Postischemic Blood Flow Research Articles

Inhibition of transient receptor potential cation channel 6 promotes capillary arterialization during post-ischaemic blood flow recovery.

Capillary arterialization, characterized by the coverage of pre-existing or nascent capillary vessels with vascular smooth muscle cells (VSMCs), is critical for the development of collateral arterioles to improve post-ischaemic blood flow. We previously demonstrated that the inhibition of transient receptor potential 6 subfamily C, member 6 (TRPC6) channels facilitate contractile differentiation of VSMCs under ischaemic stress. We here investigated whether TRPC6 inhibition promotes post-ischaemic blood flow recovery through capillary arterialization in vivo. Mice were subjected to hindlimb ischaemia by ligating left femoral artery. The recovery rate of peripheral blood flow was calculated by the ratio of ischaemic left leg to non-ischaemic right one. The number and diameter of blood vessels were analysed by immunohistochemistry. Expression and phosphorylation levels of TRPC6 proteins were determined by western blotting and immunohistochemistry. Although the post-ischaemic blood flow recovery is reportedly dependent on endothelium-dependent relaxing factors, systemic TRPC6 deletion significantly promoted blood flow recovery under the condition that nitric oxide or prostacyclin production were inhibited, accompanying capillary arterialization. Cilostazol, a clinically approved drug for peripheral arterial disease, facilitates blood flow recovery by inactivating TRPC6 via phosphorylation at Thr69 in VSMCs. Furthermore, inhibition of TRPC6 channel activity by pyrazole-2 (Pyr2; BTP2; YM-58483) promoted post-ischaemic blood flow recovery in Apolipoprotein E-knockout mice. Suppression of TRPC6 channel activity in VSMCs could be a new strategy for the improvement of post-ischaemic peripheral blood circulation.

Endothelial S1pr2 regulates post-ischemic angiogenesis via AKT/eNOS signaling pathway.

Aims: It is important to understand the mechanism that regulates post-ischemic angiogenesis and to explore a new therapeutic target for an effective improvement of revascularization in peripheral artery disease (PAD) patients. Post-ischemic angiogenesis is a highly orchestrated process, which involves vascular endothelial cells (ECs) proliferation, migration and assembly into capillaries. We found a significant reduction of S1pr2 (sphingosine 1-phosphate receptor 2) in endothelial cells after hindlimb ischemia (HLI). We thus hypothesized that EC-S1pr2 might be involved in the regulation of post-ischemic angiogenesis and blood flow recovery during peripheral arterial disease (PAD).Methods and Results: We generated both EC-specific S1pr2 loss-of-function and S1pr2 gain-of-function mice. Our study showed that EC-specific S1pr2 loss-of-function significantly enhanced post-ischemic angiogenesis and improved blood flow recovery upon femoral artery ligation, whereas the EC-specific S1pr2 gain-of-function severely hindered post-ischemic angiogenesis and reduced blood flow recovery in ischemic limbs. We next identified that S1pr2 inhibited AKT/eNOS signaling pathway, and thus inhibited EC proliferation/migration and angiogenic activity. As expected, pharmacological inhibition of S1pr2 by JTE013 improved post-ischemic angiogenesis and improved blood flow perfusion after femoral artery ligation. Moreover, we developed RGD-peptide magnetic nanoparticles packaging S1pr2-siRNA which specifically targeted ECs and achieved an efficient silencing of S1pr2 expression in ECs in vivo. This EC-targeted strategy to dampen S1pr2 significantly enhanced post-ischemic angiogenesis and boosted blood perfusion after HLI, supplying a novel therapy target for patients with peripheral arterial disease.Conclusions: This present study demonstrates that EC-expressing S1pr2 tightly controls post-ischemic angiogenesis and blood flow perfusion recovery. This research provides a novel strategy for EC-target knockdown of S1pr2 as a new therapeutic intervention for patients with peripheral artery disease.

Peripheral microvascular dysfunction is also present in patients with ischemia and no obstructive coronary artery disease (INOCA).

In patients with ischemia and no obstructive coronary artery disease (INOCA), coronary microvascular dysfunction is associated with higher rate of major adverse cardiovascular events. To demonstrate if microvascular dysfunction present in coronary microcirculation of patients with INOCA may be detected noninvasively in their peripheral circulation. 25 patients with INOCA and 25 apparently healthy individuals (controls) were subjected to nailfold videocapillaroscopy (NVC) and venous occlusion plethysmography (VOP) to evaluate peripheral microvascular function and blood collection for biomarkers analysis, including soluble vascular cell adhesion molecule-1 (sVCAM-1), endothelin-1 (ET-1) and C-reactive protein (CRP). Red blood cell velocity (RBCV) before and after ischemia (RBCVmax) were significantly lower in patients with INOCA (p = 0.0001). Time to reach maximal red blood cell velocity (TRBCVmax) was significantly longer in INOCA group (p = 0.0004). Concerning VOP, maximal blood flow (p = 0.004) and its relative increment were significantly lower in patients with INOCA (p = 0.0004). RBCVmax showed significant correlations with sVCAM-1 (r = -0.38, p < 0.05), ET-1 (r = -0.73, p < 0.05) and CRP (r = -0.33, p < 0.05). Relative increment of maximal post-ischemic blood flow was significantly correlated with sVCAM-1 (r = -0.42, p < 0.05) and ET-1 (r = -0.48, p < 0.05). The impairment of microvascular function present in coronary microcirculation of patients with INOCA can be also detected in peripheral microcirculation.

One-day acceptance and commitment therapy (ACT) workshop improves anxiety but not vascular function or inflammation in adults with moderate to high anxiety levels in a randomized controlled trial

ObjectiveAcceptance and Commitment Therapy (ACT) is a behavioral intervention demonstrating sustained improvements in anxiety in individuals with chronic anxiety and psychological distress. Because anxiety disorders are associated with the development of cardiovascular disease (CVD), we hypothesized that a novel 1-day ACT workshop would both lower anxiety and improve vascular function in persons with moderate/high anxiety. MethodsIn a randomized controlled study, 72 adults (age 33.9 ± 8.6 (SD) years) with baseline moderate/high anxiety completed a one-day ACT intervention (n = 44, age 33.9 ± 8.7 years) or control (n = 28, age 37.1 ± 10.1 years). Pre-specified secondary outcomes were measured over 12 weeks: aortic stiffness (carotid-femoral pulse wave velocity [cfPWV]), forearm vascular endothelial function (post-ischemic peak forearm blood flow [FBF] via plethysmography), and brachial artery flow-mediated dilation (FMD). Carotid artery stiffness (β-stiffness index), and inflammatory markers (C-reactive protein and tumor necrosis factor-alpha) were also explored. ResultsAlthough the intervention had a significant and sustained effect on the primary outcome of anxiety as measured by the Beck Anxiety Inventory, the 1-day ACT workshop was not associated with improvement in vascular or inflammatory endpoints. The intervention was unexpectedly associated with increases in β-stiffness index that were also associated with changing trait anxiety. ConclusionAnxiety improvements did not translate into improvements in any of the vascular function outcomes. This may reflect a less-than-robust effect of the intervention on anxiety, failure in design to select those with vascular dysfunction, or not intervening on a relevant causal pathway. (Trial registration NCT02915874 at www.clinicaltrials.gov).

Rivaroxaban compared with low-dose aspirin in individuals with type 2 diabetes and high cardiovascular risk: a randomised trial to assess effects on endothelial function, platelet activation and vascular biomarkers

Aims/hypothesisIndividuals with type 2 diabetes mellitus and subclinical inflammation have stimulated coagulation, activated platelets and endothelial dysfunction. Recent studies with the direct factor Xa inhibitor rivaroxaban in combination with low-dose aspirin demonstrated a significant reduction of major cardiovascular events, especially in individuals with type 2 diabetes and proven cardiovascular disease. Therefore, we asked the question of whether treatment with rivaroxaban could influence endothelial function, arterial stiffness and platelet activation.MethodsWe conducted a multi-centre, prospective, randomised, open-label trial in 179 participants with type 2 diabetes (duration 2–20 years), subclinical inflammation (high-sensitivity C-reactive protein 2–10 mg/l) and at least two traits of the metabolic syndrome to compare the effects of the direct factor Xa inhibitor rivaroxaban (5 mg twice daily) vs aspirin (100 mg every day) on endothelial function (assessed by forearm occlusion plethysmography), skin blood flow (assessed by laser-Doppler fluxmetry), arterial stiffness (assessed by pulse wave velocity) and serum biomarkers of endothelial function and inflammation. Furthermore, we investigated phosphorylation of vasodilator-stimulated phosphoprotein (VASP) in platelets, the concentration of platelet-derived microparticles (PMPs) and the effects of isolated PMPs on HUVEC proliferation in vitro.ResultsRivaroxaban treatment for 20 weeks (n = 89) resulted in a significant improvement of post-ischaemic forearm blood flow (3.6 ± 4.7 vs 1.0 ± 5.2 ml/100 ml, p = 0.004), a numerically increased skin blood flow and reduced soluble P-Selectin plasma level vs aspirin. We did not find significant differences of arterial stiffness or further biomarkers. Neither rivaroxaban nor aspirin influenced VASP phosphorylation of platelets. The number of PMPs increased significantly with both rivaroxaban (365.2 ± 372.1 vs 237.4 ± 157.1 μl−1, p = 0.005) and aspirin (266.0 ± 212.7 vs 201.7 ± 162.7 μl−1, p = 0.021). PMPs of rivaroxaban-treated participants stimulated HUVEC proliferation in vitro compared with aspirin. Rivaroxaban was associated with a higher number of bleeding events.Conclusions/interpretationOur findings indicate that the direct factor Xa inhibitor rivaroxaban improved endothelial function in participants with type 2 diabetes and subclinical inflammation but also increased the risk of bleeding.Trial registration:ClinicalTrials.gov NCT02164578.FundingThe study was supported by a research grant from Bayer Vital AG, Germany.Graphical abstract

Impaired Endothelial Function in Patients With Postural Tachycardia Syndrome.

The purpose of this study is to evaluate endothelial function in postural tachycardia syndrome (PoTS), a poorly understood chronic condition characterized by a state of consistent orthostatic tachycardia (delta heart rate ≥30 beats per minute) upon standing without orthostatic hypotension. Nineteen patients with PoTS and 9 healthy controls were studied after 3 days of a fixed, caffeine-free, normal sodium (150 milliequivalents/day) diet. All participants underwent autonomic function testing, including sinus arrhythmia, valsalva maneuver, hyperventilation, cold pressor, handgrip, and a standing test with catecholamine measurements, followed by endothelial function testing. We analyzed 3 measures of endothelial function: percent brachial flow-mediated dilation, digital pulsatile arterial tonometry, and postischemic percent leg blood flow. Flow-mediated dilation was significantly lower in patients with PoTS (6.23±3.54% for PoTS) than in healthy controls (10.6±4.37% for controls versus, P=0.014). PoTS and controls had similar digital pulsatile arterial tonometry (1.93±0.40 arbitrary units for controls versus 2.13±0.63 arbitrary units for PoTS). PoTS had similar but suggestive percent leg blood flow to controls (313±158% for PoTS versus 468±236% for controls, P=0.098). Patients with PoTS have significantly reduced flow-mediated dilation compared with healthy controls, suggesting that PoTS is characterized by endothelial dysfunction in conduit arteries. Registration: URL: https://www.clinicaltrials.gov. Unique identifier: NCT01308099.

A randomized controlled trial of treatment with intermittent negative pressure for intermittent claudication

ObjectiveWe investigated the effects of lower extremity intermittent negative pressure (INP) treatment for 1 hour two times daily for 12 weeks on the walking distance of patients with intermittent claudication (IC). MethodsPatients with IC were randomized to treatment with −40 mm Hg INP (treatment group) or −10 mm Hg INP (sham control group). Pain-free walking distance (PWD) and maximal walking distance (MWD) on a treadmill, resting and postexercise ankle-brachial index, resting and postischemic blood flow (plethysmography), and quality of life (EQ-5D-5L and Vascuqol-6) were measured at baseline and after 12 weeks of treatment. ResultsA total of 72 patients were randomized, and 63 had data available for the intention-to-treat analyses. The between-group comparisons showed a significant change in the PWD, favoring the treatment group over the sham control group (estimated treatment effect, 50 m; 95% confidence interval [CI], 11-89; P = .014). The PWD had increased by 68 m (P < .001) in the treatment group and 18 m (P = .064) in the sham control group. No significant difference was found in the change in the MWD between the two groups (estimated treatment effect, 42 m; 95% CI, −14 to 97; P = .139). The MWD had increased by 62 m (P = .006) in the treatment group and 20 m (P = .265) in the sham control group. For patients with a baseline PWD of <200 m (n = 56), significant changes had occurred in both PWD and MWD between the two groups, favoring the treatment group (estimated treatment effect, 42 m; 95% CI, 2-83; P = .042; and estimated treatment effect, 62 m; 95% CI, 5-118; P = .032; respectively). Both overall and for the group of patients with a PWD <200 m, no significant differences were found in the changes in the resting and postexercise ankle-brachial index, resting and postischemic blood flow, or quality of life parameters between the two groups. ConclusionsTreatment with −40 mm Hg INP increased the PWD compared with sham treatment in patients with IC. For the patients with a baseline PWD of <200 m, an increase was found in both PWD and MWD compared with sham treatment.

Influence of Macitentan on the Vascular Tone and Recruitment of Finger Capillaries Under Hypobaric Hypoxia in High Altitude.

Betge, Stefan, Stefan Drinda, Thomas Neumann, Laura Bäz, Alexander Pfeil, Christian Schulze, Ralf Mrowka, Christian Jung, and Marcus Franz. Influence of macitentan on the vascular tone and recruitment of finger capillaries under hypobaric hypoxia in high altitude. High Alt Med Biol. 21:336-345, 2020. Introduction: Acute normobaric (NH) and hypobaric hypoxia (HH) has effects on the vascular tone of larger arteries and may have effects on the microcirculation. These effects may be noninvasively detectable by automated devices. A part of these effects may be mediated by endothelin (ET) and should be influenced by macitentan (MAC), a dual endothelin receptor antagonist (ERA). Methods: We used photoplethysmographic sensors, fingertip volume sensors, nailfold capillaroscopy, and laser Doppler probes at rest and after a 5-minute forearm ischemia in healthy study subjects under NH, under HH, and under HH plus a single dose of MAC. Results: NH at simulated 4000 m led to increased heart rates (HR) and pulse wave velocities (PWV) and reduced augmentation index (AIX). The values for the AIX showed a high SD and differed between the used devices. At simulated 5500 m, only baseline mean value (BMV; EndoPAT) showed a further change, indicating less filled capillaries of the fingertips. HH (2978 m) increased HR, blood pressure values, and PWV. Focusing on the microcirculation of the fingertips, HH reduced the BMV and the nailfold capillary density and the postischemic capillary recruitment. MAC had no effect on the BMV, but antagonized the effects of HH on the nailfold capillaries and led to a strongly increased postischemic diameter of the arterial limbs. Concordantly, the postischemic blood flow velocity increment, measured through ultrasound Doppler, was increased at ALT+MAC. Conclusions: The BMV may be a parameter for changes of the microcirculation of the finger tips. A single dose of MAC blocked hypoxia-induced capillary rarefaction and enhanced postischemic hyperemia of the fingertips. These results indicate the importance of ET-1 for the regulation of the microcirculation under hypoxia. The German Registry of Clinical Studies (DRKS) ID: 00005459.

Environmental enrichment enhances post-ischemic cerebral blood flow and functional hyperemia in the ipsilesional somatosensory cortex

Environmental enrichment has been reported to promote functional recovery in an ischemic stroke. However, the underlying mechanism remains unclear. This study aimed to investigate the effect of environmental enrichment treatment on post-ischemic cerebral blood flow and functional hyperemia in the ipsilesional primary somatosensory cortex of rats. With laser speckle imaging, we were able to monitor the resting cerebral blood flow alteration in the middle cerebral artery occlusion model. Both 3- and 28-day post-ischemic infarct volumes were then examined with triphenyltetrazolium chloride and cresyl violet staining, respectively. We found that an exposure to environmental enrichment was associated with higher post-ischemic cerebral blood flow and less brain tissue loss in the ipsilesional primary somatosensory cortex compared with the standard cage environment. Furthermore, environmental enrichment also enhanced the cerebral blood flow response to whisker stimulation in the ipsilesional barrel cortex when measured 28 days after the middle cerebral artery occlusion. Together, the data suggested that an exposure to environmental enrichment promoted the restoration of cerebral blood flow in the ipsilesional cortex and contributed to a better coupling between functional activation and cerebral blood flow change, which might be the possible mechanisms underlying the neuroprotective effects of EE after ischemia.

Androgens Stimulate EPC-Mediated Neovascularization and Are Associated with Increased Coronary Collateralization.

Endothelial progenitor cells (EPCs) play a key role in neovascularization and have been linked to improved cardiovascular outcomes. Although there is a well-established inverse relationship between androgen levels and cardiovascular mortality in men, the role of androgens in EPC function is not fully understood. In this study, we investigated the effects of androgens on 2 subpopulations of EPCs, early EPCs (EEPCs) and late outgrowth EPCs (OECs), and their relationships with coronary collateralization. Early EPCs and OECs were isolated from the peripheral blood of young healthy men and treated with dihydrotestosterone (DHT) with or without androgen receptor (AR) antagonist, hydroxyflutamide, in vitro. Dihydrotestosterone treatment enhanced AR-mediated proliferation, migration, and tubulogenesis of EEPCs and OECs in a dose-dependent manner. Furthermore, DHT augmented EPC sensitivity to extracellular stimulation by vascular endothelial growth factor (VEGF) via increased surface VEGF receptor expression and AKT activation. In vivo, xenotransplantation of DHT pretreated human EPCs augmented blood flow recovery and angiogenesis in BALB/c nude male mice, compared to mice receiving untreated EPCs, following hindlimb ischemia. In particular, DHT pretreated human OECs exhibited higher reparative potential than EEPCs in augmenting postischemic blood flow recovery in mice. Furthermore, whole blood was collected from the coronary sinus of men with single vessel coronary artery disease (CAD) who underwent elective percutaneous intervention (n = 23). Coronary collateralization was assessed using the collateral flow index. Serum testosterone and EPC levels were measured. In men with CAD, circulating testosterone was positively associated with the extent of coronary collateralization and the levels of OECs. In conclusion, androgens enhance EPC function and promote neovascularization after ischemia in mice and are associated with coronary collateralization in men.

The cardiovascular phenotype of adult patients with phenylketonuria

BackgroundPatients with Phenylketonuria (PKU) are exposed to multiple cardiovascular risk factors, but the clinical significance of these abnormalities is yet unknown. The purpose of this study was to characterize the cardiovascular phenotype in adult patients with PKU by clinical and dietary data, measurements of biochemical markers, and non-invasive examination of vascular functions.ResultsTwenty-three adult patients with PKU (age: 18–47 y; 30.8 ± 8.4 y) and 28 healthy controls (age: 18–47 y; 30.1 ± 9.1 y) were included in this study. PKU patients had significantly higher systolic and diastolic blood pressure, increased resting heart rate and a higher body mass index. Total cholesterol and non-HDL cholesterol levels were significantly increased in PKU patients, whereas plasma levels of HDL cholesterol and its subfraction HDL2 (but not HDL3) were significantly decreased. The inflammatory markers C-reactive protein and serum amyloid A protein and the serum oxidative stress marker malondialdehyde were significantly higher in patients with PKU. Venous occlusion plethysmography showed marked reduction in post-ischemic blood flow and the carotid to femoral pulse wave velocity was significantly increased demonstrating endothelial dysfunction and increased vascular stiffness.ConclusionsThis study shows that the cardiovascular phenotype of adult PKU patients is characterized by an accumulation of traditional cardiovascular risk factors, high levels of inflammatory and oxidative stress markers, endothelial dysfunction and vascular stiffness. These data indicate the need for early cardiovascular risk reduction in patients with PKU.

IRF3 and IRF7 mediate neovascularization via inflammatory cytokines.

ObjectiveTo elucidate the role of interferon regulatory factor (IRF)3 and IRF7 in neovascularization.MethodsUnilateral hind limb ischaemia was induced in Irf3−/−, Irf7−/− and C57BL/6 mice by ligation of the left common femoral artery. Post‐ischaemic blood flow recovery in the paw was measured with laser Doppler perfusion imaging. Soleus, adductor and gastrocnemius muscles were harvested to investigate angiogenesis and arteriogenesis and inflammation.ResultsPost‐ischaemic blood flow recovery was decreased in Irf3−/−and Irf7−/− mice compared to C57BL/6 mice at all time points up to and including sacrifice, 28 days after surgery (t28). This was supported by a decrease in angiogenesis and arteriogenesis in soleus and adductor muscles of Irf3−/− and Irf7−/− mice at t28. Furthermore, the number of macrophages around arterioles in adductor muscles was decreased in Irf3−/−and Irf7−/− mice at t28. In addition, mRNA expression levels of pro‐inflammatory cytokines (tnfα, il6, ccl2) and growth factor receptor (vegfr2), were decreased in gastrocnemius muscles of Irf3−/− and Irf7−/− mice compared to C57BL/6 mice.ConclusionDeficiency of IRF3 and IRF7 results in impaired post‐ischaemic blood flow recovery caused by attenuated angiogenesis and arteriogenesis linked to a lack of inflammatory components in ischaemic tissue. Therefore, IRF3 and IRF7 are essential regulators of neovascularization.

Artery targeted photothrombosis widens the vascular penumbra, instigates peri-infarct neovascularization and models forelimb impairments

The photothrombotic stroke model generates localized and reproducible ischemic infarcts that are useful for studying recovery mechanisms, but its failure to produce a substantial ischemic penumbra weakens its resemblance to human stroke. We examined whether a modification of this approach, confining photodamage to arteries on the cortical surface (artery-targeted photothrombosis), could better reproduce aspects of the penumbra. Following artery-targeted or traditional photothrombosis to the motor cortex of mice, post-ischemic cerebral blood flow was measured using multi-exposure speckle imaging at 6, 48, and 120 h post-occlusion. Artery-targeted photothrombosis produced a more graded penumbra at 48 and 120 h. The density of isolectin B4+ vessels in peri-infarct cortex was similarly increased after both types of infarcts compared to sham at 2 weeks. These results indicate that both models instigated post-ischemic vascular structural changes. Finally, we determined whether the strength of the traditional photothrombotic approach for modeling upper-extremity motor impairments extends to the artery-targeted approach. In adult mice that were proficient in a skilled reaching task, small motor-cortical infarcts impaired skilled-reaching performance for up to 10 days. These results support that artery-targeted photothrombosis widens the penumbra while maintaining the ability to create localized infarcts useful for modeling post-stroke impairments.

Dobutamine Versus Vasopressin After Mesenteric Ischemia

BackgroundGastrointestinal blood flow may be compromised during and after vasopressor support. Endothelin expression may lead to microcirculatory dysfunction. The aim of this study was to analyze the effect of vasopressin and dobutamine after mesenteric ischemia on the gastrointestinal mucosal microcirculation, endothelin expression, and morphologic injury. Materials and methodsPigs were studied in four groups (six pigs in each group): 1, sham; 2-4 ischemia (1 h superior mesenteric artery occlusion with 30 min reperfusion and 30 min of vehicle [2], dobutamine [3], or vasopressin [4] administration, followed by 30-min break and thiopental-induced hypotension [3, 4]). Blood flow of the gastric, jejunal, and rectosigmoidal mucosa was measured. At the end of the experiment, the mucosal expression of endothelin-1 (ET-1) and its receptor subtypes A (ETA) and B were determined by polymerase chain reaction. Mucosal injury, apoptotic cell death, and leukocytic infiltration were determined by histology and immunohistochemical analysis of cleaved caspase-3 and myeloperoxidase. ResultsMesenteric ischemia increased jejunal mucosal ET-1 gene expression, arterial ET-1, intestinal fatty acid binding protein, and jejunal mucosal injury compared with sham. Dobutamine increased arteriovenous shunting at the cost of the jejunal mucosal blood perfusion. This was associated with an increased expression of ET-1 and ETA and mucosal leukocytic infiltration. In contrast, vasopressin increased postischemic capillary density and tissue blood flow. This was associated with a lower ET-1 gene expression. Vasopressin did not induce jejunal mucosal leukocytic infiltration. ConclusionsVasopressin reduces mesenteric ischemia-associated alterations of the microcirculation and tissue integrity, whereas dobutamine does not.

MiR-210 Enhances the Therapeutic Potential of Bone-Marrow-Derived Circulating Proangiogenic Cells in the Setting of Limb Ischemia.

Therapies based on circulating proangiogenic cells (PACs) have shown promise in ischemic disease models but require further optimization to reach the bedside. Ischemia-associated hypoxia robustly increases microRNA-210 (miR-210) expression in several cell types, including endothelial cells (ECs). In ECs, miR-210 represses EphrinA3 (EFNA3), inducing proangiogenic responses. This study provides new mechanistic evidences for a role of miR-210 in PACs. PACs were obtained from either adult peripheral blood or cord blood. miR-210 expression was modulated with either an inhibitory complementary oligonucleotide (anti-miR-210) or a miRNA mimic (pre-miR-210). Scramble and absence of transfection served as controls. As expected, hypoxia increased miR-210 in PACs. Invivo, migration toward and adhesion to the ischemic endothelium facilitate the proangiogenic actions of transplanted PACs. Invitro, PAC migration toward SDF-1α/CXCL12 was impaired by anti-miR-210 and enhanced by pre-miR-210. Moreover, pre-miR-210 increased PAC adhesion to ECs and supported angiogenic responses in co-cultured ECs. These responses were not associated with changes in extracellular miR-210 and were abrogated by lentivirus-mediated EFNA3 overexpression. Finally, ex-vivo pre-miR-210 transfection predisposed PACs to induce post-ischemic therapeutic neovascularization and blood flow recovery in an immunodeficient mouse limb ischemia model. In conclusion, miR-210 modulates PAC functions and improves their therapeutic potential in limb ischemia.

Enriched Environment Elicits Proangiogenic Mechanisms After Focal Cerebral Ischemia

Brain has limited capacity for spontaneous recovery of lost function after stroke. Exposure to enriched environment (EE) can facilitate functional recovery, but mechanisms underlying this effect are poorly understood. Here, we used a middle cerebral artery occlusion (MCAO) model to investigate the impact of EE on angiogenesis in the post-ischemic brain in adult male Sprague Dawley rats, and examined whether blood-borne factors may contribute. Compared with standard cage (SC), exposure to EE was associated with greater improvement in neurological function, higher peri-infarct vascular density, and higher chronic post-ischemic cerebral blood flow assessed by laser speckle imaging. The effect persisted for at least 28days. EE also enhanced the expression of hepatocyte growth factor in the peri-ischemic cortex when measured 15days after MCAO. Interestingly, serum from rats exposed to EE after MCAO showed elevated levels of hepatocyte growth factor, and plasma or serum from rats exposed to EE after MCAO enhanced the survival and proliferation of cultured endothelial cells, in vitro, when compared with control plasma or serum from SC group after MCAO. Together, our data suggest that exposure to EE promotes angiogenesis in the ischemic brain that may in part be mediated by blood-borne factors.

The role of CD27-CD70-mediated T cell co-stimulation in vasculogenesis, arteriogenesis and angiogenesis

BackgroundT cells have a distinctive role in neovascularization, which consists of arteriogenesis and angiogenesis under pathological conditions and vasculogenesis under physiological conditions. However, the role of co-stimulation in T cell activation in neovascularization has yet to be established. The aim of this study was to investigate the role T cell co-stimulation and inhibition in angiogenesis, arteriogenesis and vasculogenesis. Methods and resultsHind limb ischemia was induced by double ligation of the left femoral artery in mice and blood flow recovery was measured with Laser Doppler Perfusion Imaging in control, CD70−/−, CD80/86−/−, CD70/80/86−/− and CTLA4+/− mice. Blood flow recovery was significantly impaired in mice lacking CD70 compared to control mice, but was similar in CD80/86−/−, CTLA4+/− and control mice. Mice lacking CD70 showed impaired vasculogenesis, since the number of pre-existing collaterals was reduced as observed in the pia mater compared to control mice. In vitro an impaired capability of vascular smooth muscle cells (VSMC) to activate T cells was observed in VSMC lacking CD70. Furthermore, CD70−/−, CD80/86−/− and CD70/80/86−/− mice showed reduced angiogenesis in the soleus muscle 10 days after ligation. Arteriogenesis was also decreased in CD70−/− compared to control mice 10 and 28 days after surgery. ConclusionsThe present study is the first to describe an important role for T cell activation via co-stimulation in angiogenesis, arteriogenesis and vasculogenesis, where the CD27-CD70 T cell co-stimulation pathway appears to be the most important co-stimulation pathway in pre-existing collateral formation and post-ischemic blood flow recovery, by arteriogenesis and angiogenesis.

AMP and adenosine are both ligands for adenosine 2B receptor signaling

Adenosine is considered the canonical ligand for the adenosine 2B receptor (A2BR). A2BR is upregulated following kidney ischemia augmenting post ischemic blood flow and limiting tubular injury. In this context the beneficial effect of A2BR signaling has been attributed to an increase in the pericellular concentration of adenosine. However, following renal ischemia both kidney adenosine monophosphate (AMP) and adenosine levels are substantially increased. Using computational modeling and calcium mobilization assays, we investigated whether AMP could also be a ligand for A2BR.The computational modeling suggested that AMP interacts with more favorable energy to A2BR compared with adenosine. Furthermore, AMPαS, a non-hydrolyzable form of AMP, increased calcium uptake by Chinese hamster ovary (CHO) cells expressing the human A2BR, indicating preferential signaling via the Gq pathway. Therefore, a putative AMP-A2BR interaction is supported by the computational modeling data and the biological results suggest this interaction involves preferential Gq activation. These data provide further insights into the role of purinergic signaling in the pathophysiology of renal IRI.

A pro-angiogenic degradable Mg-poly(lactic-co-glycolic acid) implant combined with rhbFGF in a rat limb ischemia model.

Magnesium (Mg)-based implant has been already used in patients with critical limb ischemia. Site-specific controlled release of recombinant human basic fibroblast growth factor (rhbFGF), has become a promising approach to improve peripheral vascular disease. We report here on a novel combination implant composed of spiral magnesium and a coating made using poly(lactic-co-glycolic acid) (PLGA) with encapsulated rhbFGF (Mg-PLGA-rhbFGF). The preparation method does not involve any complex processes and results in a high encapsulation efficiency (approximately 100%). The degradation of metal Mg raise the microclimate pH in the PLGA polymer, which could well preserve the bioactivity of rhbFGF incorporated in the implant. Mg-PLGA-based, sustained local delivery of rhbFGF promotes post-ischemic angiogenesis and blood flow recovery in rat limb ischemic model. This work marks the first report for controlled release of rhbFGF in combination with metal Mg, and suggests potential therapeutic usefulness of Mg-PLGA-rhbFGF for tissue ischemia.

Equilibrative nucleoside transporter 1 (ENT1) regulates postischemic blood flow during acute kidney injury in mice.

Equilibrative nucleoside transporter 1 (ENT1) regulates postischemic blood flow during acute kidney injury in mice.