Cigarette smoke extract mediates cell premature senescence in chronic obstructive pulmonary disease patients by up-regulating USP7 to activate p300-p53/p21 pathway.

OBJECTIVES: Tobacco hazard is one of the most severe public health issues in the world. It is believed that smoking is the most important factor leading to chronic obstructive pulmonary disease (COPD). Endothelial progenitor cells (EPCs) originate from the bone marrow and can effectively repair vascular endothelial damage and improve vascular endothelial function. Current studies suggest that EPCs senescence and EPCs depletion exist in smoking-related COPD, but the molecular mechanism remains unclear. METHODS: Co-immunoprecipitation was used to detect the interaction between USP7 and p300. EPCs from smoking COPD patients were isolated, and the expressions of USP7 and p300 were detected by RT-PCR and Western Blot. Different concentrations of cigarette smoke extract (CSE) and USP7 or p300 inhibitors were used to treat EPCs, then the expression of p53, p53 target genes and aging-related genes were detected. Cell Counting Kit - 8 (CCK8) was used to detect cell proliferation, flow cytometry was used to detect cell cycle distribution, ß-galactosidase (ß-gal) staining and Lamp1 immunofluorescence was used to detect the proportion of aging cells. COPD mouse models were used to confirm the molecular mechanism. RESULTS: USP7 and p300 interacted with each other, and USP7 affected the protein stability of p300 by regulating the ubiquitination of p300. There existed high expressions of USP7 and p300 proteins in EPCs of smoking COPD patients and COPD mouse model. CSE promoted the high expressions of USP7 and p300 in EPCs. Further studies showed that CSE mediated the USP7/p300-dependent high expression of p53 and activated the expression of p53 target genes especially p21. Activation of p53 - p21 pathway finally inhibited cell activity, led to cell cycle arrest and premature senescence of EPCs. CONCLUSION: CSE mediated up-regulation of USP7 and p300 activated p53 - p21 pathway was a molecular mechanism that might lead to COPD.

Effects of calcitriol on peripheral endothelial progenitor cells and renal renovation in rats with chronic renal failure.

BACKGROUND: The role of calcitriol (1,25-dihydroxyvitamin D3 or 1,25-(OH)2D3) in physiological processes, such as anti-fibrosis, anti-inflammation, and immunoregulation is known; however, its role in the remodeling of the glomerular capillary endothelium in rats with chronic renal failure (CRF) remains unclear. METHODS: Here, we analyzed the role/number of endothelial progenitor cells (EPCs), renal function, and pathological alterations in rats with CRF, and compared the results before and after supplementation with calcitriol in vivo. RESULTS: Amongst the three experimental groups (sham group, CRF group, and calcitriol-treated group (0.03 µg/kg/d), we observed substantially elevated cell adhesion and vasculogenesis in vivo in the calcitriol-treated group. Additionally, lower levels of serum creatinine (Scr) and blood urea nitrogen (BUN) was recorded in the calcitriol-treated group than the CRF group (p > 0.05). Calcitriol treatment also resulted in an improvement in renal pathological injury. CONCLUSIONS: Thus, calcitriol could ameliorate the damage of glomerular arterial structural and renal tubules vascular network integrity, maybe through regulating the number and function of EPCs in the peripheral blood of CRF rats. Treatment with it may improve outcomes in patients with renal insufficiency or combined cardiac insufficiency. Calcitriol could ameliorate CRF-induced renal pathological injury and renal dysfunction by remodeling of the glomerular capillary endothelium, thus, improving the function of glomerular endothelial cells.

Angiogenic Actions of Paeoniflorin on Endothelial Progenitor Cells and in Ischemic Stroke Rat Model.

Ischemic stroke is one of the major diseases with high morbidity, mortality, and disability rate all over the world. Chinese herb-derived active components would provide valuable candidate compounds for ischemic stroke therapy. Paeoniflorin (PF) is an active ingredient from Paeoniae Radix which possesses neurovascular effect after ischemia. However, so far, few studies are reported on the efficacy and mechanism of PF from angiogenesis aspects. Results from our in vitro studies showed that the ability for proliferation, migration, and tube formation in bone marrow-derived endothelial progenitor cells (BM-EPCs) was promoted by coculturing with PF (100 [Formula: see text]M). Furthermore, to investigate the angiogenic effects of PF in vivo, we constructed an ischemic stroke model in rats and found that PF could reduce cerebral infarction, alleviate pathological injury, and increase the secretion of pro-angiogenic factors and cerebral vascular density after intraperitonially administration of 40 mg ⋅ kg[Formula: see text] ⋅ day[Formula: see text] for 14 days. Up-regulating the expression of VEGF/VEGF-R2 might be the mechanism of PF's angiogenic action. In conclusion, the present study provides evidence that PF is an active monomer of Traditional Chinese Medicine which shows angiogenic actions on endothelial progenitor cells and in ischemic stroke rat model.

Panax notoginseng saponins promote endothelial progenitor cell angiogenesis via the Wnt/ß-catenin pathway.

BACKGROUND: Distraction osteogenesis (DO) is an effective treatment in craniomaxillofacial surgery. However, the issue of sufficient blood supply at the regeneration tissue has limited its wide application. Panax notoginseng saponins (PNS) is a Traditional Chinese Medicine that is commonly used to treat a range of angiogenic diseases. However, the mechanisms whereby PNS alters angiogenesis in endothelial progenitor cells (EPCs) have yet to be clarified. METHODS: EPCs were identified by immunofluorescence, confirmed by their uptake of fluorescently labeled Dil-ac-LDL and FITC-UEA-1. EPCs were treated with different concentrations of PNS, and the effects of PNS on cell proliferation were measured on the optimal concentration of PNS determined. The effects of PNS on angiogenesis and migration, angiogenic cytokines mRNA expression and the proteins of the Wnt pathway were investigated. Then knocked down ß-catenin in EPCs and treated with the optimum concentrational PNS, their angiogenic potential was evaluated in tube formation and migration assays. In addition, the expression of cytokines associated with angiogenesis and Wnt/ß-catenin was then assessed via WB and RT-qPCR. RESULTS: We were able to determine the optimal concentration of PNS in the promotion of cell proliferation, tube formation, and migration to be 6.25 mg/L. PNS treatment increased the mRNA levels of VEGF, bFGF, VE-Cadherin, WNT3a, LRP5, ß-catenin, and TCF4. After knocked down ß-catenin expression, we found that PNS could sufficient to partially reverse the suppression of EPC angiogenesis. CONCLUSIONS: Overall, 6.25 mg/L PNS can promote EPC angiogenesis via Wnt/ß-catenin signaling pathway activation.

Comparative Analysis of the Rabbit Endothelial Progenitor Cells from Bone Marrow and Peripheral Blood Treated with Selenium Nanoparticles.

BACKGROUND: Selenium Nanoparticles (Se-NPs) are known for their antioxidant and anti-inflammatory activities, which are effective in preventing oxidative damage and improving physiological processes. OBJECTIVES: This study aimed at investigating the effects of biosynthesized Se-NPs on bone marrow-derived Endothelial Progenitor Cells (bone marrow-derived EPCs) and blood-derived endothelial progenitor cells (blood-derived EPCs) isolated from rabbits in vitro. METHODS: The cultured EPCs incubated with biosynthesized Se-NPs at the concentrations of 0.19, 0.38, 0.76, 1.71, 3.42, 7.03, 14.25, 28.50, 57, 114, and 228µg/ml for 48h. After screening the proliferative potential of the Se-NPs by the MTT assay, the best concentrations were selected for Real-Time quantitative Polymerase Chain Reaction (RT-qPCR). Real-time quantification of Vascular Cell Adhesion Molecule 1 (VCAM-1), lectin-like oxidized Low-Density Lipoprotein (LDL) receptor-1 (LOX-1), endothelial Nitric Oxide Synthase (eNOS), and Monocyte Chemoattractant Protein-1 (MCP-1) gene expressions were analyzed by normalizing with Glyceraldehyde- 3-Phosphate Dehydrogenase (GAPDH) as an endogenous reference gene. RESULTS: Blood-derived EPCs and bone marrow-derived EPCs showed morphological differences before treatment in vitro. Se-NPs treated EPCs indicated a significant dose-dependent proliferative activity (p<0.01). In general, the expression levels of VCAM-1, LOX-1, and MCP-1 mRNA were significantly decreased (p<0.01), whereas that of the eNOS expression was significantly increased at the concentrations of 7.3 and 14.25µg/ml (p<0.01). Although the expressions of MCP-1, LOX-1, and eNOS mRNA were decreased at certain concentrations of Se-NPs (p<0.01 and p<0.05, respectively) in the treated bone marrow-derived EPCs, no significant differences were observed in the VCAM-1 mRNA expression levels in bone marrow-derived EPCs compared with the control group (p>0.05). CONCLUSION: This was the first report to demonstrate the effects of Se-NPs on proliferative, anti-oxidative, and anti-inflammatory activities for bone marrow-derived EPCs and blood-derived EPCs. Our findings suggested that Se-NPs could be considered as an effective agent that may ameliorate vascular problems.

Pinellia ternata attenuates carotid artery intimal hyperplasia and increases endothelial progenitor cell activity via the PI3K/Akt signalling pathway in wire-injured rats.

CONTEXT: Clinically, Pinellia ternata (Thunb.) Breit. (Araceae) (PT) has been widely used in the treatment of atherosclerosis and hyperlipidaemia, but the underlying mechanisms are still not clearly understood. OBJECTIVE: This research was conducted to confirm the mechanism by which PT affects carotid artery intimal hyperplasia. MATERIALS AND METHODS: An intestinal hyperplasia Sprague-Dawley rat model was established by carotid artery injury. The rats were randomly divided into five groups (n = 8): sham, model, PT (with daily intragastric administration of 10 g/mL/kg PT tubers water extract), PT+LY294002 (with intraperitoneal injection of 50 mg/kg LY294002 + 10 g/mL/kg PT) and endothelial progenitor cells (EPCs) (with injection of 5 × 105/cells), and treated for 4 or 8 weeks. RESULTS: HE staining showed that PT attenuated intimal hyperplasia. RT-PCR, Western blotting and immunohistochemistry showed that PT increased the expression of vascular endothelial growth factor (VEGF) and eNOS in the atherosclerotic carotid artery. PT increased the Dil-acLDL+/FITC-UEA-1+ population (from 0.41 ± 0.085% to 0.60 ± 0.092%) in the blood, decreased TCHO, TG, LDL-C, IL-6 and TNF-α levels, and increased HDL-C and IL-10 levels in the blood. However, these changes were reversed by the PI3K/Akt pathway inhibitor LY294002. DISCUSSION AND CONCLUSIONS: PT can be developed as an atherosclerosis and carotid intimal hyperplasia treatment drug. Therefore, further study will focus on the effects of PT on intimal hyperplasia in wire-injured atherosclerosis patients and explore in depth some other relevant molecular mechanisms.

The Role of Vitamin D in Modulating Mesenchymal Stem Cells and Endothelial Progenitor Cells for Vascular Calcification.

Vascular calcification, which involves the deposition of calcifying particles within the arterial wall, is mediated by atherosclerosis, vascular smooth muscle cell osteoblastic changes, adventitial mesenchymal stem cell osteoblastic differentiation, and insufficiency of the calcification inhibitors. Recent observations implied a role for mesenchymal stem cells and endothelial progenitor cells in vascular calcification. Mesenchymal stem cells reside in the bone marrow and the adventitial layer of arteries. Endothelial progenitor cells that originate from the bone marrow are an important mechanism for repairing injured endothelial cells. Mesenchymal stem cells may differentiate osteogenically by inflammation or by specific stimuli, which can activate calcification. However, the bioactive substances secreted from mesenchymal stem cells have been shown to mitigate vascular calcification by suppressing inflammation, bone morphogenetic protein 2, and the Wingless-INT signal. Vitamin D deficiency may contribute to vascular calcification. Vitamin D supplement has been used to modulate the osteoblastic differentiation of mesenchymal stem cells and to lessen vascular injury by stimulating adhesion and migration of endothelial progenitor cells. This narrative review clarifies the role of mesenchymal stem cells and the possible role of vitamin D in the mechanisms of vascular calcification.

Dietary Supplemental Glutamine Enhances the Percentage of Circulating Endothelial Progenitor Cells in Mice with High-Fat Diet-Induced Obesity Subjected to Hind Limb Ischemia.

This study investigated whether glutamine (GLN) pretreatment can enhance circulating endothelial progenitor cells (EPCs) and attenuate inflammatory reaction in high-fat diet-induced obese mice with limb ischemia. Mice were assigned to a normal control (NC), high-fat control (HC), limb ischemia (HI), and GLN limb ischemia (HG) groups. The NC group provided chow diet and treated as a negative control. Mice in the HC and HI groups were fed a high-fat diet which 60% energy provided by fat for 8 weeks. Mice in the HG group were fed the same diet for 4 weeks and then transferred to a high-fat diet with 25% of total protein nitrogen provided as GLN to replace part of the casein for the subsequent 4 weeks. After feeding 8 weeks, mice in the HC group were sham-operated, while the HI and HG groups underwent an operation to induce limb ischemia. All mice except the NC group were euthanized on either day 1 or 7 after the operation. The results showed that the 8 weeks' high-fat diet feeding resulted in obesity. The HG group had higher circulating EPCs on day 1 while muscle vascular endothelial growth factor, matrix metalloproteinase-9, and hypoxia-inducible factor-1 gene expressions were higher on day 7 postischemia than those of the HI group. The superoxide dismutase activity and reduced glutathione content in affected muscles were higher, whereas mRNA expressions of interleukin-6 and tumor necrosis factor-α were lower in the HG than those in the HI group. These findings suggest that obese mice pretreated with GLN-supplemented high-fat diet increased circulating EPC percentage, enhanced the antioxidant capacity, and attenuated inflammatory reactions in response to limb ischemia.

Garcimultiflorone K inhibits angiogenesis through Akt/eNOS- and mTOR-dependent pathways in human endothelial progenitor cells.

Background Garcimultiflorone K is a novel polyprenylated polycyclic acylphloroglucinol isolated from the stems of Garcinia multiflora that exhibits promising anti-angiogenic activity in human endothelial progenitor cells (EPCs). Purpose This study sought to determine the underlying anti-angiogenic mechanisms and pharmacological properties of garcimultiflorone K. Methods We examined the anti-angiogenic effects of garcimultiflorone K and its mechanisms of action using in vitro EPC models and in vivo zebrafish embryos. Results EPCs proliferation, migration, differentiation and capillary-like tube formation were effectively and concentration-dependently inhibited by garcimultiflorone K without any signs of cytotoxicity. Our investigations revealed that garcimultiflorone K suppressed EPCs angiogenesis through Akt, mTOR, p70S6K, and eNOS signaling cascades. Notably, garcimultiflorone K dose-dependently impeded angiogenesis in zebrafish embryos. Conclusion Our data demonstrate the anti-angiogneic effects of garcimultiflorone K in both in vitro and in vivo models. Garcimultiflorone K appears to have potential in the treatment of angiogenesis-related diseases.

Vitamin D (1,25-(OH)2D3) regulates the gene expression through competing endogenous RNAs networks in high glucose-treated endothelial progenitor cells.

Vitamin D (vit-D) supplementation can improve endothelial cell function in type 2 diabetes mellitus patients with vit-D insufficiency or deficiency. In the present study, we aimed to compare the expression profiles of circRNAs, lncRNAs, miRNAs, and mRNAs between 1,25-(OH)2D3-treated endothelial progenitor cells (EPCs) and control cells, and to further construct the 1,25-(OH)2D3-regulated ceRNA networks in EPCs. RNA sequencing was performed on the 1,25-(OH)2D3-treated EPCs and control cells derived from the bone marrow (BM). Bioinformatics analyses were performed to identify differentially expressed (DE) microRNAs (miRNAs), circular RNAs (circRNAs), mRNAs, and long non-coding RNAs (lncRNAs). Then Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were conducted to predict the function of genes. Competing endogenous RNA (ceRNA) networks were constructed with Cytoscape software. 1,25-(OH)2D3 application induced changes in the expression profiles of 1791 mRNAs, 2726 lncRNAs, 205 circRNAs, and 45 miRNAs in EPCs treated with high levels of glucose. These DE RNAs were associated with MMP and GTPase activities, specific signaling pathways, and components of actin, extracellular matrix, or adherens junction. DE circRNAs, which functioned independently of their linear host genes, interacted with miRNAs to serve as miRNA sponges in complex ceRNA networks. The data indicated that circRNAs and lncRNAs comprised ceRNAs to sponge effects of miRNAs on the expressions of mRNAs following 1,25-(OH)2D3 application in EPCs. 1,25-(OH)2D3 improved the function of EPCs via associated ceRNA interaction networks in diabetes patients.

Danhong injection mobilizes endothelial progenitor cells to repair vascular endothelium injury via upregulating the expression of Akt, eNOS and MMP-9.

BACKGROUD: Endothelial progenitor cells (EPCs) have been characterized as one of the key effectors of endothelial healing. The effect of Danhong injection (DHI), the most widely prescribed Chinese medicine for coronary heart disease (CHD), on EPCs mobilization remains unclear. PURPOSE: We aimed to assess the effect of DHI on EPCs mobilization to repair percutaneous coronary intervention (PCI) induced vascular injury, and to investigate the characteristics and potential mechanism of DHI on EPCs mobilization. METHOD: Forty-two patients with CHD underwent PCI and received stent implantation were enrolled in a Phase II clinical trials. All patients received routine western medical treatment after PCI, patients of DHI group received DHI in addition. The levels of CECs, cytokines (vWF, IL-6, CRP) and EPCs were analyzed at baseline, post-PCI and after treatment. To investigate the characteristics of DHI on EPCs mobilization, 12 healthy volunteers received intravenous infusion of DHI once and the other 12 received for 7 days. EPCs enumeration were done at a series of time points. At last we tested the effect of DHI and three chemical constituents of DHI (danshensu; lithospermic acid, LA; salvianolic acid D, SaD) on EPCs level and expression of Akt, eNOS and MMP-9 in bone marrow cells of myocardial infarction (MI) mice. RESULTS: In the DHI group the angina symptoms were improved, the levels of cytokines and CECs were reduced; while EPCs population was increased after treatment. In the phase I clinical trials, EPCs counts reached a plateau phase in 9 h and maintained for more than 10 h after a single dose. After continuous administration, EPCs levels plateaued on the 3rd or 4th day, and maintain till 1 day after the withdrawal, then its levels gradually declined. DHI treatment induced a timely dependent mobilization of EPCs. DHI promoted EPCs mobilization via upregulating the expression of Akt, eNOS and MMP-9 in BM. LA and SaD have played a valuable role in EPCs mobilization. CONCLUSION: These initial results demonstrated that DHI is effective in alleviating endothelial injury and promoting endothelial repair through enhancing EPCs mobilization and revealed the effect feature and possible mechanisms of DHI in mobilizing EPCs.

Berberine-Promoted CXCR4 Expression Accelerates Endothelial Repair Capacity of Early Endothelial Progenitor Cells in Persons with Prehypertension.

OBJECTIVE: To evaluate whether the berberine treatment can improve endothelial repair capacity of early endothelial progenitor cells (EPCs) from prehypertensive subjects through increasing CXC chemokine receptor 4 (CXCR4) signaling. METHODS: EPCs were isolated from prehypertensive and healthy subjects and cultured. In vivo reendothelialization capacity of EPCs from prehypertensive patients with or without in vitro berberine treatment was examined in a nude mouse model of carotid artery injury. The protein expressions of CXCR4/Janus kinase-2 (JAK-2) signaling of in vitro EPCs were detected by Western blot analysis. RESULTS: CXCR4 signaling and alteration in migration and adhesion functions of EPCs were evaluated. Basal CXCR4 expression was significantly reduced in EPCs from prehypertensive patients compared with normal subjects (P<0.01). Also, the phosphorylation of JAK-2 of EPCs, a CXCR4 downstream signaling, was significantly decreased (P<0.01). Berberine promoted CXCR4/JAK-2 signaling expression of in vitro EPCs (P<0.01). Transplantation of EPCs pretreated with berberine markedly accelerated in vivo reendothelialization (P<0.01). The increased in vitro function and in vivo reendothelialization capacity of EPCs were inhibited by CXCR4 neutralizing antibody or pretreatment with JAK-2 inhibitor AG490, respectively (P<0.01). CONCLUSION: Berberinemodified EPCs via up-regulation of CXCR4 signaling contributes to enhanced endothelial repair capacity in prehypertension, indicating that berberine may be used as a novel potential primary prevention means against prehypertension-related atherosclerotic cardiovascular disease.

Supplementation with a selective amino acid formula ameliorates muscular dystrophy in mdx mice.

Duchenne muscular dystrophy (DMD) is one of the most common and severe forms of muscular dystrophy. Oxidative myofibre content, muscle vasculature architecture and exercise tolerance are impaired in DMD. Several studies have demonstrated that nutrient supplements ameliorate dystrophic features, thereby enhancing muscle performance. Here, we report that dietary supplementation with a specific branched-chain amino acid-enriched mixture (BCAAem) increased the abundance of oxidative muscle fibres associated with increased muscle endurance in dystrophic mdx mice. Amelioration of the fatigue index in BCAAem-treated mdx mice was caused by a cascade of events in the muscle tissue, which were promoted by endothelial nitric oxide synthase (eNOS) activation and vascular endothelial growth factor (VEGF) expression. VEGF induction led to recruitment of bone marrow (BM)-derived endothelial progenitors (EPs), which increased the capillary density of dystrophic skeletal muscle. Functionally, BCAAem mitigated the dystrophic phenotype of mdx mice without inducing dystrophin protein expression or replacing the dystrophin-associated glycoprotein (DAG) complex in the membrane, which is typically lost in DMD. BCAAem supplementation could be an effective adjuvant strategy in DMD treatment.

Components from the Leaves and Twigs of Mangrove Lumnitzera racemosa with Anti-Angiogenic and Anti-Inflammatory Effects.

One new neolignan, racelactone A (1), together with seven known compounds (2-8) were isolated from the methanolic extract of the leaves and twigs of Lumnitzera racemosa. The structure of racelactone A (1) was determined on the basis of the mass and NMR spectroscopic data interpretation. With respect to bioactivity, compound 1 displayed an anti-angiogenic effect by suppressing tube formation. Furthermore, compounds 1, 4, and 5 showed significant anti-inflammatory effects with IC50 values of 4.95 ± 0.89, 1.95 ± 0.40, and 2.57 ± 0.23 µM, respectively. The plausible biosynthesis pathway of racelactone A (1) was proposed.

Construction of a small-caliber tissue-engineered blood vessel using icariin-loaded ß-cyclodextrin sulfate for in situ anticoagulation and endothelialization.

The rapid endothelialization of tissue-engineered blood vessels (TEBVs) can effectively prevent thrombosis and inhibit intimal hyperplasia. The traditional Chinese medicine ingredient icariin is highly promising for the treatment of cardiovascular diseases. ß-cyclodextrin sulfate is a type of hollow molecule that has good biocompatibility and anticoagulation properties and exhibits a sustained release of icariin. We studied whether icariin-loaded ß-cyclodextrin sulfate can promote the endothelialization of TEBVs. The experimental results showed that icariin could significantly promote the proliferation and migration of endothelial progenitor cells; at the same time, icariin could promote the migration of rat vascular endothelial cells (RAVECs). Subsequently, we used an electrostatic force to modify the surface of the TEBVs with icariin-loaded ß-cyclodextrin sulfate, and these vessels were implanted into the rat common carotid artery. After 3 months, micro-CT results showed that the TEBVs modified using icariin-loaded ß-cyclodextrin sulfate had a greater patency rate. Scanning electron microscopy (SEM) and CD31 immunofluorescence results showed a better degree of endothelialization. Taken together, icariin-loaded ß-cyclodextrin sulfate can achieve anticoagulation and rapid endothelialization of TEBVs to ensure their long-term patency.

Monotropein promotes angiogenesis and inhibits oxidative stress-induced autophagy in endothelial progenitor cells to accelerate wound healing.

Attenuating oxidative stress-induced damage and promoting endothelial progenitor cell (EPC) differentiation are critical for ischaemic injuries. We suggested monotropein (Mtp), a bioactive constituent used in traditional Chinese medicine, can inhibit oxidative stress-induced mitochondrial dysfunction and stimulate bone marrow-derived EPC (BM-EPC) differentiation. Results showed Mtp significantly elevated migration and tube formation of BM-EPCs and prevented tert-butyl hydroperoxide (TBHP)-induced programmed cell death through apoptosis and autophagy by reducing intracellular reactive oxygen species release and restoring mitochondrial membrane potential, which may be mediated viamTOR/p70S6K/4EBP1 and AMPK phosphorylation. Moreover, Mtp accelerated wound healing in rats, as indicated by reduced healing times, decreased macrophage infiltration and increased blood vessel formation. In summary, Mtp promoted mobilization and differentiation of BM-EPCs and protected against apoptosis and autophagy by suppressing the AMPK/mTOR pathway, improving wound healing in vivo. This study revealed that Mtp is a potential therapeutic for endothelial injury-related wounds.

Evaluation of velvet antler total protein effect on bone marrow­derived endothelial progenitor cells.

Lu Rong, velvet antler (VA), is a traditional Chinese medicine, which is used as a food supplement and therapeutic drug in China, Japan, Russia, New Zealand and Southeast Asia. The regenerative characteristics of VA have resulted in great research interest, particularly regarding the fields of organ grafting and stem cell differentiation. Various VA proteomic studies verified that proteins act as the primary bioactive components of VA. The present study aimed to investigate if VA proteins (VA­pro) influence endothelial progenitor cell (EPC) viability. Various methods have previously been used to investigate VA­pro, including freeze­drying technology, ultrasonic wave methods, high performance liquid chromatography­mass spectrometry, EPCs extraction and culture. Results demonstrated that VA­pro promoted EPCs proliferation and migration, particularly at a concentration of 1 mg/ml. Furthermore, VA­pro increased the activation level of Notch1 intracellular domain and Hes1, and the level of phosphorylated­Akt and phosphorylated­mechanistic target of rapamycin. VA­pro may therefore affect EPC viability via regulation of the Notch and Akt signaling pathways. The present study revealed the effects and potential molecular mechanism of VA­pro on EPCs, and suggested an association between VA regeneration characteristics and the optimization of EPC viability. These findings may contribute to EPC transplantation research and aid in providing a novel treatment method for vascular diseases in the future.

Shexiang Baoxin pills promotes angiogenesis in myocardial infarction rats via up-regulation of 20-HETE-mediated endothelial progenitor cells mobilization.

BACKGROUND AND AIMS: Therapeutic angiogenesis is a pivotal strategy for ischemic heart disease. The aim of the present study was to determine the effect and molecular mechanism of Shexiang Baoxin pills, a widely-used traditional Chinese medicine for ischemic heart disease, on angiogenesis in a rat model of myocardial infarction (MI). METHODS: We used the occlusion of left anterior descending coronary artery of Sprague-Dawley rats as a model of MI. The MI rats were treated with distilled water, Shexiang Baoxin pills, or Shexiang Baoxin pills + HET0016 (a selective blocker of the biosynthesis of 20-hydroxyeicosatetraenoic acid (20-HETE) at 10 mg/kg/day), respectively. Sham-operated rats were used as controls. RESULTS: Treatment with Shexiang Baoxin pills increases the level of serum 20-HETE in MI rats, which can be suppressed by HET0016 treatment. Shexiang Baoxin pills shows cardio-protective effects on MI rats, including improving cardiac function, decreasing infarction area, and promoting angiogenesis in peri-infarct area. The protective effects of Shexiang Baoxin pills are partly inhibited by HET0016. Furthermore, Shexiang Baoxin pills enhances the number of circulating endothelial progenitor cells (EPCs) and the expression of the vascular endothelial growth factor (VEGF), based on immunohistochemical analysis, in peri-infarct area of MI rats, which is partly suppressed by HET0016. CONCLUSIONS: Shexiang Baoxin pills may partially participate in angiogenesis in MI rats. The protective mechanism of Shexiang Baoxin pills may be mediated via up-regulation of 20-HETE, which promotes EPCs mobilization and VEGF expression.

Effect of vitamin D on endothelial progenitor cells function.

BACKGROUND: Endothelial 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. AIM: To examine the in vitro effects of Calcitriol on EPCs from healthy subjects and patients with diabetes. METHODS: Fifty-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. RESULTS: In 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. CONCLUSION: In 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.

The acute impact of high-dose lipid-lowering treatment on endothelial progenitor cells in patients with coronary artery disease-The REMEDY-EPC early substudy.

RATIONALE AND OBJECTIVE: Endothelial progenitor cells (EPCs) play a role in vascular repair, while circulating endothelial cells (CECs) are biomarkers of vascular damage and regeneration. Statins may promote EPC/CEC mobilization in the peripheral blood. We evaluated whether pre-procedural exposure to different lipid-lowering drugs (statins±ezetimibe) can acutely increase levels/activity of EPCs/CECs in patients with stable coronary artery disease (CAD). METHODS: In a planned sub-analysis of the Rosuvastatin For REduction Of Myocardial DamagE During Coronary AngioplastY (REMEDY) trial, 38 patients with stable CAD on chronic low-dose statin therapy were randomized, in a double-blind, placebo-controlled design, into 4 groups before PCI: i. placebo (n = 11); ii. atorvastatin (80 mg+40 mg, n = 9); iii. rosuvastatin (40 mg twice, n = 9); and iv. rosuvastatin (5 mg) and ezetimibe (10 mg) twice, (n = 9). At baseline and 24 h after treatment-before PCI-, patients underwent blinded analyses of EPCs [colony forming units-endothelial cells (CFU-ECs), endothelial colony-forming cells (ECFCs) and tubulization activity] and CECs in peripheral blood. RESULTS: We found no significant treatment effects on parameters investigated such as number of CECs [Median (IQR): i. 0(0), ii. 4.5(27), iii. 1.9(2.3), iv. 1.9(2.3)], CFU-ECs [Median (IQR): i. 27(11), ii. 19(31), iii. 47(36), iv. 30(98)], and ECFCs [Median (IQR): i. 86(84), ii. 7(84), iii. 8/(42.5), iv. 5(2)], as well as tubulization activity [total tubuli (well), Median (IQR): i. 19(7), ii. 5(4), iii. 25(13), iv. 15(24)]. CONCLUSIONS: In this study, we found no evidence of acute changes in levels or activity of EPCs and CECs after high-dose lipid-lowering therapy in stable CAD patients.