Impaired Endothelial Progenitor Cells Research Articles

Endothelial progenitor cells and peripheral neuropathy in subjects with type 2 diabetes mellitus

AimsTo examine for differences in circulating progenitor cells (CPCs) and endothelial progenitor cells (EPCs) in patients with and without diabetic peripheral neuropathy (DPN). MethodsA total of 105 participants were included: 50 patients with type 2 diabetes (T2DM) and DPN, 30 patients with T2DM without DPN and 25 healthy individuals. CPCs and 6 different EPCs phenotypes were assessed with flow cytometry. We also measured plasma levels of vascular endothelial growth factor (VEGF), stromal cell-derived factor 1 (SDF-1), vascular cell adhesion protein-1 (VCAM-1), intracellular adhesion molecule-1 (ICAM) and tumor necrosis factor a (TNFa). ResultsNo difference was observed in the number of CPCs among the 3 groups. Patients with DPN had higher numbers of all 6 EPCs phenotypes when compared with patients without DPN and higher number of 5 EPCs phenotypes when compared with healthy individuals. Plasma VEFG, VCAM-1, ICAM-1 and TNFa levels did not differ among the 3 groups. Patients with DPN had lower SDF-1 levels in comparison with healthy individuals. ConclusionCirculating EPCs are increased while SDF-1 levels are decreased in the presence of DPN. Our findings suggest that DPN may be associated with impaired trafficking of EPCs and impaired EPCs homing to the injured endothelium.

B lymphocyte stimulator modulates number and function of endothelial progenitor cells in systemic lupus erythematosus

BackgroundCirculating endothelial progenitor cells (EPCs) are biologic markers of endothelial function. In patients with systemic lupus erythematosus (SLE), the numerical reduction and functional impairment of EPCs contribute to the endothelial dysfunction.Through ex vivo and in vitro studies, we aimed at evaluating the effects of B lymphocyte stimulator (BLyS) on EPC colonies and endothelial cells and also investigating BLyS receptor expression on these cells.MethodsEPCs were isolated from peripheral blood mononuclear cells (PBMC). In order to evaluate their ability to form colonies, EPCs were cultured on fibronectin-coated dishes and incubated with BlyS alone or BlyS and belimumab. Apoptosis of EPCs and endothelial cell line EA.hy926 was evaluated after 6, 12, and 24 h of incubation with BLyS and after 6 h with BLyS and belimumab. The expression of B cell activating factor-receptor (BAFF-R), B cell maturation antigen (BCMA), and transmembrane activator and calcium modulator and cyclophilin ligand (CAML) interactor (TACI) on EPCs and EA.hy926 was analyzed by cytofluorimetry.ResultsThe number of EPC colonies was lower in patients than in controls. Moreover, the colonies from SLE patients were poorly organized compared to controls; the addition of belimumab restored the colony structure. Incubation with BLyS induced apoptosis of EPCs and EA.hy926 that was inhibited by the co-incubation with belimumab. BAFF-R and BCMA were expressed on both EPCs and EA.hy926, while TACI was expressed only on EPCs.ConclusionsEPCs and endothelial cells preferentially express BAFF-R which could be involved in the pro-apoptotic effect of BlyS. Belimumab administration seems to restore the quantitative and qualitative changes of EPC colonies both ex vivo and in vitro.

Icariin reduces high glucose-induced endothelial progenitor cell dysfunction via inhibiting the p38/CREB pathway and activating the Akt/eNOS/NO pathway.

High glucose (HG) impairs endothelial progenitor cell (EPC) function. The activation of p38 mitogen-activated protein kinase and the inhibition of the Akt/eNOS/NO pathway serve central roles in this process. Icariin has protective effects in endothelial cells. The aim of the present study was to investigate the effects of icariin on HG-induced EPC dysfunction, including proliferation, migration and tube formation. Experiments were performed with EPCs isolated from the femurs and tibias of Sprague-Dawley rats in vitro. In a dose-dependent manner, icariin reversed the inhibition of EPC proliferation induced by HG treatment, and the maximal effective concentration of icariin was 1 µM [Fold change (FC):0.90±0.07, P=0.0124 vs. HG group]. The impaired EPC migration and tube formation induced by glucose was partially restored by 1 µM icariin treatment (FC: 0.81±0.08, P=0.0148 vs. HG group for migration; 0.82±0.03, P=0.0214 vs. HG group for tube formation). Furthermore, icariin significantly suppressed HG-induced p38 and cAMP response element binding protein (CREB) phosphorylation in EPCs (FC: 1.84±0.21, P=0.0238 vs. HG group for p38; FC: 2.24±0.15, P=0.0068 vs. HG group for CREB). Increased Akt and endothelial nitric oxide (NO) synthase (eNOS) activation was also observed after icariin treatment (FC: 0.64±0.08, P=0.0047 vs. HG group for Akt; FC:0.53±0.05, P=0.0019 vs. HG group for eNOS), which was followed by increased NO production (FC: 0.69±0.06, P=0.0064 vs. HG group). In conclusion, icariin attenuated HG-induced EPC dysfunction, which may be partially attributed to the inhibition of the p38/CREB pathway and the activation of the Akt/eNOS/NO pathway. Icariin may be a therapeutic candidate for improving the function of EPC.

Impairment of circulating endothelial progenitor cells (EPCs) in patients with glucocorticoid-induced avascular necrosis of the femoral head and changes of EPCs after glucocorticoid treatment in vitro

BackgroundAvascular necrosis of the femoral head (ANFH) is a severe complication after high-dose glucocorticoid (GC) administration. The pathogenesis of GC-induced ANFH remains unclear. Though the important role of endothelial progenitor cells (EPCs) in the progression of GC-induced ANFH has been noticed, the effects of GCs on EPCs and the underlying mechanism still need further study.MethodsCirculating EPCs were obtained from the peripheral blood of ANFH patients and healthy controls by Ficoll-density gradient centrifugation. CD133+CD34+ cells with DiI-Ac-LDL uptake and FITC-UEA-1 binding were considered as EPCs. Number and functions of EPCs were analyzed by flow cytometry, chemotaxis assay, and tube formation assay. EPCs from healthy controls were also treated by different concentrations of methylprednisolone and prednisolone in vitro, and cell growth and angiogenic function were evaluated. Expression of CXCR7 and its downstream Akt/GSK-3β/Fyn pathway were also analyzed by western blots after cells treated by methylprednisolone in vitro.ResultsThe number and functions of EPCs in patients with GC-induced ANFH were significantly decreased. In vitro study showed for the first time that except extremely high concentrations, low to medium concentrations of GCs did not have significant effects on EPCs’ growth. Methylprednisolone and prednisolone both inhibited angiogenesis of EPCs even at low concentrations. Mechanism studies found CXCR7 was downregulated in EPCs after methylprednisolone treatment in vitro. Expression and phosphorylation of Akt and GSK-3β were also decreased with an upregulation of Fyn expression after steroid treatment.ConclusionsOur study showed that GC-induced ANFH patients have reduced the number and impaired functions of circulating EPCs. GCs did not show a significant effect on the growth of EPCs in vitro except extremely high concentrations of GCs. However, GCs significantly impaired EPC angiogenic function in vitro, even at low concentrations. Our study also suggested that downregulation of CXCR7 and its downstream Akt/GSK-3β/Fyn pathway in EPCs might be a novel mechanism of how GCs suppress EPCs’ angiogenesis.

Preliminary Study: Purple Sweet Potato Extract Seems to Be Superior to Increase the Migration of Impaired Endothelial Progenitor Cells Compared to l-Ascorbic Acid

Impairment of the endothelial progenitor cells (EPCs) ability to proliferate and migrate in the patients with coronary heart disease (CHD) is partly caused by oxidative stress. This research evaluates the effect of treatment with Ipomoea batatas L./purple sweet potato (PSP) extract and l-ascorbic acid on the proliferation and migration of impaired EPCs. EPCs were isolated from CHD patient’s peripheral blood. EPCs culture were cultivated and divided into control (untreated), PSP extract treatment (dose 1 and 25 μg/mL), and l-ascorbic acid treatment (dose 10 and 250 μg/mL) groups for 48 h. EPCs proliferation was analyzed with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) cell proliferation assay, and migration was evaluated with the cell migration assay kit. Statistical tests were evaluated using SPSS 25.0. This research showed that EPCs proliferation and migration was significantly higher in all PSP extract and l-ascorbic acid treatment compared to the control (p < 0.001). EPCs migration on treatment with a PSP extract dose of 25 μg/mL was significantly higher compared to the treatment with l-ascorbic acid dose of 250 μg/mL (303,000 ± 1000 compared to 215,000 ± 3000 cells, p< 0.001). In conclusion, both treatments with PSP extract and l-ascorbic acid can improve the proliferation and migration of impaired EPCs. At the dose of 25 μg/mL, PSP extract seems to be superior to the l-ascorbic acid dose of 250 μg/mL to improve EPCs migration.

Astragaloside IV protects endothelial progenitor cells from the damage of ox-LDL via the LOX-1/NLRP3 inflammasome pathway.

Purpose: Functional impairment of endothelial progenitor cells (EPCs) is frequently observed in patients with diabetic vascular complications. Astragaloside IV (ASV) has a significant protective effect against vascular endothelial dysfunction. Thus, this study aimed to investigate the role of ASV on oxidized low-density lipoprotein (ox-LDL)-induced EPCs dysfunction and its potential mechanisms.Methods: EPCs were isolated from the peripheral blood of mice and treated with different concentration of ASV (10, 20, 40, 60, 80, 100 and 200 µM). ox-LDL was served as a stimulus for cell model. The proliferation and migration, and improved tube formation ability of EPCs were determined. Reactive oxygen species (ROS) production and the levels of inflammatory cytokines, including interleukin 1β (IL-1β), IL-6, IL-10 and tumor necrosis factor (TNF-α) were measured. The expression oflectin-like oxidized LDL receptor (LOX-1) andNod-like receptor nucleotide-binding domain leucine rich repeat containing protein 3 (NLRP3) inflammasome were detected by Western blot analysis.Results: We found ASV treatment alleviated ox-LDL-induced cellular dysfunction, as evidenced by promoted proliferation and migration, and improved tube formation ability. Besides, ASV treatment significantly suppressed ox-LDL-induced ROS production and the levels of inflammatory cytokines. ASV inhibited ox-LDL-induced expression of LOX-1 in a concentration-dependent manner. Overexpression of LOX-1 in EPCs triggered NLRP3inflammasome activation, while inhibition of LOX-1 or treatment with ASV suppressed ox-LDL-induced NLRP3 inflammasome activation. Furthermore, overexpression of LOX-1 in ox-LDL-induced EPCs furtherly impaired cellular function, which could be ameliorated by ASV treatment.Conclusion: Our study showed that ASV may protect EPCs against ox-LDL-induced dysfunction via LOX-1/NLRP3 pathway.

Dipeptidyl dipeptidase-4 inhibitor recovered ischemia through an increase in vasculogenic endothelial progenitor cells and regeneration-associated cells in diet-induced obese mice.

Metabolic syndrome (MS), overlapping type 2 diabetes, hyperlipidemia, and/or hypertension, owing to high-fat diet, poses risk for cardiovascular disease. A critical feature associated with such risk is the functional impairment of endothelial progenitor cells (EPCs). Dipeptidyl dipeptidase-4 inhibitors (DPP-4 i) not only inhibit degradation of incretins to control blood glucose levels, but also improve EPC bioactivity and induce anti-inflammatory effects in tissues. In the present study, we investigated the effects of such an inhibitor, MK-06266, in an ischemia model of MS using diet-induced obese (DIO) mice. EPC bioactivity was examined in MK-0626-administered DIO mice and a non-treated control group, using an EPC colony-forming assay and bone marrow cKit+ Sca-1+ lineage-cells, and peripheral blood-mononuclear cells. Our results showed that, in vitro, the effect of MK-0626 treatment on EPC bioactivities and differentiation was superior compared to the control. Furthermore, microvascular density and pericyte-recruited arteriole number increased in MK-0626-administered mice, but not in the control group. Lineage profiling of isolated cells from ischemic tissues revealed that MK-0626 administration has an inhibitory effect on unproductive inflammation. This occurred via a decrease in the influx of total blood cells and pro-inflammatory cells such as neutrophils, total macrophages, M1, total T-cells, cytotoxic T-cells, and B-cells, with a concomitant increase in number of regeneration-associated cells, such as M2/M ratio and Treg/T-helper. Laser Doppler analysis revealed that at day 14 after ischemic injury, blood perfusion in hindlimb was greater in MK-0626-treated DIO mice, but not in control. In conclusion, the DPP-4 i had a positive effect on EPC differentiation in MS model of DIO mice. Following ischemic injury, DPP-4 i sharply reduced recruitment of pro-inflammatory cells into ischemic tissue and triggered regeneration and reparation, making it a promising therapeutic agent for MS treatment.

Hyperinsulinemia impairs functions of circulating endothelial progenitor cells.

Circulating endothelial progenitor cells (EPCs) play a key role in maintaining endothelial function. Dysfunction of EPCs is associated with the cardiovascular complication of diabetes. The purpose of this study is to investigate the direct effects of hyperinsulinemia on EPCs and the underlying mechanisms. EPCs isolated from healthy adults were cultured with various concentrations of insulin (control group, without insulin; physiological insulin group, 10nM insulin and hyperinsulinemia group, 100nM insulin) with or without phosphatidylinositol-3-kinase (PI3-K) inhibitor (LY294002, 5µM), endothelial nitric oxide synthase (eNOS) inhibitor (L-NG-nitro-arginine methyl ester (L-NAME), 100µM), sodium nitroprusside (SNP, 25µM), p38 mitogen-activated protein kinase(MAPK) inhibitor (SB203580, 5µM) or extracellular signal-regulated kinases (ERK) 1/2 inhibitor (PD98059, 10µM). Proliferation, tube formation, and apoptosis of EPCs were determined. Expressions of eNOS, PI3-K, protein kinase B (Akt), p38 MAPK, and ERK 1/2 were assessed. Hyperinsulinemia caused a significant decrease in proliferation and tube formation abilities than control group. Hyperinsulinemia increased apoptosis rate of EPCs than control group. Furthermore, hyperinsulinemia downregulated phosphorylation of eNOS, PI3-K and Akt, and upregulated phosphorylation of p38 MAPK and ERK. SNP could restore impaired tube formation induced by hyperinsulinemia. P38 MAPK inhibitor but not ERK inhibitor could decrease apoptosis induced by hyperinsulinemia. Hyperinsulinemia impaired EPCs' tube formation ability by downregulation of PI-3K/Akt/eNOS pathway. Hyperinsulinemia induced apoptosis of EPCs via upregulation of p38 MAPK.

Endothelial progenitor cell impairment mediated vasodilation dysfunction via diminishing nitric oxide production in postmenopausal females.

Vascular endothelial dysfunction is the major contributing factor to hypertension. Endothelial progenitor cells (EPCs) are essential for endogenous vascular endothelial renovation. The activity and number of circulating EPCs are preserved in prehypertensive premenopausal females according to our previous research. However, the changes of EPCs in prehypertensive postmenopausal females are poorly understood, and the mechanisms responsible for the loss of the gender protection advantage of cardiovascular disease remain unexplored. In order to determine the effects of EPCs in prehypertensive postmenopausal females, the number and activity of circulating EPCs were tested in the present study. Next, the function of EPCs secreting nitric oxide (NO), vascular endothelial growth factor (VEGF) and granulocyte‑macrophage colony‑stimulating factor (GM‑CSF), as well as their concentration in the plasma, were measured. The association between flow‑mediated dilation (FMD) and EPC secretion was also assessed. Attenuation of proliferation and migration of EPCs was observed in prehypertensive patients in comparison with normotensive subjects. In addition, a reduced NO production secreted by EPCs was detected in prehypertensive patients as compared with that in normotensive patients. There was no significant difference in EPC function between postmenopausal females and age‑matched males. Finally, the association between FMD and NO production was validated. Collectively, these data indicated that impaired EPCs mediated vasodilation dysfunction via decreasing NO production. Therefore, EPC function enhancement and NO level augmentation are emerging as novel therapeutic strategies for prehypertension therapy.

Rosuvastatin is Superior Compared to Simvastatin and Atorvastatin to Induce Endothelial Progenitor Cells Migration

Introduction Statins have shown to improve Endothelial Progenitor Cells (EPCs) function, however no comparison has been done between various statins effectivity to induce EPCs migration. Aim This study compared simvastatin, atorvastatin and rosuvastatin treatment on impaired EPCs from Coronary Artery Disease (CAD) patients. Materials and Methods EPCs were isolated, cultivated and divided into untreated group (control), simvastatin (dose 0.1, 0.25, 0.5 mM), atorvastatin (0.1, 0.25, 0.5 mM), rosuvastatin (0.1, 0.25, 0.5 mM). EPCs migration was evaluated with boyden chamber assay. ANOVA, pearson correlation and linear regression test were done using SPSS 25.0. Results This research showed that simvastatin, atorvastatin, and rosuvastatin increased EPCs migration in dose dependent manner (p<0.05). Regression test showed that simvastatin treatment was responsible for 92.9% of EPCs migration, while atorvastatin was 75.5% and rosuvastatin was 65.6%. Rosuvastatin treatment dose 0.5 mM have the highest EPCs migration effect (195,750.00±5,809.48) compared to simvastatin (123,750.00±9,367.50, p≤0.001) and atorvastatin (156,375.00±12,392.03, p≤0.001) at the same dose. Conclusion Rosuvastatin treatment has higher EPCs migration effect compared to simvastatin and atorvastatin. This suggests that rosuvastatin might be preferable to improve EPCs migration in CAD patients.

Endothelial Progenitor Cells as Mediators of the Crosstalk between Vascular Repair and Immunity: Lessons from Systemic Autoimmune Diseases.

From the discovery of Endothelial Progenitor Cells (EPC), these bone marrowderived precursors have been placed as crucial mediators of the endothelial repair. Accordingly, altered levels and function of EPC have been found in different scenarios of CV risk. Despite the fact that EPC exhibit important characteristics which support a link of this cell subset with a number of inflammatory and immune networks, little is known on the actual mediators involved and the clinical relevance of these features. Systemic diseases are generally hallmarked by a vascular repair failure and increased cardiovascular disease occurrence, EPC impairment having a pivotal role. Because of their immunemediated etiology, this group of conditions represents an invaluable scenario to unravel the connections between immune dysregulation and EPC dysfunction. In the present review, we summarize the current knowledge regarding the cutting-edge area of the modulation of EPC levels and function by inflammatory cytokines in systemic diseases. We also address the possibility of the available immunomodulatory drugs to counteract this situation. Finally, due to the emerging role of the vitamin D as a common mediator in the immune system and the cardiovascular axis, we cover the topic of the role of vitamin D as a potential player in the inflammatory-mediated EPC dysfunction in systemic diseases.

Association between Type I interferon and depletion and dysfunction of endothelial progenitor cells in C57BL/6 mice deficient in both apolipoprotein E and Fas ligand

Patients with systemic lupus erythematosus (SLE) have a tremendously increased risk for cardiovascular disease (CVD), which could not be accounted in entirety by traditional Framingham risk factors. To study whether the accelerated atherosclerosis in SLE patients is mediated by type I interferon (IFN-I) through the regulation of endothelial progenitor cells (EPCs), we created a line of C57BL/6 mice with deficiency in both apolipoprotein E (ApoE−/−) and fas ligand (FasL−/−, gld.). As expected, the resultant gld. ApoE−/− mice exhibited both aggravated lupus-like disease and atherosclerosis under normal diet. Increased expression of IFN-I-stimulated genes (ISGs) was closely associated with depletion and dysfunction of EPCs, as well as with accelerated atherosclerotic lesion in gld. ApoE−/− mice. While only IFN-α instead of other interventions, including tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IRS423 and IRS661, impaired EPC function in vitro. Mechanistically, activation or inhibition of the TLR7/9 signaling could modulate EPC number and function in vivo. Decreased proliferation rate and increased apoptotic rate of EPCs induced by IFN-α might contribute to the results to a certain extent. Thus, our data suggest that excessive expression of IFN-I through the activation of TLR7/9 signaling may induce accelerated atherosclerosis in lupus through the depletion or dysfunction of EPCs, suggesting that targeting IFN-I might have potential therapeutic effects on both lupus disease and premature atherosclerosis in SLE patients.

Inhibition of macrophage inflammatory protein-1β improves endothelial progenitor cell function and ischemia-induced angiogenesis in diabetes.

Systemic inflammation might contribute to the impairment of neovasculogenesis and endothelial progenitor cell (EPC) function in clinical diabetes mellitus (DM). Macrophage inflammatory protein-1β (MIP-1β) is an inflammatory chemokine that may be up-regulated in clinical DM. Its role in diabetic vasculopathy was not clarified. This study aimed to investigate the role of MIP-1β in human EPCs and in neovasculogenesis in different diabetic animal models with hindlimb ischemia. EPCs chamber assay and in vitro tube formation assay were used to estimate the degree of EPC migration and tube formation abilities. Leprdb/JNarl mice, C57BL/6 mice fed a high-fat diet, and streptozotocin-induced diabetic mice were used as different diabetic animal models. Laser Doppler imaging and flow cytometry were used to evaluate the degree of neovasculogenesis and the circulating levels of EPCs, respectively. MIP-1β impaired human EPC function for angiogenesis in vitro. Plasma MIP-1β levels were up-regulated in type 2 DM patients. MIP-1β inhibition enhanced the function and the C-X-C chemokine receptor type 4 expression of EPCs from type 2 diabetic patients, and improved EPC homing for ischemia-induced neovasculogenesis in different types of diabetic animals. MIP-1β directly impaired human EPC function. Inhibition of MIP-1β improved in vitro EPC function, and enhanced in vivo EPC homing and ischemia-induced neovasculogenesis, suggesting the critical role of MIP-1β for vasculopathy in the presence of DM.

Impairment of bone marrow endothelial progenitor cells in acute graft-versus-host disease patients after allotransplant.

Graft-versus-host disease (GVHD) is a major complication after allogeneic haematopoietic stem cell transplantation (allo-HSCT) that is frequently associated with bone marrow (BM) suppression, and clinical management is challenging. BM endothelial progenitor cells (EPCs) play crucial roles in the regulation of haematopoiesis and thrombopoiesis. However, little is known regarding the functional roles of BM EPCs in acute GVHD (aGVHD) patients. In the current prospective case-control study, reduced and dysfunctional BM EPCs, characterized by decreased migration and angiogenesis capacities and increased levels of reactive oxygen species (ROS) and apoptosis, were found in aGVHD patients compared with those without aGVHD. Moreover, lower frequency and increased levels of ROS, apoptosis and DNA damage, but reduced colony-forming unit-plating efficiency were found in BM CD34+ cells of aGVHD patients compared with those without aGVHD. The severity of aGVHD and GVHD-mediated cytopenia was associated with BM EPC impairment in aGVHD patients. In addition, the EPC impairment positively correlated with ROS level. Taken together, our results suggest that reduced and dysfunctional BM EPCs may be involved in the pathogenesis of aGVHD. Although these findings require validation, our data indicate that improvement of BM EPCs may represent a promising therapeutic approach for aGVHD patients.

Melatonin Improves Neovascularization after Hind-Limb Ischemia Surgery in Streptozocin-Induced Diabetic Mice by Enhancing Endothelial Progenitor Cells Functions

Objective: Diabetic patients show poor endothelial progenitor cell (EPC) functions and are prone to ischemic micro-vascular or macro-vascular complications. Melatonin has been shown to have protective effects in ischemic injury, possibly in part by modulating the functioning of EBCs. We investigated whether pretreatment with melatonin can restore the impaired functions of EPCs in diabetes. Methods: Human EPC were isolated and cultured in a high-glucose medium for functional testing. Cell proliferation, nitric oxide (NO) production, and apoptosis assay were examined. A streptozocin (STZ)-induced diabetic mouse model was established to evaluate the actions of chronic hyperglycemia on ischemia-induced blood flow recovery. The circulating EPC number in the peripheral blood was determined by flow cytometry (Sca-1+/Flk-1+). Results: Incubation with a high-glucose medium (25 mM) significantly suppressed EPC proliferation, reduced NO production, and lessened phosphorylations of Akt and eNOS. Moreover, EPC treated with high-glucose medium increased reactive oxygen species production, promoted cellular apoptosis and senescence, and also inhibited EPC tube formation. Treatment of melatonin could recover these harmful effects. Four weeks after hindlimb ischemia surgery, the STZ-induced diabetic mice had significantly reduced tissue reperfusion, EPC mobilization, and impaired neovascularization in the ischemic hindlimbs compared with the nondiabetic mice. In STZ-induced diabetic mice, the melatonin group showed significantly increased ischemic/non-ischemic limb blood perfusion ratios, higher capillary densities, and improved EPC mobilization. Conclusion: Chronic hyperglycemia impaired blood flow recovery and EPC mobilization in response to tissue ischemia, and pretreatment of melatonin could reduce these effects. Disclosure C. Kuo: None. P. Huang: None. S. Lin: None.

Atorvastatin enhances bone marrow endothelial cell function in corticosteroid-resistant immune thrombocytopenia patients

AbstractThe pathogenesis of corticosteroid-resistant immune thrombocytopenia (ITP), a clinically challenging condition in which patients exhibit either no response to corticosteroids or are corticosteroid-dependent, remains poorly understood. Murine studies suggest that bone marrow (BM) endothelial progenitor cells (EPCs) play a crucial role in regulating megakaryocytopoiesis. However, little is known regarding the number and function of BM EPCs or how to improve impaired BM EPCs in corticosteroid-resistant ITP patients. In the current case-control study, we evaluated whether the BM EPCs in corticosteroid-resistant ITP differed from those in corticosteroid-sensitive ITP. Moreover, whether atorvastatin could enhance the number and function of BM EPCs derived from corticosteroid-resistant ITP patients was investigated in vitro and in vivo. Reduced and dysfunctional BM EPCs, characterized by decreased capacities of migration and angiogenesis as well as higher levels of reactive oxygen species and apoptosis, were observed in corticosteroid-resistant ITP patients. In vitro treatment with atorvastatin quantitatively and functionally improved BM EPCs derived from corticosteroid-resistant ITP patients by downregulating the p38 MAPK pathway and upregulating the Akt pathway, and rescued the impaired BM EPCs to support megakaryocytopoiesis. Subsequently, a pilot cohort study showed that atorvastatin was safe and effective in corticosteroid-resistant ITP patients. Taken together, these results indicate that reduced and dysfunctional BM EPCs play a role in the pathogenesis of corticosteroid-resistant ITP, and the impaired BM EPCs could be improved by atorvastatin both in vitro and in vivo. Although requiring further validation, our data indicate that atorvastatin represents a promising therapeutic approach for repairing impaired BM EPCs in corticosteroid-resistant ITP patients.

N-acetyl-L-cysteine improves bone marrow endothelial progenitor cells in prolonged isolated thrombocytopenia patients post allogeneic hematopoietic stem cell transplantation.

Prolonged isolated thrombocytopenia (PT) is a serious complication following allogeneic hematopoietic stem cell transplantation (allo-HSCT). According to murine studies, endothelial progenitor cells (EPCs) play a crucial role in the regulation of hematopoiesis and thrombopoiesis in the bone marrow (BM) microenvironment. We previously showed that the reduced frequency of BM EPCs was an independent risk factor for the occurrence of PT following allo-HSCT. However, the functional role of BM EPCs and methods to improve the impaired BM EPCs in PT patients are unknown. In the current case-control study, we investigated whether the BM EPCs in PT patients differed from those in good graft function patients. Moreover, we evaluated whether N-acetyl-L-cysteine (NAC, a reactive oxygen species [ROS] scavenger) could enhance BM EPCs from PT patients in vitro and in vivo. The PT patients exhibited dysfunctional BM EPCs characterized by high levels of ROS and apoptosis and decreased migration and angiogenesis capabilities. In vitro treatment with NAC improved the quantity and function of the BM EPCs cultivated from the PT patients by downregulating the p38 MAPK pathway and rescued the impaired BM EPCs to support megakaryocytopoiesis. Furthermore, according to the results of a preliminary clinical study, NAC is safe and effective in PT patients. In summary, these results suggested that the reduced and dysfunctional BM EPCs are involved in the occurrence of PT. The defective BM EPCs in the PT patients can be quantitatively and functionally improved by NAC, indicating that NAC is a promising therapeutic approach for PT patients following allo-HSCT.

Chronic hyperuricemia impairs blood flow recovery in the ischemic hindlimb through suppression of endothelial progenitor cells.

ObjectiveChronic hyperuricemia is associated with cardiovascular disease, but its impact on endothelial progenitor cells (EPC) and ischemia-induced neovascularization remains unclear. Herein we investigated whether chronic hyperuricemia could impede blood flow recovery in response to tissue ischemia by suppression of EPC.MethodsHuman EPC were isolated and cultured in a high-level uric acid medium for functional testing. Cell proliferation, nitric oxide (NO) production and apoptosis assay were examined. A chronic hyperuricemia mouse model was established by potassium oxonate treatment and/or a high-level uric acid diet to evaluate the actions of chronic hyperuricemia on ischemia-induced blood flow recovery. After 4 weeks of drug treatment, hindlimb ischemia surgery was performed in the control and hyperuricemia mice. Blood flow recovery was followed up every week before and after ischemic surgery using a laser Doppler Perfusion Imager System. The circulating EPC number in the peripheral blood was determined by flow cytometry (Sca-1+/Flk-1+).ResultsIncubation with a high-level uric acid medium (10 mg/dL) significantly suppressed EPC proliferation, reduced NO production, and lessened phosphorylation of Akt and eNOS. Moreover, EPC treated with high-level uric acid increased reactive oxygen species production, promoted cellular apoptosis and senescence, and also inhibited EPC tube formation. Four weeks after hindlimb ischemia surgery, the chronic hyperuricemia mice had significantly reduced tissue reperfusion, EPC mobilization, and impaired neovascularization in the ischemic hindlimbs compared with the control mice.ConclusionsChronic hyperuricemia impaired blood flow recovery and EPC mobilization in response to tissue ischemia, and these effects could have occurred through suppression of EPC.

Puerarin protects endothelial progenitor cells from damage of angiotensin II via activation of ERK1/2‑Nrf2 signaling pathway.

Endothelial progenitor cell (EPC) dysfunction is associated with the formation of carotid atherosclerosis. It has been demonstrated that angiotensin II (AngII) may impair the function of EPCs and puerarin, a natural product, possesses cardiovascular protective effects against oxidative stress and inflammation. Therefore, the present study aimed to investigate the beneficial effects of puerarin in AngII‑induced EPC injury, and to elucidate the underlying mechanisms. Treatment with AngII suppressed EPC proliferation and migration, increased the expression of the senescence marker β‑galactosidase, and the adhesion molecules intracellular adhesion molecule‑1 and vascular cell adhesion molecule‑1. However, the above effects were markedly alleviated by treatment with puerarin in a dose‑dependent manner (1, 10 and 100µM). In addition, AngII significantly increased reactive oxygen species production and the levels of the inflammatory cytokine tumor necrosis factor‑α and interleukin‑6. Notably, these effects were reversed by puerarin. However, it was identified that the impaired EPC functions were due to inhibition of the phosphorylation of extracellular signal‑regulated kinase 1 and 2 (ERK1/2) and the degradation of nuclear factor erythroid 2 like 2 (Nrf2), and treatment with puerarin activated the ERK1/2‑Nrf2 signaling pathway. The results of the present study indicated that puerarin protected AngII‑induced EPC dysfunction via activation of the ERK1/2‑Nrf2 signaling pathway.

Effect of vitamin D on endothelial progenitor cells function

BackgroundEndothelial progenitor cells (EPCs) are a population of bone marrow-derived cells, which have an important role in the process of endothelialization and vascular repair following injury. Impairment of EPCs, which occurs in patients with diabetes, was shown to be related to endothelial dysfunction, coronary artery disease (CAD) and adverse clinical outcomes. Recent evidence has shown that calcitriol, the active hormone of vitamin D, has a favorable impact on the endothelium and cardiovascular system. There is limited data on the effect of vitamin D on EPCs function.AimTo examine the in vitro effects of Calcitriol on EPCs from healthy subjects and patients with diabetes.MethodsFifty-one patients with type 2 diabetes (60±11 years, 40% women, HbA1C: 9.1±0.8%) and 23 healthy volunteers were recruited. EPCs were isolated and cultured with and without calcitriol. The capacity of the cells to form colony-forming units (CFUs), their viability (measured by MTT assay), KLF-10 levels and angiogenic markers were evaluated after 1 week of culture.ResultsIn diabetic patients, EPC CFUs and cell viability were higher in EPCs exposed to calcitriol vs. EPCs not exposed to calcitriol [EPC CFUs: 1.25 (IQR 1.0–2.0) vs. 0.5 (IQR 0.5–1.9), p < 0.001; MTT:0.62 (IQR 0.44–0.93) vs. 0.52 (IQR 0.31–0.62), p = 0.001]. KLF-10 levels tended to be higher in EPCs exposed to vitamin D, with no differences in angiopoietic markers. In healthy subjects, calcitriol supplementation also resulted in higher cell viability [MTT: 0.23 (IQR 0.11–0.46) vs. 0.19 (0.09–0.39), p = 0.04], but without differences in CFU count or angiopoietic markers.ConclusionIn patients with diabetes mellitus, in vitro vitamin D supplementation improved EPCs capacity to form colonies and viability. Further studies regarding the mechanisms by which vitamin D exerts its effect are required.