Blood Endothelial Progenitor Cells Research Articles

Highly-Expressed MiR-221-3p Distinctly Increases the Incidence of Diabetic Retinopathy in Patients With Type 2 Diabetes Mellitus.

Diabetic retinopathy (DR) is the leading cause of blindness in patients with diabetes mellitus (DM). MiR-221-3p is implicated in microvascular dysfunction in DR, and we explored their relationship. Patients with type 2 diabetes mellitus (T2DM) were allocated to the non-DR (NDR)/nonproliferative DR (NPDR)/proliferative DR (PDR) groups, with their clinical baseline and pathological data collected. The miR-221-3p and VEGF levels were determined by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and ELISA, respectively. Peripheral blood endothelial progenitor cell (EPC) and endothelial cell (EC) ratios were ascertained by flow cytometry. The correlations between miR-221-3p levels and VEGF/EPCs/ECs, the predictive value of serum miR-221-3p levels in DR, and the independent risk factors for DR occurrence in T2DM were analyzed by Pearson's correlation analysis, receiver operating characteristic (ROC) curve, and multifactorial logistic regression analysis. Serum miR-221-3p was highly expressed in DR. Clinical severity of DR was positively correlated with miR-221-3p levels. Endothelial function was impaired in DR. Serum miR-221-3p levels in DR were favorably correlated with VEGF and ECs and negatively associated with EPCs. The area under the curve of serum miR-221-3p in evaluating DR occurrence in patients with T2DM was 0.8178 (1.235 cutoff value, 69.62% sensitivity, and 82.35% specificity). High expression of miR-221-3p increased DR incidence in patients with T2DM. Diabetes course, VEGF, EPCs, ECs, and miR-221-3p levels were independent risk factors for DR development in patients with T2DM. Serum miR-221-3p levels in patients with DR were positively correlated with VEGF and ECs and negatively linked with EPCs. Highly expressed miR-221-3p distinctly increased DR incidence in patients with T2DM and was an independent risk factor for DR development in patients with T2DM. This study assessed serum miR-221-3p level and endothelial function indicators (VEGF, EPCs, and ECs) in patients with DR and analyzed the correlation between each indicator. We found that high serum miR-221-3p expression prominently increased the incidence of DR in patients with T2DM and was an independent risk factor for the development of DR in patients with T2DM. This study provided a scientific basis for further clarification of the pathogenesis of DR, and also provided new ideas for clinical prediction and management of DR.

RNA N6‑methyladenosine methyltransferase WTAP promotes the differentiation of endothelial progenitor cells.

N6-methyladenosine (m6A) serves a critical role in regulating gene expression and has been associated with various diseases; however, its role in the differentiation of endothelial progenitor cells (EPCs) remains unclear. The present study used liquid chromatography with tandem mass spectrometry and immunofluorescence assays to quantify the levels of m6A in human peripheral blood-derived EPCs (HPB-EPCs) before and after differentiation into mature cells. The present study performed Cell Counting Kit 8, Transwell, and tube formation assays to determine the effects of overexpression and knockdown of Wilms' tumor 1-associated protein (WTAP) on HPB-EPCs. The results revealed that the level of m6A modification was significantly increased during HPB-EPCs differentiation, and WTAP exhibited the most significant alteration among the enzymes involved in m6A regulation. When WTAP was overexpressed in HPB-EPCs, cell proliferation, invasion, and the formation of tubes were improved, whereas WTAP knockdown yielded the opposite effects. In conclusion, the present study highlighted the involvement of m6A in regulating EPC differentiation, with WTAP acting as a promoter of EPC differentiation.

Verification of Therapeutic Effect through Accelerator Mass Spectrometry-Based Single Cell Level Quantification of hESC-Endothelial Cells Distributed into an Ischemic Model.

As the potential of pluripotent stem cell-derived differentiated cells has been demonstrated in regenerative medicine, differentiated vascular endothelial cells (ECs) are emerging as a therapeutic agent for the cardiovascular system. To verify the therapeutic efficacy of differentiated ECs in an ischemic model, human embryonic stem cells (hESCs) are induced as EC lineage and produce high-purity ECs through fluorescence-activated cell sorting (FACS). When hESC-ECs are transplanted into a hindlimb ischemic model, it is confirmed that blood flow and muscle regeneration are further improved by creating new blood vessels together with autologous ECs than the primary cell as cord blood endothelial progenitor cells (CB-EPCs). In addition, previously reported studies show the detection of transplanted cells engrafted in blood vessels through various tracking methods, but fail to provide accurate quantitative values over time. In this study, it is demonstrated that hESC-ECs are engrafted approximately sevenfold more than CB-EPCs by using an accelerator mass spectrometry (AMS)-based cell tracking technology that can perform quantification at the single cell level. An accurate quantification index is suggested. It has never been reported in in vivo kinetics of hESC-ECs that can act as therapeutic agents.

Cord Blood–Derived Endothelial Progenitor Cells Promote In Vivo Regeneration of Human Hematopoietic Bone Marrow

Victims of acute radiation exposure are susceptible to hematopoietic toxicity due to bone marrow damage and loss of mature blood elements. Here, we evaluated cord blood-derived endothelial progenitor cells (CB-EPCs) as a potential cellular therapy for mitigation of hematologic acute radiation syndrome. CB-EPCs express endothelial cell markers and maintain their growth characteristics beyond 10+ passages without diminishing their doubling capacity. Further, CB-EPCs can be cryopreserved in vapor-phase liquid nitrogen and easily recovered for propagation, making them an attractive nonimmunogenic cellular therapy for off-the-shelf use. Importantly, we show CB-EPCs have the capacity to potently expand adult human bone marrow hematopoietic progenitor cells both in vitro and in vivo. To demonstrate the role of CB-EPCs in promoting in vivo human immune reconstitution after irradiation, we employed a novel humanized mouse model established by transplant of CD34+ bone marrow cells from 9 unique adult organ donors into immunocompromised NSG-SGM3 mice. The response of the humanized immune system to ionizing irradiation was then tested by exposure to 1 Gy followed by subcutaneous treatment of CB-EPCs, Food and Drug Administration-approved growth factor pegfilgrastim (0.3 mg/kg), or saline. At day 7, total human bone marrow was decreased by 80% in irradiated controls. However, treatment with either growth factor pegfilgrastim or CB-EPCs increased recovery of total human bone marrow by 2.5-fold compared with saline. Notably, CB-EPCs also increased recovery of both human CD34+ progenitors by 5-fold and colony-forming capacity by 3-fold versus saline. Additionally, CB-EPCs promoted recovery of endogenous bone marrow endothelial cells as observed by both increased vessel area and length compared with saline. These findings indicate the feasibility of using humanized mice engrafted with adult bone marrow for radiation research and the development of CB-EPCs as an off-the-shelf cellular therapy for mitigation of hematologic acute radiation syndrome.

Human Cord Blood-Endothelial Progenitor Cells Alleviate Intimal Hyperplasia of Arterial Damage in a Rat Stroke Model.

Human cord blood-endothelial progenitor cells (hCB-EPCs) isolated from the human umbilical cord can be used to repair damaged arteries. In this study, we used an animal model with pathological changes that mimics artery wall damage caused by stent retrievers in humans. We injected hCB-EPCs to investigate their effect on endothelial hyperplasia and dysfunction during intimal repair. Four groups were established based on the length of reperfusion (3 and 28 days), as well as the presence or absence of hCB-EPC therapy. Damage to the internal carotid artery was evaluated by hematoxylin-eosin and immunohistochemical staining. Stroke volume was not significantly different between non-EPC and EPC groups although EPC treatment alleviated intimal hyperplasia 28 days after intimal damage. Vascular endothelial growth factor (VEGF) and eNOS expression were significantly higher in the EPC-treated group than in the non-EPC group 3 days after intimal damage. In addition, MMP9 and 4HNE expression in the EPC-treated group was significantly lower than in the non-EPC group. Ultimately, this study found that venous transplantation of hCB-EPCs could inhibit neointimal hyperplasia, alleviate endothelial dysfunction, suppress intimal inflammation, and reduce oxidative stress during healing of intimal damage.

Biofunctionalization of cardiovascular stents to induce endothelialization: Implications for in- stent thrombosis in diabetes.

Stent thrombosis remains one of the main causes that lead to vascular stent failure in patients undergoing percutaneous coronary intervention (PCI). Type 2 diabetes mellitus is accompanied by endothelial dysfunction and platelet hyperactivity and is associated with suboptimal outcomes following PCI, and an increase in the incidence of late stent thrombosis. Evidence suggests that late stent thrombosis is caused by the delayed and impaired endothelialization of the lumen of the stent. The endothelium has a key role in modulating inflammation and thrombosis and maintaining homeostasis, thus restoring a functional endothelial cell layer is an important target for the prevention of stent thrombosis. Modifications using specific molecules to induce endothelial cell adhesion, proliferation and function can improve stents endothelialization and prevent thrombosis. Blood endothelial progenitor cells (EPCs) represent a potential cell source for the in situ-endothelialization of vascular conduits and stents. We aim in this review to summarize the main biofunctionalization strategies to induce the in-situ endothelialization of coronary artery stents using circulating endothelial stem cells.

Fetal sex and maternal fasting glucose affect neonatal cord blood-derived endothelial progenitor cells

BackgroundMaternal cardiovascular risk factors (CVRF) in pregnancy, i.e., obesity and hyperglycemia, transmit to the fetus and affect placental and fetal endothelial function. Moreover, a sex dimorphism in endothelial function and susceptibility towards CVRF exists already in utero. Endothelial colony-forming cells (ECFC) are circulating endothelial progenitors highly present in neonatal cord blood and sensitive to CVRF. This study investigated whether fetal sex or subtle maternal metabolic changes within healthy range alter fetal ECFC outgrowth.MethodsOutgrowth of ECFC from cord blood of male (n = 31) and female (n = 26) neonates was analyzed after healthy pregnancies and related to fetal sex and maternal metabolic parameters.ResultsMale ECFC grew out earlier (−20.57% days; p = 0.031) than female. Although all women were non-diabetic, higher levels of fasting plasma glucose (FPG) at midpregnancy increased the time required for colony outgrowth (OR: 1.019; p = 0.030), which, after stratifying for fetal sex, was significant only in the males. Gestational weight gain and BMI did not affect outgrowth. Colony number was unchanged by all parameters.ConclusionsFetal sex and maternal FPG within normal range alter ECFC function in utero. A role of ECFC in postnatal angiogenesis and vasculogenesis has been suggested, which may be affected by altered outgrowth dynamics.ImpactThis study is the first to report that a sexual dimorphism exists in ECFC function, as cells of female progeny require a longer period of time until colony outgrowth than ECFC of male progeny.Our data show that ECFC function is highly sensitive and affected by maternal glucose levels even in a normal, non-diabetic range.Our data raise the question of whether maternal plasma glucose in pregnancy should be considered to play a critical role even in the non-diabetic setting.

Microvascular Permeability and Texture Analysis of the Skeletal Muscle of Diabetic Rabbits With Critical Limb Ischaemia Based on DCE-MRI

BackgroundWe evaluated skeletal muscle vascular permeability in diabetic rabbits with critical limb ischaemia using quantitative dynamic contrast agent-enhanced (DCE) magnetic resonance imaging (MRI) and explored the feasibility of using DCE-MRI Ktrans-based texture analysis for assessing early slight ischaemia-related skeletal muscle structural changes.MethodTwenty-four male New Zealand white rabbits (2.7 ± 0.3 kg; n = 12 each in sham-operated and experimental groups) underwent axial MRI of the vastus lateralis muscle at 1, 2, and 3 weeks after alloxan injection. Between-group and intra-group postoperative permeability and texture parameters were compared. Texture features of experimental groups in the third week were modelled by receiver operating characteristic (ROC) curve analysis. Correlations of permeability and of statistical texture parameters with peripheral blood endothelial progenitor cells (EPCs) and microvascular density (MVD) were analysed.ResultsIn the experimental group, the transfer constant (Ktrans) was statistically significant at all time-points (F = 5.800, P = 0.009). Their vastus lateralis muscle Ktrans was significantly lower in the third than in the first week (P = 0.018) and correlated positively with peripheral blood EPCs in the experimental group [r = 0.598, (95% CI: 0.256, 0.807)]. The rate constant was negatively associated with vastus lateralis muscle MVD [r = -0.410, (95% CI: -0.698, -0.008)]. The area under the ROC curve of texture parameters based on Ktrans in ischaemic limbs was 0.882.ConclusionsQuantitative DCE-MRI parameters could evaluate microvascular permeability of ischaemic limb skeletal muscle, and texture analysis based on DCE-MRI Ktrans allowed evaluation of early slight skeletal muscle structural changes.

Endothelial Progenitor Cell Levels and Extent of Post-prandial Lipemic Response.

Background and ObjectiveDue to the frequency of meal ingestion, individuals spend the majority of the day, ~18 h, in a status of post-prandial (PP) stress. Remnant-like lipoprotein particles (RLPs) are predominant in PP phase playing an important role in the development of atherosclerosis. Endothelial progenitor cells (EPCs) have been suggested to play a role in vessel wall homeostasis and in reducing atherosclerosis. However, there is no information about peripheral blood EPCs number following PP stress. We investigated the association between circulating EPCs levels and extent of PP lipemia in healthy subjects following a high-fat meal.Materials and MethodsA total of 84 healthy subjects (42 men, 42 women) aged 17–55 years were included in the study. PP lipemic response of subjects was determined by Oral Fat-Loading Test (OFLT). All the subjects were classified on the basis of their plasma TG levels after PP lipemic stressors in categories 1 (low), 2 (moderate), and 3 (high). Circulating EPCs numbers were measured by the flow cytometry method.ResultsThere was a significant difference in terms of lipid parameters between men and women: high-density lipoprotein cholesterol (HDL-C) was significantly lower in men than in women (p < 0.001). Total cholesterol (TC) (p = 0.004), low-density lipoprotein cholesterol (LDL-C) (p < 0.001), triglyceride (TG) (p < 0.001), and TG-AUC (p < 0.001) were significantly higher in men than in women. There was no significant difference between the genders in terms of CD34+KDR+ and CD34+KDR+CD133+cell number and MMP-9 levels. Vascular endothelial growth factor (VEGF) levels were significantly higher in men than women (p = 0.004). TC, LDL-C, and TG were significantly higher in the 3rd category than 1st and 2nd categories (p < 0.001) in women. Age, body mass index (BMI), fat rate, TG, TC, and LDL-C were significantly higher in the 3rd category than 1st category (p < 0.001, p = 0.002, p = 0.002, p = 0.01, p = 0.007, p = 0.004; respectively), in men. Circulating numbers of EPCs in men were significantly higher in the PP hyperlipidemia group than in the low TG levels category, independently from age (p < 0.05). Circulating EPC levels showed a positive correlation with OFLT response in men (r = 0.414, p < 0.05). Also, OFLT response showed a strong positive correlation with fasting TG levels (r = 0.930, p < 0.001). EPC levels in categories of women were not different.ConclusionIncreased EPCs levels in subjects with different PP hyperlipidemia may be associated with a response to endothelial injury, related to increased atherogenic remnant particles at the PP phase.

A combined cell and growth factor delivery for the repair of a critical size tibia defect using biodegradable hydrogel implants.

The ability to repair critical‐sized long‐bone injuries using growth factor and cell delivery was investigated using hydrogel biomaterials. Physiological doses of the recombinant human bone morphogenic protein‐2 (rhBMP2) were delivered in a sustained manner from a biodegradable hydrogel containing peripheral human blood‐derived endothelial progenitor cells (hEPCs). The biodegradable implants made from polyethylene glycol (PEG) and denatured fibrinogen (PEG‐fibrinogen, PF) were loaded with 7.7 μg/ml of rhBMP2 and 2.5 × 106 cells/ml hEPCs. The safety and efficacy of the implant were tested in a rodent model of a critical‐size long‐bone defect. The hydrogel implants were formed ex‐situ and placed into defects in the tibia of athymic nude rats and analyzed for bone repair after 13 weeks following surgery. The hydrogels containing a combination of 7.7 μg/ml of rhBMP2 and 2.5 × 106 cells/ml hEPCs were compared to control hydrogels containing 7.7 μg/ml of rhBMP2 only, 2.5 × 106 cells/ml hEPCs only, or bare hydrogels. Assessments of bone repair include histological analysis, bone formation at the site of implantation using quantitative microCT, and assessment of implant degradation. New bone formation was detected in all treated animals, with the highest amounts found in the treatments that included animals that combined the PF implant with rhBMP2. Moreover, statistically significant increases in the tissue mineral density (TMD), trabecular number and trabecular thickness were observed in defects treated with rhBMP2 compared to non‐rhBMP2 defects. New bone formation was significantly higher in the hEPC‐treated defects compared to bare hydrogel defects, but there were no significant differences in new bone formation, trabecular number, trabecular thickness or TMD at 13 weeks when comparing the rhBMP2 + hEPCs‐treated defects to rhBMP2‐treated defects. The study concludes that the bone regeneration using hydrogel implants containing hEPCs are overshadowed by enhanced osteogenesis associated with sustained delivery of rhBMP2.

Effects of Shen-Yuan-Dan on Periprocedural Myocardial Injury and the Number of Peripheral Blood Endothelial Progenitor Cells in Patients with Unstable Angina Pectoris Undergoing Elective Percutaneous Coronary Intervention.

Objectives We aimed to investigate the effects of Shen-Yuan-Dan (SYD), a Chinese medicine preparation, on periprocedural myocardial injury (PMI) and the number of peripheral blood endothelial progenitor cells (EPCs) in patients with unstable angina pectoris (UA) who underwent elective percutaneous coronary intervention (PCI). Methods Patients were randomly divided into the experimental (group A) and control (group B) groups through the random number table method. In group A, patients concurrently received the conventional western treatment and SYD orally (4 capsules/time, 3 times/d, from 3 d before surgery to 7 d after surgery). In group B, patients received conventional Western medicine treatment. Both groups underwent coronary angiography, and patients undergoing PCI were eventually included in the study. The following patient data were collected: incidence of PMI, serum CK-MB content before PCI, 4 h, 24 h, and 7 d after PCI, number of CD45dim/-CD34+CD309+ peripheral venous EPCs, and number of CD184 coexpressed EPCs. The incidence of adverse reactions and 30-day major adverse cardiovascular events (MACEs) were also recorded. Results Sixty-two patients were finally included in this study, with 32 and 30 in groups A and B, respectively. In group A, the number of peripheral blood EPCs and the number of CD184 coexpressed EPCs at 1 h before surgery were higher than those at 3 d before surgery (37.24 ± 25.20 vs. 22.78 ± 9.60/ml; P < 0.001 and 23.38 ± 15.30 vs. 13.54 ± 8.08/ml; P < 0.001, resp.). The number of peripheral blood EPCs and number of CD184 coexpressed EPCs at 4 h after surgery were lower than those at 1 h before surgery (25.30 ± 11.90 vs. 37.24 ± 25.20/ml; P=0.019 and 15.38 ± 8.78 vs. 23.38 ± 15.30/ml; P=0.013, resp.), but there was no difference at 24 h and at 7 d after surgery in comparison with that at 1 h before surgery (P > 0.05). In group B, compared with that at 1 h before surgery, there existed a decline in the number of EPCs in peripheral blood and the number of CD184 coexpressed EPCs at 4 h after surgery, but without a statistical difference (P > 0.05). Comparing both groups, it was found that the incidence of PMI in group A was lower (6.25% vs. 26.67%; P=0.04), and the serum CK-MB content at 4 and 24 h after surgery was also lower than that in group B (17.33 ± 5.83 vs. 20.38 ± 4.32 U/l; P=0.048 and 15.79 ± 5.32 vs. 19.10 ± 4.93 U/l; P=0.030, resp.). The number of EPCs in peripheral blood and the number of CD184 coexpressed EPCs in group A were higher than those in group B at 1 h before surgery (37.24 ± 25.20 vs. 22.36 ± 12.26/ml; P=0.034 and 23.38 ± 15.30 vs. 13.12 ± 14.62/ml; P=0.013, resp.). In addition, there were no obvious adverse reactions and no 30-day MACEs in both groups during the trial. Conclusion SYD can reduce PMI and promote the mobilization of EPCs in the perioperative period of elective PCI in patients with UA.

The impact of cardiomotor rehabilitation on endothelial function in elderly patients with chronic heart failure

BackgroundTo explore the effects of cardiac exercise rehabilitation on peripheral blood endothelial progenitor cells (EPC) in elderly patients with chronic heart failure.Methods80 elderly patients with chronic heart failure were selected from March 2017 to March 2019 and randomly divided into two groups (N = 40). The control group was treated routinely and walked freely for 30–60 min every day. The patients in the exercise rehabilitation group developed a cardiac exercise rehabilitation plan. Then, cardiac function and peripheral blood B-natriuretic peptide (BNP) levels in the two groups were compared. The cell viability, proliferation, apoptosis, and invasion ability of EPCs were detected. The levels of the PI3K/AKT pathway and eNOS and VEGF were compared.ResultsThere were no significant differences in all indexes between the two groups before treatment (P > 0.05), and both improved significantly after treatment (P < 0.05). After treatment, LVEF and LVFS in the exercise rehabilitation group were significantly higher than those in the control group (P < 0.05), and LVEDD and LVESD were significantly lower than those in the control group (P < 0.05). The BNP level in the exercise rehabilitation group was significantly lower than that in the control group (P < 0.05). The cell viability, proliferation, invasion ability of EPC, and the levels of PI3K, AKT, eNOS, and VEGF mRNA and protein in the exercise rehabilitation group were significantly higher than those in the control group. Apoptosis rate was significantly lower than those in the control group (P < 0.05).ConclusionsVisceral exercise rehabilitation can improve cardiac ejection and myocardial function in elderly patients with chronic heart failure, and can promote the vitality, proliferation, and invasion of peripheral blood EPC, and promote the expression of eNOS and VEGF by upregulating the PI3K/AKT pathway to promote angiogenesis and endothelial function.

Bradykinin Protects Human Endothelial Progenitor Cells from High-Glucose-Induced Senescence through B2 Receptor-Mediated Activation of the Akt/eNOS Signalling Pathway.

Background Circulating endothelial progenitor cells (EPCs) play important roles in vascular repair. However, the mechanisms of high-glucose- (HG-) induced cord blood EPC senescence and the role of B2 receptor (B2R) remain unknown. Methods Cord blood samples from 26 patients with gestational diabetes mellitus (GDM) and samples from 26 healthy controls were collected. B2R expression on circulating CD34+ cells of cord blood mononuclear cells (CBMCs) was detected using flow cytometry. The plasma concentrations of 8-isoprostaglandin F2α (8-iso-PGF2α) and nitric oxide (NO) were measured. EPCs were treated with HG (40 mM) alone or with bradykinin (BK) (1 nM). The B2R and eNOS small interfering RNAs (siRNAs) and the PI3K antagonist LY294002 were added to block B2R, eNOS, and PI3K separately. To determine the number of senescent cells, senescence-associated β-galactosidase (SA-β-gal) staining was performed. The level of mitochondrial reactive oxygen species (ROS) in EPCs was assessed by Mito-Sox staining. Cell viability was evaluated by Cell Counting Kit-8 (CCK-8) assays. Mitochondrial DNA (mtDNA) copy number and the relative length of telomeres were detected by real time-PCR. The distribution of human telomerase reverse transcriptase (hTERT) in the nucleus, cytosol, and mitochondria of EPCs was detected by immunofluorescence. The expression of B2R, p16, p21, p53, P-Ser473AKT, T-AKT, eNOS, and hTERT was demonstrated by Western blot. Results B2R expression on circulating CD34+ cells of CBMCs was significantly reduced in patients with GDM compared to healthy controls. Furthermore, B2R expression on circulating CD34+ cells of CBMCs was inversely correlated with plasma 8-iso-PGF2α concentrations and positively correlated with plasma NO levels. BK treatment decreased EPC senescence and ROS generation. Furthermore, BK treatment of HG-exposed cells led to elevated P-Ser473AKT and eNOS protein expression compared with HG treatment alone. BK reduced hTERT translocation in HG-induced senescent EPCs. B2R siRNA, eNOS siRNA, and antagonist of the PI3K signalling pathway blocked the protective effects of BK. Conclusion BK, acting through PI3K-AKT-eNOS signalling pathways, reduced hTERT translocation, increased the relative length of telomeres while reducing mtDNA copy number, and finally protected against EPC senescence induced by HG.

P300/Sp1-Mediated High Expression of p16 Promotes Endothelial Progenitor Cell Senescence Leading to the Occurrence of Chronic Obstructive Pulmonary Disease

Objective Chronic obstructive pulmonary disease (COPD) is a common chronic disease and develops rapidly into a grave public health problem worldwide. However, what exactly causes the occurrence of COPD remains largely unclear. Here, we are trying to explore whether the high expression of p16 mediated by p300/Sp1 can cause chronic obstructive pulmonary disease through promoting the senescence of endothelial progenitor cells (EPCs). Methods Peripheral blood EPCs were isolated from nonsmoking non-COPD, smoking non-COPD, and smoking COPD patients. The expressions of p16, p300, and senescence-related genes were detected by RT-PCR and Western Blot. Then, we knocked down or overexpressed Sp1 and p300 and used the ChIP assay to detect the histone H4 acetylation level in the promoter region of p16, CCK8 to detect cell proliferation, flow cytometry to detect the cell cycle, and β-galactosidase staining to count the proportion of senescent cells. Results The high expression of p16 was found in peripheral blood EPCs of COPD patients; the cigarette smoke extract (CSE) led to the increase of p16. The high expression of p16 in EPCs promoted cell cycle arrest and apoptosis. The CSE-mediated high expression of p16 promoted cell senescence. The expression of p300 was increased in peripheral blood EPCs of COPD patients. Moreover, p300/Sp1 enhanced the histone H4 acetylation level in the promoter region of p16, thereby mediating the senescence of EPCs. And knockdown of p300/Sp1 could rescue CSE-mediated cell senescence. Conclusion p300/Sp1 enhanced the histone H4 acetylation level in the p16 promoter region to mediate the senescence of EPCs.

Advances of Endothelial Progenitor Cells in the Development of Depression.

Depression is a major psychological disease of human beings. With the severity of depression, it elevates the risk of cardiovascular disease (CVD), especially acute coronary syndrome (ACS), resulting in serious harm to human health. The number of endothelial progenitor cells (EPCs) is closely related to the development of depression. It has been reported that the number of peripheral blood EPCs in patients with depression was reduced. However, effects on the function of EPCs in depression are still unclear. This paper aims to analyze and summarize the research of EPCs in depression, and we envision that EPCs might act as a new target for evaluating the severity of depression and its complications.

Endothelial Colony Forming Cells from Individuals with Low VWF Levels Demonstrate Impairment in Stimulated VWF Release

BackgroundVon Willebrand factor (VWF) is an essential hemostatic protein and deficiencies of VWF predispose to mucocutaneous bleeding (MCB) such as seen in von Willebrand disease (VWD). VWD type 1 is characterized by incomplete penetrance and variable expressivity even in individuals and families with the same mutation in VWF . In addition to those patients with VWD type 1, there is a large subset of individuals that have VWF levels below 50 IU/dL, who suffer from MCB but do not meet the full criteria for VWD (VWF levels less than 30 IU/dL). The mechanisms for low VWF levels in these patients are poorly understood but are thought to be due to decreased synthesis and/or decreased secretion, increased clearance, or altered proteolysis. Endothelial colony forming cells (ECFCs, also known as blood outgrowth endothelial cells or endothelial progenitor cells) are a source of donor-specific endothelial cells and have demonstrated defects in VWF release and packaging in VWD. ECFCs have been previously studied in patients with VWD, but have not been evaluated in individuals with low VWF levels.Hypothesis/ObjectiveWe hypothesize that the study of ECFCs from individuals with low VWF levels will reveal deficiencies of abnormal VWF release and shed light on the mechanisms of low VWF.MethodsECFC Derivation: Patients with low VWF levels and MCB (30-50 IU/dL) were enrolled in a IRB-approved study and whole blood was collected into sodium heparin vacutainers, layered on Ficoll and centrifuged to isolate the mononuclear cell layer, which is then plated in a twelve-well collagen coated plate. After extended incubation, the presence of phenotypically ECFCs was confirmed by visual morphology and by single stain flow cytometry with appropriate (positive/negative) staining for the endothelial cells markers CD146, CD105, CD31 and negative non-endothelial cell markers CD34, CD90, CD133, and CD45.Stimulated Release of VWF from ECFC: ECFCs were plated in 12 well tissue culture plates. For stimulated release, the cells were exposed to 1mL serum-free media ± phorbol-12-myristate-13-acetate (PMA) (50ng/mL) for 1 hour. The conditioned media samples were then collected and assayed via the VWF:Ag ELISA.Enzyme-Linked ImmunoSorbant Assay (VWF:Ag ELISA): A 96-well plate was be coated with anti-VWF antibodies and after blocking and incubation with respective samples VWF levels were determined anti-VWF primary and HRP-linked secondary antibodies; values were compared to a standard curve and normalized to cell surface area. Results are analyzed via ANOVA with correction for multiple comparisons.ResultsECFC Derivation: A total of seven ECFC lines were created, 5 from individuals with MCB and VWF levels between 30-50 IU/dL (4:1 female: male ratio, age range 11-54 years) and 2 from healthy controls (2 female, age range 22-39 years, control ECFC 1, control ECFC 2) without symptoms of MCB.VWF release from patient derived endothelial cells: When treated with 0ng/mL of PMA, Low VWF derived ECFCs showed no demonstrable differences in VWF release into the conditioned media (Figure A). However, when treated with 50ng/mL of PMA, there is a noticeable trend of increased VWF release in the HUVEC and Control 1 and 2 ECFCs in comparison to the Low VWF ECFCs 1-5, although the results were not statistically significant (Figure B). When respective cohorts are grouped, the low VWF cohort demonstrates a statistically significant decrease in VWF at 50ng/mL as compared to HUVEC (p=0.0001) or control ECFC (p=0.0147) (Figure C).ConclusionsWe have demonstrated the ability to derive ECFCs from patients with Low VWF levels and MCB. Patient derived ECFCs from individuals with Low VWF levels demonstrate impairments in VWF release when stimulated with PMA and as a group, those individuals with low VWF levels demonstrate a significant decrease in VWF release. This decrease in stimulated VWF release is seen with several different individual patient samples and suggests that individuals with low VWF levels may have defects in VWF synthesis and or release that correspond to their lower plasma levels of VWF. [Display omitted] DisclosuresNg:Shire: Consultancy; CSL Behring Heimburger Award: Research Funding.

MicroRNA Profiles of Maternal and Neonatal Endothelial Progenitor Cells in Preeclampsia.

Preeclampsia is associated with an increased cardiovascular morbidity of mother and offspring, thus contributing to a substantial burden in women and children’s health. It has been proven that endothelial progenitor cell (EPC) numbers and functional characteristics are impaired in cardiovascular disease and preeclampsia, although causative factors for the latter have remained elusive. MicroRNA (miRNA) modifications are a potential mechanism through which exposure to an altered environment translates into the development of chronic disease. In this study, we examined whether development of preeclampsia corresponds to alterations of miRNAs in maternal- and cord-blood-derived EPC. To test this end, we analyzed maternal and neonatal miRNAs via RNA sequencing from endothelial cells of preeclamptic and healthy controls in different cell culture passages. We were able to demonstrate differentially represented miRNAs in all groups. Hsa-miR-1270 showed significantly different levels in cord blood EPC from preeclampsia versus control and was negatively correlated with mRNA levels of its predicted targets ANGPTL7 and TFRC. Transfection with an hsa-miR-1270 inhibitor decreased the tube formation capacity and chemotactic motility but did not change proliferation in vitro. Target predictions and gene set enrichment analyses identified alternative splicing as a significantly enriched pathway for hsa-miR-1270. The top miRNAs in three other groups were predicted to target transcriptional and developmental pathways. Here, we showed for the first time significantly different levels of miRNAs and differently represented mRNA levels of predicted target genes in EPC derived from preeclampsia. Understanding the effects of preeclampsia on the epigenetic mechanisms of EPC will be crucial and may provide initial insights for further evaluation of the benefits of therapies targeting this cell population.

Blue laser-induced selective vasorelaxation by the activation of NOSs

Phototherapy has been tried for treating cardiovascular diseases. In particular, ultraviolet and blue visible lights were suggested to be useful due to their nitric oxide (NO)-production ability in the skin. However, the effects of blue light on the arterial contractility are controversial. Here, we hypothesized that appropriate protocol of blue laser can induce selective vasorelaxation by activating vasodilating signaling molecules in arteries. Using organ chamber arterial mechanics, NO assay, Matrigel assay, and microarray, we showed that a 200-Hz, 300-μs, 445-nm pulsed-laser (total energy of 600 mJ; spot size 4 mm) induced selective vasorelaxation, without vasocontraction in rat mesenteric arteries. The laser stimulation increased NO production in the cord blood-endothelial progenitor cells (CB-EPCs). Both the laser-induced vasorelaxation and NO production were inhibited by a non-selective, pan-NO synthase inhibitor, L-NG-Nitro arginine methyl ester. Microarray study in CB-EPCs suggested up-regulation of cryptochrome (CRY)2 as well as NO synthase (NOS)1 and NOSTRIN (NOS trafficking) by the laser. In conclusion, this study suggests that the 445-nm blue puled-laser can induce vasorelaxation possibly via the CRY photoreceptors and NOSs activation. The blue laser-therapy would be useful for treating systemic hypertension as well as improving local blood flow depending on the area of irradiation.

Beneficial Effects of Astragaloside IV-Treated and 3-Dimensional-Cultured Endothelial Progenitor Cells on Angiogenesis and Wound Healing.

Astragaloside IV (AS-IV) is a natural herb extract and a popular compound used in traditional Chinese medicine because of its effect on multiple biological processes, such as promotion of cell proliferation, improvement in cardiopulmonary and vascular function, and promotion of angiogenesis around wounds, leading to accelerated wound healing. Vascular regeneration primarily results from the differentiation of endothelial progenitor cells (EPCs). Biomedical acceleration of angiogenesis and differentiation of EPCs around the wound remain challenging. In this study, we treated human umbilical cord blood-derived EPCs with AS-IV and cultured them on 2-dimensional (tissue culture polystyrene) and 3-dimensional culture plates (3DPs). These cultured cells were then combined with human blood plasma gel and applied on the skin of nude mice in an attempt to repair full-thickness skin defects. The results show that using 3DP culture could increase vascular-related gene expression in EPCs. Furthermore, 12.5 μg/mL AS-IV-treaded EPCs were combined with plasma gels (P-3DP-EPC12.5) and showed enhanced vascular-related protein expression levels after 3 days of culture. Finally, P-3DP-EPC12.5s were used to repair full-thickness skin defects in nude mice, and we could register a wound healing rate greater than 90% by day 14. Based on these results, we concluded that we have developed a potential therapeutic approach for wound healing using plasma gel containing 3-dimensional surface-cultured AS-IV-treated EPCs.

Comparative Preliminary Analysis of Umbilical Cord Blood-, Healthy Adult Peripheral Blood- and Myocardial Infarct Peripheral Blood-Derived Endothelial Progenitor Cells

BACKGROUND: Cardiovascular disease is a leading cause of death globally with the 287,000 deaths per years, characterized by declining of heart function caused by the reduction of heart capacity and lead to heart failure. Cell therapy using endothelial progenitor cells (EPCs) has a big potential for cardiovascular regeneration. EPCs are cells that have ability to differentiate into endothelial cells that can be mobilized and integrated into the defected blood vessel by angiogenesis.&#x0D; AIM: We aimed to seek the superior EPCs derived from MNCs for functional improvement of advanced heart failure patient by cell therapy using EPCs.&#x0D; MATERIALS AND METHODS: We did preliminary analysis to compare umbilical cord blood (UCB), healthy adult peripheral blood (PB)-, and myocardial infarct PB-derived EPCs characteristic and surface phenotypes. Different sources of each EPCs mononuclear cells were characterized by the expression of endothelial (cluster of differentiation [CD] 31, acethylated low-density lipoprotein, and von Willebrand) and hematopoietic stem cell (CD45, CD34, and CD133) surface markers with flow cytometry.&#x0D; RESULTS: In this study, EPCs and the conditioned medium (CM) have been produced and characterized in laboratory scale by comparing several sources of EPCs for instance UCB, PB from healthy people, and patients with myocardial infarction. We have shown that EPC characterizations from each group were not significantly different, but vascular endothelial growth factor and hepatocyte growth factor in UBC-derived CM-EPCs were higher than in PB.&#x0D; CONCLUSION: In conclusion, the UBC-derived EPCs might have a better potential for cardiovascular regeneration.