Endothelial Progenitor Cells Apoptosis Research Articles

TRIM27 Promotes Endothelial Progenitor Cell Apoptosis in Patients with In-Stent Restenosis by Ubiquitinating TBK1.

Approximately 2-10% in-stent restenosis (ISR) may occur following percutaneous coronary intervention (PCI) despite the use of modern drug-eluting stents (DES); thus, our study aimed to explore the effects of tripartite motif-containing (TRIM) 27 on ISR and the underlying mechanism. For this purpose, a total of 42 patients undergoing coronary angiography who had prior coronary angiography with DES implantation were recruited. Endothelial progenitor cells (EPCs) markers (defined as CD34 and vascular endothelial growth factoreceptor-2 (VEGFR-2)) in peripheral blood were measured to asses the circulating EPC level. The TRIM family-related gene expressions were detected by reverse transcription-quantitative polymerase chain reaction. Results suggested that ISR patients had reduced CD34+VEGFR-2+ and increased apoptosis rate of EPCs, along with upregulated TRIM27 and TRIM37 and downregulated TRIM28. TRIM27 promoted and TBK1 inhibited the apoptosis rate of EPCs. Mechanically, TRIM27 interacted with TBK1 to ubiquitinate TBK1 in in vitro study. In summary, TRIM27 promoted the progression of ISR in patients after PCI by ubiquitinating TBK1, which might provide novel ideas for the clinical treatment of ISR.

Cigarette smoke extract induces the senescence of endothelial progenitor cells by upregulating p300.

Endothelial progenitor cells (EPCs) are the main source of endothelial cells. The senescence of EPCs is involved in the pathogenesis of chronic obstructive pulmonary disease (COPD). Cigarette smoke extract (CSE) can directly induce the dysfunction and increased expression of senescence-related markers in EPCs cultured in vitro. Histone acetyltransferase p300 is a transcriptional activator, and its changes can lead to cell senescence. The present study investigated whether CSE can induce the senescence of EPCs by upregulating p300. EPCs were isolated from bone marrow of C57BL/6J mice by density gradient centrifugation. The p300 inhibitor C646 and agonist CTPB were used to interfere with EPCs, cell cycle and apoptosis were detected by flow cytometry, the proportion of senile cells was counted by β-galactosidase staining, the protein expression of p300, H4K12, Cyclin D1, TERT and Ki67 were detected by western blot. Compared with the control group, the cell cycle of CSE group and CTPB group were blocked, the apoptosis rate and early apoptosis rate were increased, the proportion of senile cells counted by β-galactosidase staining was increased, the expression of p300 and H4K12 protein were increased, the expression of Cyclin D1, TERT and Ki67 protein were decreased. C646 could partly alleviate the damages caused by CSE. CSE may promote the apoptosis and senescence of EPCs by upregulating the expression of p300 and H4K12 protein, thus preventing the transition of EPCs from G1 phase to S phase, affecting telomerase synthesis, and reducing EPCs proliferation.

TWEAK Inhibits Venous Thrombosis Progression Through the NF-κB Signaling Pathway in Lung Cancer

Deep vein thrombosis (DVT) is a common complication of malignancy, which greatly increases the mortality rate of tumor patients. Therefore, it is critical to understand the mechanism of malignancy and DVT. TWEAK expression in lung adenocarcinoma tissues from TCGA dataset was performed by using GEPIA and detected in 58 lung cancer patients with DVT by qRT-PCR. TWEAK shRNAs were transfected into endothelial progenitor cells (EPCs) to analyze the consequent alteration in EPCs behaviors through CCK-8, cloning formation, transwell, and flow cytometry assays. TWEAK was obviously declined in lung cancer patients with DVT and low expression of TWEAK was related to poor overall survival. The function experiments revealed that TWEAK over-expression facilitated EPCs proliferation, migration, invasion, and attenuated cell apoptosis. However, TWEAK inhibition showed the opposite effects on EPCs behavior. Mechanistically, TWEAK over-expression promoted the activation of p-p65 and p-IkBα. Moreover, NF-κB pathway inhibitor overturned the effects of TWEAK on EPCs proliferation, metastasis and apoptosis. TWEAK might inhibit venous thrombosis progression through the NF-κB signaling pathway in lung cancer.

Loxenatide attenuates ROS-mediated vascular endothelial progenitor cell damage and mitochondrial dysfunction via SIRT3/Foxo3 signaling pathway.

Diabetes mellitus (DM), becomes a main public health issue worldwide due to the rapid increase in DM patient numbers. The dysfunction of endothelial progenitor cells (EPCs) in DM patients plays a critical role in endothelial repair and the progression of DM-related vascular complications. Loxenatide is an a glucagon-like peptide 1 receptor agonist, which is used to control glycemic in type 2 diabetes patients. However, the role of Loxenatide in EPCs remains to be investigated. EPCs were isolated, characterized, and treated with Loxenatide, high-glucose, or 3-TYP. quantitative real-time polymerase chain reaction, flow cytometry, western blot, and cell counting kit-8 assay were employed to validate the expression of gene and protein expressions and cell viability, respectively. Application of Seahorse XFp to measure oxygen consumption and mitochondrial membrane potential (MMP) were measured by Seahorse XFp and MMP assay. Loxenatide attenuated high-glucose-induced reactive oxygen species (ROS) production and mitochondrial-dependent apoptosis of EPCs in a concentration-dependent manner. The EPC mitochondrial respiration dysfunction induced by high glucose was also repressed by the loxenatide treatment. The protection effect of Loxenatide on EPCs against high-glucose was applied by activating the sirtuin 3 (SIRT3)/Foxo3 signaling pathway. We demonstrated the regulatory role of Loxenatide in mitochondrial dysfunction and apoptosis of EPCs. We elucidated that Loxenatide protects EPC from high-glucose-induced apoptosis via ROS-mediated mitochondrial pathway through the SIRT3/Foxo3 signing pathway. This may provide a new therapeutical target for the treatment of DM-related vascular complications.

Matrine Protects Endothelial Progenitor Cells against Apoptosis and Promotes Their Migration, Invasion, and Tube Formation Abilities via Modulating miR-126b/FOXO4 Axis

Introduction and Objective: Endothelial progenitor cells (EPCs) have been proven to exhibit a therapeutic effect in deep vein thrombosis, but are susceptible to microenvironment. Besides, Matrine has promotive effects on EPCs, but its effects on microRNA (miR)-126 remain obscure, which are therefore discussed in the study. Methods: The cultured EPCs were extracted from Sprague-Dawley rats and identified by immunofluorescence assay. After being treated with Matrine or transfected with miR-126b inhibitor and small interfering RNA targeting forkhead box (FOXO) 4, the viability and apoptosis of EPCs were determined by cell counting kit-8 assay and flow cytometry. The migration, invasion, and tube formation abilities were detected by scratch, Transwell, and tube formation assays. The target genes of miR-126b were predicted by TargetScan, and verified by dual-luciferase reporter assay. The expressions of miR-126b, FOXO4, matrix metalloproteinase (MMP) 2, MMP9, and vascular endothelial growth factor (VEGF) A were determined by quantitative real-time polymerase chain reaction and Western blot. Results: The EPCs were successfully extracted and cultured, as evidenced by positive reaction cluster of differentiation (CD) 34 and CD133. Matrine promoted the viability, migration, invasion, and tube formation while inhibiting the apoptosis of EPCs, and upregulated the expression of miR-126b. Besides, miR-126b inhibitor reversed the effects of Matrine on EPCs and downregulated the expression levels of MMP2, MMP9, and VEGFA. MiR-126b targeted the FOXO4, and siFOXO4 reversed the abovementioned effects of miR-126b inhibitor on EPCs. Conclusion: Matrine protects EPCs from apoptosis and promotes their migration, invasion, and tube formation abilities via regulating miR-126b/FOXO4 axis.

Downregulation of miR-125a-5p Promotes Endothelial Progenitor Cell Migration and Angiogenesis and Alleviates Deep Vein Thrombosis in Mice Via Upregulation of MCL-1.

Endothelial progenitor cells (EPCs) contribute to recanalization of deep vein thrombosis (DVT). MicroRNAs (miRNAs) play regulatory roles in functions of EPCs, which is becoming a promising therapeutic choice for thrombus resolution. The main purpose of this study was to explore the effect of miR-125a-5p on EPC functions in deep vein thrombosis (DVT). EPCs were isolated from the peripheral blood of patients with DVT. In DVT mouse models, DVT was induced by stenosis of the inferior vena cava (IVC). The levels of miR-125a-5p and myeloid cell leukemia sequence 1 (MCL-1) in EPCs and thrombi of DVT mice were detected by RT-qPCR. EPC migration, angiogenesis, and apoptosis were estimated by Transwell assay, tube formation assay, and flow cytometry analysis. Luciferase reporter assay was utilized for detecting the binding of miR-125a-5p and MCL-1. The phosphorylation of PI3K and AKT was estimated by western blot. DVT formation in vivo was observed through hematoxylin-eosin (H&E) staining. The expression of thrombus resolution marker, CD34 molecule (CD34), in the thrombi was measured by immunofluorescence staining. MiR-125a-5p upregulation repressed EPC migration and angiogenesis and facilitated apoptosis. MiR-125a-5p downregulation showed the opposite effect. MCL-1 was targeted and negatively regulated by miR-125a-5p. Additionally, miR-125a-5p inhibited the PI3K/AKT pathway in EPCs. Inhibition of MCL-1 or PI3K/AKT pathway reversed the effect of miR-125a-5p knockdown on EPC functions. The in vivo experiments revealed that miR-125a-5p downregulation repressed thrombus formation and promoted the homing capability of EPCs to the thrombosis site, thereby alleviating DVT mice. Downregulation of miR-125a-5p promotes EPC migration and angiogenesis by upregulating MCL-1, thereby enhancing EPC homing to thrombi and facilitating thrombus resolution.

Nitric Oxide Generation and Endothelial Progenitor Cells Recruitment for Improving Hemocompatibility and Accelerating Endothelialization of Tissue Engineering Heart Valve

AbstractTissue engineering heart valve (TEHV) offers great potential to overcome the limitations of commercial artificial valves used in clinical practice as a permanent prosthetic valve. Currently, decellularized heart valve (DHV) is the most widely used scaffold for TEHV, but showed suboptimal performance due to difficulty of endothelialization. Facilitating endothelialization of DHV is indispensable for better valve performance, and excellent hemocompatibility guarantees enough time windows for endothelialization process. Herein, a dual‐functional TEHV scaffold with improving hemocompatibility and accelerating endothelialization is constructed by modifying DHV with copper ions (Cu) and growth differentiation factor 11 (GDF11). Results show the newly‐constructed scaffold successfully generates endogenous nitric oxide (NO) through catalysis of Cu, and possesses improved hemocompatibility by down‐regulating platelets activation and adhesion. Furthermore, GDF11 immobilization significantly accelerates scaffold endothelialization through facilitating recruitment, supporting growth, and alleviating apoptosis of endothelial progenitor cells . This TEHV scaffold shows favorable performance under in vivo hemodynamic environment with intact endothelial coverage and adaptive ECM remodeling, and without thrombus or calcification formation. This newly‐constructed TEHV scaffold is expected to make up for the shortcomings of currently available prosthetic valves in clinical practice and has the potential possibility of rapid translation to the clinic as a better prosthetic valve.

Melatonin Improves Ischemia-Induced Circulation Recovery Impairment in Mice with Streptozotocin-Induced Diabetes by Improving the Endothelial Progenitor Cells Functioning.

Patients with diabetes mellitus tend to develop ischemia-related complications and have compromised endothelial progenitor cell (EPC) function. Melatonin protects against ischemic injury, possibly via EPC modulation. We investigated whether melatonin pretreatment could restore EPC function impairment and improve circulation recovery in a diabetic critical limb ischemia mouse model. Under 25 mM high-glucose medium in vitro, EPC proliferation, nitric oxide production, tube formation, and endothelial nitric oxide synthase (eNOS) phosphorylation were significantly suppressed. Hyperglycemia promoted EPC senescence and apoptosis as well as increased reactive oxygen species (ROS) production. Melatonin treatment reversed the harmful effects of hyperglycemia on EPC through adenosine monophosphate–activated protein kinase-related mechanisms to increase eNOS phosphorylation and heme oxygenase-1 expression. In an in-vivo study, after a 4-week surgical induction of hindlimb ischemia, mice with streptozotocin (STZ)-induced diabetes showed significant reductions in new vessel formation, tissue reperfusion, and EPC mobilization in ischemic hindlimbs compared to non-diabetic mice. Mice with STZ-induced diabetes that received melatonin treatment (10 mg/kg/day, intraperitoneal) had significantly improved blood perfusion ratios of ischemic to non-ischemic limb, EPC mobilization, and densities of capillaries. In addition, a murine bone marrow transplantation model to support these findings demonstrated that melatonin stimulated bone marrow-originated EPCs to differentiate into vascular endothelial cells in femoral ligation-induced ischemic muscles. In summary, this study suggests that melatonin treatment augments EPC function along with neovascularization in response to ischemia in diabetic mice. We illustrated the protective effects of melatonin on EPC H2O2 production, senescence, and migration through melatonin receptors and modulating eNOS, AMPK, and HO-1 activities at the cellular level. Thus, melatonin might be used to treat the impairment of EPC mobilization and circulation recuperation in response to ischemic injury caused by chronic hyperglycemia. Additional studies are needed to elucidate the applicability of the results in humans.

Nrf2 transcriptional upregulation of IDH2 to tune mitochondrial dynamics and rescue angiogenic function of diabetic EPCs

Endothelial progenitor cells (EPCs) are reduced in number and impaired in function in diabetic patients. Whether and how Nrf2 regulates the function of diabetic EPCs remains unclear. In this study, we found that the expression of Nrf2 and its downstream genes were decreased in EPCs from both diabetic patients and db/db mice. Survival ability and angiogenic function of EPCs from diabetic patients and db/db mice also were impaired. Gain- and loss-of-function studies, respectively, showed that knockdown of Nrf2 increased apoptosis and impaired tube formation in EPCs from healthy donors and wild-type mice, while Nrf2 overexpression decreased apoptosis and rescued tube formation in EPCs from diabetic patients and db/db mice. Additionally, proangiogenic function of Nrf2-manipulated mouse EPCs was validated in db/db mice with hind limb ischemia. Mechanistic studies demonstrated that diabetes induced mitochondrial fragmentation and dysfunction of EPCs by dysregulating the abundance of proteins controlling mitochondrial dynamics; upregulating Nrf2 expression attenuated diabetes-induced mitochondrial fragmentation and dysfunction and rectified the abundance of proteins controlling mitochondrial dynamics. Further RNA-sequencing analysis demonstrated that Nrf2 specifically upregulated the transcription of isocitrate dehydrogenase 2 (IDH2), a key enzyme regulating tricarboxylic acid cycle and mitochondrial function. Overexpression of IDH2 rectified Nrf2 knockdown- or diabetes-induced mitochondrial fragmentation and EPC dysfunction. In a therapeutic approach, supplementation of an Nrf2 activator sulforaphane enhanced angiogenesis and blood perfusion recovery in db/db mice with hind limb ischemia. Collectively, these findings indicate that Nrf2 is a potential therapeutic target for improving diabetic EPC function. Thus, elevating Nrf2 expression enhances EPC resistance to diabetes-induced oxidative damage and improves therapeutic efficacy of EPCs in treating diabetic limb ischemia likely via transcriptional upregulating IDH2 expression and improving mitochondrial function of diabetic EPCs.

Abstract P3011: Nrf2 Transcriptional Upregulation Of Idh2 To Tune Mitochondrial Dynamics And Rescue Angiogenic Function Of Diabetic Epcs

Endothelial progenitor cells (EPCs) are reduced in number and impaired in function in diabetic patients. Whether and how Nrf2 regulates the function of diabetic EPCs remains unclear. In this study, we found that the expression of Nrf2 and its downstream genes were decreased in EPCs from both diabetic patients and db/db mice. Survival ability and angiogenic function of EPCs from diabetic patients and db/db mice also were impaired. Gain- and loss-of-function studies, respectively, showed that knockdown of Nrf2 increased apoptosis and impaired tube formation in EPCs from healthy donors and wild-type mice, while Nrf2 overexpression decreased apoptosis and rescued tube formation in EPCs from diabetic patients and db/db mice. Additionally, proangiogenic function of Nrf2-manipulated mouse EPCs was validated in db/db mice with hind limb ischemia. Mechanistic studies demonstrated that diabetes induced mitochondrial fragmentation and dysfunction of EPCs by dysregulating the abundance of proteins controlling mitochondrial dynamics; upregulating Nrf2 expression attenuated diabetes-induced mitochondrial fragmentation and dysfunction and rectified the abundance of proteins controlling mitochondrial dynamics. Further RNA-sequencing analysis demonstrated that Nrf2 specifically upregulated the transcription of isocitrate dehydrogenase 2 (IDH2), a key enzyme regulating tricarboxylic acid cycle and mitochondrial function. Overexpression of IDH2 rectified Nrf2 knockdown- or diabetes-induced mitochondrial fragmentation and EPC dysfunction. In a therapeutic approach, supplementation of an Nrf2 activator sulforaphane enhanced angiogenesis and blood perfusion recovery in db/db mice with hind limb ischemia. Collectively, these findings indicate that Nrf2 is a potential therapeutic target for improving diabetic EPC function. Thus, elevating Nrf2 expression enhances EPC resistance to diabetes-induced oxidative damage and improves therapeutic efficacy of EPCs in treating diabetic limb ischemia likely via transcriptional upregulation of IDH2 expression and improving mitochondrial function of diabetic EPCs.

Synergistic effect of VEGF and SDF-1α in endothelial progenitor cells and vascular smooth muscle cells.

Vascular endothelial growth factor (VEGF) is a potent agonist of angiogenesis that induces proliferation and differentiation of endothelial progenitor cells (EPCs) after vascular injury. Previous studies have suggested that stromal cell-derived factor 1-alpha (SDF-1α) and VEGF have a synergistic effect on vascular stenosis. The aim of the present study was to investigate whether VEGF and SDF-1α act synergistically in EPCs and vascular smooth muscle cells (VSMCs). In this study, EPCs were isolated from rat bone marrow and their morphology and function were studied. Subsequently, VEGF was delivered into EPCs using an adenoviral vector. Tube formation, migration, proliferation, and apoptosis of VEGF-overexpressing EPCs was analyzed. Then, EPCs were co-cultured with VSMCs in the presence or absence of SDF-1α, the migration, proliferation, apoptosis, and differentiation capacity of EPCs and VSMCs were analyzed respectively. The isolated EPCs showed typical morphological features, phagocytic capacity, and expressed surface proteins. While stable expression of VEGF remarkably enhanced tube formation, migration, and proliferation capacity of EPCs, apoptosis was decreased. Moreover, the proliferation, migration, and differentiation capacity of EPCs in the co-cultured model was enhanced in the presence of SDF-1α, and apoptosis was decreased. However, these effects were reversed in VSMCs. Therefore, our results showed that VEGF and SDF-1α synergistically increased the migration, differentiation, and proliferation capabilities of EPCs, but not VSMCs. This study suggests a promising strategy to prevent vascular stenosis.

CXCR7 Agonist TC14012 Improves Angiogenic Function of Endothelial Progenitor Cells via Activating Akt/eNOS Pathway and Promotes Ischemic Angiogenesis in Diabetic Limb Ischemia.

Endothelial progenitor cells (EPCs) play a critical role in repairing damaged vessels and triggering ischemic angiogenesis, but their number is reduced and function is impaired under diabetic conditions. Improving EPC function has been considered a promising strategy to ameliorate diabetic vascular complications. In the present study, we aim to investigate whether and how CXCR7 agonist TC14012 promotes the angiogenic function of diabetic EPCs. High glucose (HG) treatment was used to mimic the hyperglycemia in diabetes. Tube formation, cell scratch recovery and transwell assay, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, and cleaved-caspase3 expression were used to evaluate the angiogenic capability, cell migration, and apoptosis of EPCs, respectively. Hind limb ischemia (HLI) model was used to appraise the ability of TC14012 in promoting diabetic ischemic angiogenesis in vivo. HG treatment impaired EPC tube formation and migration, and induced EPC apoptosis and oxidative damage, while TC14012 rescued tube formation and migration, and prevented HG-induced apoptosis and oxidative damage of EPCs. Furthermore, these beneficial effects of TC14012 on EPCs were attenuated by specific siRNAs against CXCR7, validating that CXCR7 is a functional target of TC14012 in EPCs. Mechanistic studies demonstrated that HG treatment reduced CXCR7 expression in EPCs, and impaired Akt and endothelial nitric oxide synthase (eNOS) phosphorylation and nitric oxide (NO) production; similarly, these signal impairments in HG-exposed EPCs could be rescued by TC14012. However, the protective effects of TC14012 on tube formation and migration, Akt and eNOS phosphorylation, and NO production in HG-treated EPCs were almost completely abolished by siRNAs against CXCR7 or Akt specific inhibitor wortmannin. More importantly, in vivo study showed that TC14012 administration enhanced blood perfusion recovery and angiogenesis in the ischemic hind limb and increased the EPC number in peripheral circulation of db/db mice, demonstrating the capability of TC14012 in promoting EPC mobilization and ischemia angiogenic function. TC14012 can prevent EPCs from HG-induced dysfunction and apoptosis, improve eNOS activity and NO production via CXCR7/Akt signal pathway, and promote EPC mobilization and diabetic ischemia angiogenesis.

Upregulated miR-206 Aggravates Deep Vein Thrombosis by Regulating GJA1-Mediated Autophagy of Endothelial Progenitor Cells.

Background Deep vein thrombosis (DVT) is the third most prevalent vascular disease worldwide. MicroRNAs (miRNAs) play regulatory roles in functions of endothelial progenitor cells (EPCs), which is becoming a promising therapeutic choice for thrombus resolution. Nevertheless, the role of miR-206 in EPCs is unclear. Methods EPCs were isolated from the peripheral blood of patients with DVT. In DVT mouse models, DVT was induced by stenosis of the inferior vena cava (IVC). The levels of miR-206 and gap junction protein alpha 1 (GJA1) in EPCs and vascular tissues of DVT mice were detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The proliferation, migration, apoptosis, and angiogenesis were tested by cell counting kit-8 (CCK-8) assay, Transwell assay, flow cytometry analysis, and in vitro tube formation assay. The levels of autophagy-related proteins as well as the level of GJA1 in EPCs and vascular tissues were evaluated by western blotting. DVT formation in vivo was observed through hematoxylin-eosin (HE) staining. The expression of thrombus resolution markers, CD34 molecule (CD34) and matrix metallopeptidase 2 (MMP2), in the thrombi was measured by immunofluorescence staining. Results miR-206 overexpression inhibited proliferation, migration, and angiogenesis and promoted apoptosis of EPCs, while miR-206 knockdown exerted an opposite effect on EPC phenotypes. Downregulation of GJA1, the target of miR-206, abolished the influence of miR-206 on EPC phenotypes. Furthermore, silencing of miR-206 suppressed the autophagy of EPCs via upregulating GJA1. miR-206 knockdown repressed thrombus formation, enhanced the homing ability of EPCs to the thrombosis site, and facilitated thrombus resolution in DVT mouse models. Additionally, miR-206 was upregulated while GJA1 was downregulated in vascular tissues of DVT mice. miR-206 knockdown elevated GJA1 expression in vascular tissues of DVT mice. The expression of miR-206 was negatively correlated with that of GJA1 in DVT mice. Conclusion miR-206 knockdown upregulates GJA1 to inhibit autophagy of EPCs and then promote EPC proliferation, migration, and angiogenesis, thereby enhancing EPC homing to thrombi and facilitating thrombus resolution.

CD146 mediates the anti-apoptotic role of Netrin-1 in endothelial progenitor cells under hypoxic conditions.

Modulating the biological status of endothelial progenitor cells (EPCs), such as function and survival, is essential for therapeutic angiogenesis in ischemic vascular disease environments. This study aimed to explore the role and molecular mechanisms underlying Netrin-1 in the viability and angiogenic function of EPCs. EPCs were isolated from the bone barrow of adult C57/BL6 mice. The apoptosis and various functions of EPCs were analyzed in vitro by manipulating the expression of Netrin-1. The TUNEL assay was performed to detect apoptotic EPCs. Cell migration and tube formation assays were performed to detect EPC function. Trypan blue staining was performed to detect cell viability. Western blot analysis was performed to detect the protein expression levels of Netrin-1, CD146 and apoptotic factors. Quantitative PCR analysis was performed to detect the expression levels of Netrin-1 receptors. The results demonstrated that treatment with exogenous Netrin-1 promoted EPC migration and tube formation, whereas transfection with small interfering (si)RNA targeting Netrin-1 exhibited the opposite effects. Exogenous Netrin-1 protected EPCs from hypoxia-induced apoptosis, whereas the interruption of endogenous Netrin-1 enhancement under hypoxia by Netrin-1-siRNA exacerbated the apoptosis of EPCs. Furthermore, CD146, one of the immunoglobulin receptors activated by Netrin-1, was screened for in the present study. Results demonstrated that CD146 did not participate in Netrin-1-promoted EPC function, but mediated the anti-apoptotic effects of Netrin-1 in EPCs. In conclusion, Netrin-1 enhanced the angiogenic function of EPCs and alleviated hypoxia-induced apoptosis, which was mediated by CD146. This biological function of Netrin-1 may provide a potential therapeutic option to promote EPCs for the treatment of ischemic vascular diseases.

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.

Extracorporeal Cardiac Shock Waves Therapy Improves the Function of Endothelial Progenitor Cells After Hypoxia Injury via Activating PI3K/Akt/eNOS Signal Pathway.

Background: Extracorporeal cardiac shock waves (ECSW) have great potential in the treatment of coronary heart disease. Endothelial progenitor cells (EPCs) are a class of pluripotent progenitor cells derived from bone marrow or peripheral blood, which have the capacity to migrate to ischemic myocardium and differentiate into mature endothelial cells and play an important role in neovascularization and endothelial repair. In this study, we investigated whether ECSW therapy can improve EPCs dysfunction and apoptosis induced by hypoxia and explored the underlying mechanisms.Methods: EPCs were separated from ApoE gene knockout rat bone marrow and identified using flow cytometry and fluorescence staining. EPCs were used to produce in vitro hypoxia-injury models which were then divided into six groups: Control, Hypoxia, Hypoxia + ECSW, Hypoxia + LY294002 + ECSW, Hypoxia + MK-2206 + ECSW, and Hypoxia + L-NAME + ECSW. EPCs from the Control, Hypoxia, and Hypoxia + ECSW groups were used in mRNA sequencing reactions. mRNA and protein expression levels were analyzed using qRT-PCR and western blot analysis, respectively. Proliferation, apoptosis, adhesion, migration, and angiogenesis were measured using CCK-8, flow cytometry, gelatin, transwell, and tube formation, respectively. Nitric oxide (NO) levels were measured using an NO assay kit.Results: Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis showed that differentially expressed genes were enriched in cancer signaling, PI3K-Akt signaling, and Rap1 signaling pathways. We selected differentially expressed genes in the PI3K-Akt signaling pathway and verified them using a series of experiments. The results showed that ECSW therapy (500 shots at 0.09 mJ/mm2) significantly improved proliferation, adhesion, migration, and tube formation abilities of EPCs following hypoxic injury, accompanied by upregulation of p-PI3K, p-Akt, p-eNOS, Bcl-2 protein and NO, PI3K, and Akt mRNA expression, and downregulation of Bax and Caspase3 protein expression. All these effects of ECSW were eliminated using inhibitors specific to PI3K (LY294002), Akt (MK-2206), and eNOS (L-NAME).Conclusion: ECSW exerted a strong repaired effect on EPCs suffering inhibited hypoxia injury by inhibiting cell apoptosis and promoting angiogenesis, mainly through activating the PI3K/Akt/eNOS signaling pathway, which provide new evidence for ECSW therapy in CHD.

Effects of Sirt1 on proliferation, migration, and apoptosis of endothelial progenitor cells in peripheral blood of SD rats with chronic obstructive pulmonary disease

Objective: To explore the effect of Sirt1 on the function of endothelial progenitor cells (EPCs) in rats with chronic obstructive pulmonary disease (COPD). Methods: A rat COPD model was established via smoking and endotoxin administration for three months. The peripheral circulating EPCs were isolated by gradient centrifugation, and their functions, cell cycle distribution, apoptosis, and Sirt1 expression were examined. The function changes of EPCs in the presence or absence of Sirt1 agonist and inhibitor were estimated; meanwhile, the expressions of Sirt1, FOXO3a, NF-κB, and p53 were also evaluated. Results: The proliferation, adhesion, and migration of EPCs decreased while the apoptosis rate was increased in the COPD rats. The expression of Sirt1 protein in EPCs of the COPD group was significantly lower than that in the control group (P&lt;0.01). The overexpression of the Sirt1 gene using a gene transfection technique or Sirt1 agonists (SRT1720) improved the proliferation, migration, and adhesion, and decreased the apoptosis of EPC. However, Sirt1 inhibitor (EX527) decreased EPC functions in the COPD group. The effect of Sirt1 expression on EPC function may be related to reduction of FOXO3a and increase of NF-κB and p53 activity. Conclusions: Increased expression of Sirt1 can improve the proliferation and migration of EPCs and reduce their apoptosis in COPD rats. This change may be related to FOXO3a, NF-κB, and p53 signaling pathways.

Up-regulation of GLP-1R improved the dysfunction of late EPCs under hyperglycemia by regulating SIRT1 expression

The dysfunction of endothelial progenitor cells (EPCs) is closely associated with diabetic vascular complications. Both glucagonlike peptide-1 receptor (GLP-1R) and silent information regulator 1 (SIRT1) can control systemic glucose homeostasis and protect endothelial cells against hyperglycemia-induced oxidative stress. In this study, we mainly assessed the role played by SIRT1 and GLP-1R and their relationship in regulating the function of late EPCs under hyperglycemia stimulation. Human peripheral blood mononuclear cells (PBMCs) were cultured in EGM-2 medium and induced to differentiate into EPCs and 25 mM glucose was used to stimulate EPCs to obtain a hyperglycemia condition. Subsequently, the expression and location of GLP-1R and SIRT1 in EPCs were detected. After GLP-1R or SIRT1 knockdown, or the treatment by GLP-1R agonist and/or SIRT1 agonist/inhibitor, the effects of SIRT1 and GLP-1R and their relationship in regulating the function of late EPCs under hyperglycemia stimulation was studied by detecting the apoptosis, migration, adhesion and angiogenicity abilities of EPCs. Results demonstrated that, in high-glucose stimulated EPCs, the expression of GLP-1R and SIRT1 was down-regulated. The knockdown of either GLP-1R or SIRT1 could increase EPCs apoptosis and weaken the migration, adhesion and angiogenicity abilities of EPCs. In addition, the improvement effects of Exendin-4 or GLP-1R over-expression on EPCs dysfunction could be weakened to some degree under SIRT1 knockdown. In conclusion, both GLP-1R and SIRT1 expression played important roles in regulating EPCs dysfunction under hyperglycemia and the up-regulation of GLP-1R improved the dysfunction of late EPCs by regulating SIRT1 expression.

Development of a multi-arm polyrotaxanes modified mesoporous silica-coated gold nanoplatform for protecting endothelial progenitor cells against high glucose environment.

Recent study reported that endothelial progenitor cells (EPCs) have potential to treat diabetic macroangiopathy. High glucose environment of diabetes can affect the adhesion of EPCs by decreasing the expression of CXC chemokine receptor 4 (CXCR4) and affect the proliferation of EPCs by decreasing the expression of miR-126. The results showed that the cytotoxicity of GNR@MSNs@PEI to EPCs was significantly lower than PEI; the temperature of GNR@MSNs@PEI solution can be controlled between 38-40°C under 808nm laser irradiation. 25.67µg of pcDNA3.1-GFP-CXCR4 and 5.36µg of FITC-miR-126 could be loaded in 1mg of GNR@MSNs@PEI; GNR@MSNs@PEI has gene transfection almost the same as Lipofectamine 3000. Subsequent in vitro studies showed that pcDNA3.1-GFP-CXCR4 and miR-126 loaded GNR@MSNs@PEI can significantly increase the adhesion and proliferation and decrease the apoptosis of EPCs treated with high glucose under 808nm laser irradiation. In conclusion, nano-carriers (GNR@MSNs@PEI) with high pcDNA3.1-CXCR4 and miR-126 loading capacity, high biocompatibility, well cell internalization, and controllable release ability were constructed to transfer CXCR4 expression plasmid (pcDNA3.1-CXCR4) and miR-126 into EPCs efficiently. Further in vitro studies indicated that pcDNA3.1-CXCR4 and miR-126-loaded GNR@MSNs@PEI could protect EPCs against high glucose-induced injury.

Gleevec and Rapamycin Synergistically Reduce Cell Viability and Inhibit Proliferation and Angiogenic Function of Mouse Bone Marrow-Derived Endothelial Progenitor Cells

Objective: This study investigates the synergistic effects of Gleevec (imatinib) and rapamycin on the proliferative and angiogenic properties of mouse bone marrow-derived endothelial progenitor cells (EPCs). Materials and Methods: EPCs were isolated from mouse bone marrow and treated with different concentrations of Gleevec or rapamycin individually or in combination. The cell viability and proliferation were examined using the MTT assay. An analysis of cell cycle and apoptosis was performed using flow cytometry. Formation of capillary-like tubes was examined in vitro, and the protein expression of cell differentiation markers was determined using Western blot analysis. Results: Gleevec significantly reduced cell viability, cell proliferation, and induced cell apoptosis in EPCs. Rapamycin had similar effects on EPCs, but it did not induce cell apoptosis. The combination of Gleevec and rapamycin reduced the cell proliferation but increased cell apoptosis. Although rapamycin had no demonstratable effect on tube formation, the combined therapy of Gleevec and rapamycin significantly reduced tube formation when compared with Gleevec alone. Mechanistically, Gleevec, but not rapamycin, induced a significant elevation in caspase-3 activity in EPCs, and it attenuated the expression of the endothelial protein marker platelet-derived growth factor receptor α. Functionally, rapamycin, but not Gleevec, significantly enhanced the expression of endothelial differentiation marker proteins, while attenuating the expression of mammalian target of rapamycin signaling-related proteins. Conclusions: Gleevec and rapamycin synergistically suppress cell proliferation and tube formation of EPCs by inducing cell apoptosis and endothelial differentiation. Mechanistically, it is likely that rapamycin enhances the proapoptotic and antiangiogenic effects of Gleevec by promoting the endothelial differentiation of EPCs. Given that EPCs are involved in the pathogenesis of some cardiovascular diseases and critical to angiogenesis, pharmacological inhibition of EPC proliferation by combined Gleevec and rapamycin therapy may be a promising approach for suppressing cardiovascular disease pathologies associated with angiogenesis.