we examined baseline and exercise-induced levels of HSCs and EPCs in sedentary and trained young and older men. To study the role of aging in eight sedentary young (19-28 years) and eight sedentary older men (67-76 years), baseline and acute exercise-induced numbers of HSCs (CD34+-cells) and EPCs (CD34+/VEGFR-2+-cells) were quantified by fluorescence-activated cell sorter (FACS) analysis. To examine the effect of chronic training, eight age-matched trained young men (18-28 years) were compared with sedentary young men, whereas older men performed an 8-week endurance training. Older men showed significantly lower baseline and exercise-induced levels of HSCs/EPCs than the young men (P < 0.05). In young and older men, acute exercise significantly increased HSCs (P < 0.01), but not EPCs. The absolute increase in numbers of HSCs was attenuated in older men (P = 0.03). Apart from the lower baseline numbers of EPCs after chronic training in older men, training status did not alter baseline or exercise-induced levels of HSCs/EPCs in young and older men. We concluded that advancing age results in lower circulating numbers of HSCs and EPCs and attenuates the acute exercise-induced increase in HSCs. Interestingly, in young as well as in older men chronic endurance training does not affect baseline and exercise-induced numbers of HSCs and EPCs

Hyperoxia disrupts vascular endothelial growth factor-nitric oxide signaling and decreases growth of endothelial colony-forming cells from preterm infants

Exposure of preterm infants to hyperoxia impairs vascular growth, contributing to the development of bronchopulmonary dysplasia and retinopathy of prematurity. Disruption of vascular endothelial growth factor (VEGF)-nitric oxide (NO) signaling impairs vascular growth. Endothelial progenitor cells (EPCs) may play an important role in vascular growth. Endothelial colony-forming cells (ECFCs), a type of EPC, from human preterm cord blood are more susceptible to hyperoxia-induced growth impairment than term ECFCs. Therefore, we hypothesized that hyperoxia disrupts VEGF-NO signaling and impairs growth in preterm ECFCs and that exogenous VEGF or NO preserves growth in hyperoxia. Growth kinetics of preterm cord blood-derived ECFCs (gestational ages, 27–34 wk) were assessed in room air (RA) and hyperoxia (40–50% oxygen) with or without VEGF, NO, or Nω-nitro-l-arginine. VEGF, VEGF receptor-2 (VEGFR-2), and endothelial NO synthase (eNOS) protein expression and NO production were compared. Compared with RA controls, hyperoxia significantly decreased growth, VEGFR-2 and eNOS expression, and NO production. VEGF treatment restored growth in hyperoxia to values measured in RA controls and significantly increased eNOS expression in hyperoxia. NO treatment also increased growth in hyperoxia. Nω-nitro-l-arginine treatment inhibited VEGF-augmented growth in RA and hyperoxia. We conclude that hyperoxia decreases growth and disrupts VEGF-NO signaling in human preterm ECFCs. VEGF treatment restores growth in hyperoxia by increasing NO production. NO treatment also increases growth during hyperoxia. Exogenous VEGF or NO may protect preterm ECFCs from the adverse effects of hyperoxia and preservation of ECFC function may improve outcomes of preterm infants