Endothelial Progenitor Cells Treatment Research Articles

Local application reduces number of needed EPC for beneficial effects on wound healing compared to systemic treatment in mice

IntroductionStem cell transplantation is one of the most promising strategies to improve healing in chronic wounds as systemic administration of endothelial progenitor cells (EPC) enhances healing by promoting neovascularization and homing though a high amount of cells is needed. In the following study, we analysed whether local application can reduce the number of EPC needed achieving the same beneficial effect on wound healing.Material and MethodsWound healing after local or systemic treatment with EPC was monitored in vivo by creating standardized wounds on the dorsum of hairless mice measuring wound closure every second day. Systemic group received 2 × 106 EPC i.v. and locally treated group 2 × 105 EPC, locally injected. As control PBS injection was performed the same way. Expression of CD31, VEGF, CD90 and, SDF-1α was analysed immunohistochemically for evaluation of neovascularisation and amelioration of homing.ResultsLocal (7.1 ± 0.45 SD) as well as systemic (6.1 ± 0.23 SD) EPC transplantation led to a significant acceleration of wound closure compared to controls (PBS local: 9.7 ± 0.5 SD, PBS systemic 10.9 ± 0.38 SD). Systemic application enhanced CD31 expression on day 6 after wounding and local EPC on 6 and 9 in comparison to control. VEGF expression was not significantly affected. Systemic and local EPC treatment resulted in a significantly enhanced SDF-1α and CD90 expression on all days investigated.ConclusionLocal as well as systemic EPC treatment enhances wound healing. Moreover, beneficial effects are obtained with a tenfold decrease number of EPC when applied locally. Thus, local EPC treatment might be more convenient way to enhance wound healing as number of progenitor cells is limited.

Effects of endothelial progenitor cells transplantation on hyperlipidemia associated kidney damage in ApoE knockout mouse model

BackgroundHyperlipidaemia causes kidney damage over the long term. We investigated the effect of the administration of endothelial progenitor cells (EPCs) on the progression of kidney damage in a mouse model of hyperlipidaemia.MethodsApolipoprotein E-knockout (ApoE−/−) mice were treated with a high-cholesterol diet after spleen resection. Twenty-four weeks later, the mice were divided into two groups and intravenously injected with PBS or EPCs. Six weeks later, the recruitment of EPCs to the kidney was monitored by immunofluorescence. The lipid, endothelial cell, and collagen contents in the kidney were evaluated by specific immunostaining. The protein expression levels of transforming growth factor-β (TGF-β), Smad2/3, and phospho-Smad3 (p-smad3) were detected by western blot analysis.ResultsApoE−/− mice treated with a high-fat diet demonstrated glomerular lipid deposition, enlargement of the glomerular mesangial matrix, endothelial cell enlargement accompanied by vacuolar degeneration and an area of interstitial collagen in the kidney. Six weeks after EPC treatment, only a few EPCs were detected in the kidney tissues of ApoE−/− mice, mainly in the kidney interstitial area. No significant differences in TGF-β, p-smad3 or smad2/3 expression were found between the PBS group and the EPC treatment group (TGF-β expression, PBS group: 1.06 ± 0.09, EPC treatment group: 1.09 ± 0.17, P = 0.787; p-smad3/smad2/3 expression: PBS group: 1.11 ± 0.41, EPC treatment group: 1.05 ± 0.33, P = 0.861).ConclusionsOur findings demonstrate that hyperlipidaemia causes basement membrane thickening, glomerulosclerosis and the vascular degeneration of endothelial cells. The long-term administration of EPCs substantially has limited effect in the progression of kidney damage in a mouse model of hyperlipidaemia.

The Protective Effect of Adiponectin-Transfected Endothelial Progenitor Cells on Cognitive Function in D-Galactose-Induced Aging Rats.

Aging is a multifactorial process involving the cumulative effects of inflammation, oxidative stress, and mitochondrial dynamics, which can produce complex structural and biochemical alterations to the nervous system and lead to dysfunction of microcirculation, blood-brain barrier (BBB), and other problems in the brain. Long-term injection of D-galactose (D-gal) can induce chronic inflammation and oxidative stress, accelerating aging. The model of accelerated aging with long-term administration of D-gal have been widely used in anti-aging studies, due to the increase of chronic inflammation and decline of cognition that similarity with natural aging in animals. However, despite extensive researches in the D-gal-induced aging rats, studies on their microvasculature remain limited. Endothelial progenitor cells (EPCs), which are precursors to endothelial cells (ECs), play a significant role in the repair and regeneration process of endogenous blood vessel, and adiponectin (APN), a protein derived from adipocyte, has many effects on protective vascular endothelium and anti-inflammatory. Recently, many studies have shown that APN can promote improvements in cognitive function. Under these circumstances, we investigated the neuroprotective effect of the APN-transfected EPC (APN-EPC) treatment on rats after administration with D-gal and explored the likely underlying mechanisms. Compared to model group for D-gal administration, better cognitive function and denser microvessels were significantly found in the APN-EPC treatment group, and indicated APN-EPC treatment in aging rats could improve the cognitive dysfunction and microvessel density. The level of proinflammatory cytokines IL-1β, IL-6, and TNF-α, activated astrocytes and apoptosis rate were significantly reduced in the APN-EPC group compared with the model group, showed that APN-EPCs alleviated the neuroinflammation in aging rats. In addition, the APN-EPC group inhibited the decrease of BBB-related proteins claudin-5, occludin, and Zo-1 in aging rats and attenuated BBB dysfunction significantly. These results of our study indicated that APN-EPC treatment in D-gal-induced aging rats have a positive effect on improving cognitive and BBB dysfunction, increasing angiogenesis, and reducing neuroinflammation and apoptosis rate. This research suggests that cell therapy via gene modification may provide a safe and effective approach for the treatment of age-related neurogenerative diseases.

Transplantation of Endothelial Progenitor Cells in the Treatment of Coronary Artery Microembolism in Rats

As the impairment of myocardial microenvironments due to coronary microembolization (CME) compromises the treatment effect of percutaneous coronary intervention and leads to adverse prognosis, we hypothesized that endothelial progenitor cells (EPCs) transplantation could improve cardiac function in the condition of CME. Low- (2 × 105) and high- (2 × 106) dose rat bone marrow-derived EPCs were transplanted in a model of CME. To develop a CME model, rats were injected with autologous micro-blood-clots into the left ventricle. Echocardiograph was examined before and 1, 7, and 28 days after EPC transplantation; serum cardiac troponin I (cTNI), von Willebrand factor (vWF), and cardiac microRNA expression were examined one day after EPCs transplantation. Heart morphology and vascular endothelial growth factor (VEGF), vWF, and basic fibroblast growth factor (bFGF) expression were examined one day after EPC transplantation. After 10 days of culture inductions, BM-EPCs have high purity as confirmed by flow cytometry. Cardiac function reflected by left ventricular ejection fraction significantly decreased after CME treatment and rescued by low-dose EPC. Compared to the sham group, cTNI and vWF serum levels increased significantly after CME treatment and rescued by low-dose EPC and high-dose EPC. Low-dose EPC treatment decreased myocardial necrosis and fibrosis and elevated cardiac expression of VEGF and vWF, while decreasing the cardiac expression of bFGF. Low-dose EPC treatment significantly suppressed cardiac expression of microRNA-19a but significantly enhanced microRNA-21, microRNA-214, and microRNA-486-3p expression. In conclusion, our results indicate that low-dose EPC transplantation may play a proangiogenic, antifibroblast, antifibrosis, and antinecrosis role and enhance cardiac function in a rat model of CME through a microRNA-related pathway.

P5386Transplantation of endothelial progenitor cells in the treatment of coronary artery microembolism in rats

Abstract Objective As the impairment of myocardial micro-environments due to coronary micro-embolization (CME) compromises the treatment effect of percutaneous coronary intervention and leads to adverse prognosis, we hypothesized that endothelial progenitor cells (EPCs) transplantation could improve cardiac function in the condition of CME. Materials and methods Low- (2×105) and high- (2×106) dose rat bone marrow-derived EPCs were transplanted in a model of CME. To develop a CME model, rats were injected with autologous micro-blood clots into the left ventricle. Echocardiography was examined before and 1, 7, and 28 days after EPC transplantation; serum cTNI, vWF,and cardiac microRNA expression were examined one day after EPCs transplantation. Heart morphology and VEGF, vWF, and bFGF expression were examined one day after EPC transplantation. Figure 1. Results Results After 10 days of culture inductions, BM-EPCs have high purity as confirmed by flow cytometry. Cardiac function reflected by LVEF significantly decreased after CME treatment and rescued by low-dose EPC. Compared to the sham group, cTNI, and vWF serum levels increased significantly after CME treatment and rescued by low-dose EPC and high-dose EPC. Low-dose EPC treatment decreased myocardial necrosis and fibrosis and elevated cardiac expression of VEGF and vWF, while decreasing the cardiac expression of bFGF. Low-dose EPC treatment significantly suppressed cardiac expression of microRNA-19a but significantly enhanced microRNA-21, microRNA-214, and microRNA-486-3p expression. In conclusion, our results indicate that low-dose EPC transplantation may play a pro-angiogenic, anti-fibroblast, anti-fibrosis, and anti-necrosis role and enhance cardiac function in a rat model of CME through a microRNA-related pathway. Acknowledgement/Funding Supported by the State Key Laboratory of Low-Dimensional Quantum Physics Open Project Fund (No. KF201608)

Dickkopf-1 Treatment Stimulates Hematopoietic Regenerative Function in Infused Endothelial Progenitor Cells.

Acute high-dose radiation injury damages the bone marrow hematopoietic stem and progenitor cell compartment. This damage compromises the functional ability of the bone marrow to produce mature blood cells and results in an increased risk of death due to hematopoietic complications. Past work has shown that the bone marrow endothelium provides critical cues, which promote hematopoietic stem cell regeneration after injury. Additionally, transfusion of endothelial cells after radiation injury has been shown to promote recovery of both the bone marrow vasculature and hematopoietic systems. In this work, we examined the regenerative capacity of intravenous infusion of umbilical cord-blood derived endothelial progenitor cells (EPCs) since this is a cell source which is easy to obtain, expand and cryopreserve. We show that pre-treatment with the Wnt-antagonist Dickkopf1 (Dkk1) augments EPC regenerative function in an allogeneic mouse transplant model. Here, hematopoietic recovery was assessed in Balb/c mice after 5 Gy total-body irradiation and transplantation with C57/BL6-derived EPCs either with or without Dkk1 pre-treatment. The Dkk1-treated EPC group had significantly faster recovery of peripheral white blood cells, total bone marrow cellularity, bone marrow progenitors and BM endothelial cells compared to EPC treatment alone or saline controls. Importantly, after an LD50/30 dose of 8 Gy in the Balb/c mouse, Dkk1-treated EPCs were able to rescue 100% of irradiated mice versus 80% in the EPC control group and only 33% in the saline-treated group. To understand how Dkk1 induces regenerative function in the EPCs, we screened for pro-regenerative factors secreted by the EPC in response to Dkk1. Dkk1-treated EPCs were observed to secrete high levels of the anti-fibrotic protein follistatin as well as several proteins known to promote regeneration including EGF, VEGF and G-CSF. This work demonstrates the potential for Dkk1-treated EPCs as a rescue therapeutic for victims of acute radiation injury.

Endothelial Progenitor Cells Transplantation Promotes the Whole Brain Recovery after Focal Ischemic Stroke

Background: Ischemic stroke caused severe damages to patients, mostly treatments for ischemic stroke focused recovery on the peri-infarct areas. In this study, we found that focal ischemic stroke caused damages to whole brain, endothelial progenitor cells (EPCs) treatment could significantly promote the recovery of the whole brain after ischemic stroke. Methods: EPCs were infected with lentivirus carrying luc2 and eGFP cassette for bioluminescence imaging (BLI) and labeled with Alkyl-PEI-SPIO for magnetic resonance imaging (MRI). 5×10^5 EPCs were transplanted intracardially 24 hours after photothrombotic ischemic stroke onset. BLI, MRI and behavior test were carried out at different time points. Expression of BDNF, VEGF in the ipsilateral and contralateral hemisphere and infarct volume were analyzed 7 days after EPCs transplantation. Expression of myelin basic protein (MBP) and NeuN in the ipsilateral and contralateral hemisphere were detected 14 days after EPCs transplantation. Findings: EPCs transplantation significantly increased the expression of BDNF, MBP and NeuN in both ipsilateral and contralateral hemisphere, while no significant differences were found in VEGF expression. Also, in vivo MRI results showed that fiber counts and FA values in both ipsilateral and contralateral corpus callosum increased 14 days after EPCs transplantation. No significant differences were found in the infarct volume 7 days after EPCs transplantation. mNSS scores and foot-faults test indicated better functional recovery 14 days after EPCs transplantation. Interpretation: Studies should focus on the whole brain recovery after stroke onset. EPCs treatment offered a powerful therapy method for whole brain recovery after ischemic stroke which may associated with elevated BDNF expression. Funding Statement: This work was supported by the National Key Basic Research Program of China (NOs. 2013CB733800 and 2013CB733803), the National Natural Science Foundation of China (NOs. 81230034 and 81520108015), the National Natural Science Foundation of Jiangsu Province (NO. BK20130057), the National Science Foundation for Youths of China (NOs. 81601544 and 81501522), the Fundamental Research Funds for the Central Universities (NO. 2242018k30004). C. Wang acknowledged the financial support from the China Scholarship Council for his joint PhD scholarship (NO. 201706090204). The funder had no role in the study design, data analysis, write-up or in the writing of the report. The corresponding author had full access to all the data in the study and had final responsibility for the decision to submit for publication. Declaration of Interest: The authors have no conflicts of interest to disclose. Ethics Approval Statement: This study was approved by the Institutional Animal Care and Use Committee (IACUC) of Southeast University (approval ID: SYXK-2010.4987). BALB/c nude female mice (8 weeks, 20-23 g, Comparative Medicine Centre of Yangzhou University [SCXK (SU) 2012-0004], Yangzhou, China) were used in this study. Briefly, EPCs were isolated from newborns with all mothers’ informed consent and with the approval of the local hospital ethics committee.

Sustained release of endothelial progenitor cell-derived extracellular vesicles from shear-thinning hydrogels improves angiogenesis and promotes function after myocardial infarction.

Previous studies have demonstrated improved cardiac function following myocardial infarction (MI) after administration of endothelial progenitor cells (EPCs) into ischaemic myocardium. A growing body of literature supports paracrine effectors, including extracellular vesicles (EVs), as the main mediators of the therapeutic benefits of EPCs. The direct use of paracrine factors is an attractive strategy that harnesses the effects of cell therapy without concerns of cell engraftment or viability. We aim to reproduce the beneficial effects of EPC treatment through delivery of EPC-derived EVs within a shear-thinning gel (STG) for precise localization and sustained delivery. EVs were harvested from EPCs isolated from adult male Rattus norvegicus (Wistar) rats and characterized by electron microscopy, nanoparticle tracking analysis (NTA), and mass spectrometry. EVs were incorporated into the STG and injected at the border zone in rat models of MI. Haemodynamic function, angiogenesis, and myocardial remodelling were analyzed in five groups: phosphate buffered saline (PBS) control, STG control, EVs in PBS, EVs in STG, and EPCs in STG. Electron microscopy and NTA of EVs showed uniform particles of 50-200 nm. EV content analysis revealed several key angiogenic mediators. EV uptake by endothelial cells was confirmed and followed by robust therapeutic angiogenesis. In vivo animal experiments demonstrated that delivery of EVs within the STG resulted in increased peri-infarct vascular proliferation, preservation of ventricular geometry, and improved haemodynamic function post-MI. EPC-derived EVs delivered into ischaemic myocardium via an injectable hydrogel enhanced peri-infarct angiogenesis and myocardial haemodynamics in a rat model of MI. The STG greatly increased therapeutic efficiency and efficacy of EV-mediated myocardial preservation.

Noninvasive evaluation of the migration effect of transplanted endothelial progenitor cells in ischemic muscle using a multimodal imaging agent.

BackgroundEndothelial progenitor cells (EPCs) play an important role in repairing ischemia tissues. However, the survival, migration and therapeutic efficacy of EPCs after transplantation need to be better understood for further cell therapy.PurposeThis study investigated the migration effect of EPCs labeled with a multimodal imaging agent in a murine ischemic hindlimb model, using magnetic resonance imaging (MRI) and optical imaging after transplantation.MethodsEPCs derived from mouse bone marrow were labeled with a multimodal imaging agent and were administered through intracardiac delivery to mice with ischemic hindlimbs. The injected EPCs and their migration effect were observed via MRI and optical imaging in vivo, and then compared to a reference standard based on histological data. The quantification of gadolinium in tissue samples was done using inductively coupled plasma mass spectrometry (ICP-MS).ResultsUsing in vivo MRI and optical imaging, the labeled EPCs were observed to migrate to ischemic muscle on days 3–5 after injection, while ex vivo, the EPCs were observed in the capillary vessels of the injured tissue. There were significant linear correlations between the Gd contents measured using ICP-MS in samples from the ischemic hindlimbs and livers and T1 relaxation times calculated using MRI, as well as the average fluorescence signal intensities recorded in optical images (T1 relaxation time: r=0.491; average signal from optical imaging: r=0.704, P<0.01). EPC treatment upregulated the levels of C-X-C chemokine receptor 4 and vascular endothelial growth factor (VEGF) receptor 2 and enhanced the expression of stromal cell-derived factor-1 and VEGF.ConclusionTransplanted EPCs can be monitored with noninvasive MRI and optical imaging in vivo and were found to enhance the paracrine secretion of angiogenic factors.

Pro-apoptotic effect of TRAIL-transfected endothelial progenitor cells on glioma cells.

Glioma is one of the most common aggressive neuroepithelial malignant tumors in the central nervous system. It has a high recurrence rate and poor prognosis, primarily due to the fact that novel therapeutic agents cannot penetrate the blood-brain barrier (BBB). Endothelial progenitor cells (EPCs) have been reported to move across the BBB and access the tumor site. However, whether EPCs expressing the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induce glioma cell apoptosis requires further investigation. In the present study, EPCs were transfected and stably expressed with TRAIL through lentiviral infection. The pro-apoptotic effect of these TRAIL-expressing EPCs on the SHG44 glioma cell line was investigated. The migration ability of TRAIL-expressing EPCs toward SHG44 cells through the Transwell culture system was investigated via a high-content screening assay. The apoptotic rate and the expression of cleaved caspase-8 and −3 in addition to the cleaved poly(ADP-ribose) polymerase in SHG44 cells significantly increased in the TRAIL-overexpressing EPC treatment group compared with the controls. The increased apoptotic rate was reversed using a caspase inhibitor. The findings suggested that the TRAIL-expressing EPCs induced apoptosis in the SHG44 cells by activating the death receptor pathway, indicating that the TRAIL-expressing EPCs may be a useful strategy for glioma treatment.

MicroRNA-126/stromal cell-derived factor 1/C-X-C chemokine receptor type 7 signaling pathway promotes post-stroke angiogenesis of endothelial progenitor cell transplantation

Stroke is the most common cause of mortality worldwide. Post-stroke angiogenesis is of great significance to the treatment of strokes. The aim of the present study was to investigate the mechanism underlying the angiogenesis-promoting effect of microRNA‑126 (miR‑126)‑associated signaling pathways using a stroke model invivo and a cell migration model invitro. Bone marrow‑derived endothelial progenitor cells (EPCs) were extracted and identified using a density gradient method. Reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) was performed to examine the expression levels of miR‑126 and C‑X‑C chemokine receptor type 7 (CXCR7). Target genes of miR‑126 were analyzed using TargetScan software version 7.1 (www.targetscan.org/). In addition, a reporter gene assay and RT‑qPCR were performed to determine the target genes of miR‑126. The effect of miR‑126 on cell migration was examined using a cell migration model invitro and a middle cerebral artery occlusion model of mice was established invivo. The miR‑126 antagomir‑treated EPCs were infused into stroke mice. Microvessel density, nerve function score and infarction volume were assessed. Flow cytometric analysis indicated that cluster of differentiation (CD)34, CD133 and vascular endothelial growth factor receptor 2 were partly expressed on the cell surface of bone marrow‑derived EPCs. In addition, the expression levels of Di‑acetylated‑low density lipoprotein and Ulex europaeus agglutinin 1 were positive. Stromal cell‑derived factor 1 (SDF-1) was identified as a target gene of miR‑126, which was confirmed by a reporter gene assay and RT‑qPCR. Cell migration examination demonstrated that the neutralizing antibody of CXCR7 blocked miR‑126 angomir‑induced migration of EPCs. Microvessel density increased, while nerve function score and infarction volume decreased following infusion of miR-126 angomir‑treated EPCs. Furthermore, miR‑126 angomir improved the efficacy of EPC treatment. Thus, miR‑126 improved the migration of EPCs via the miR‑126/SDF‑1/CXCR7 signaling pathway.

VEGF165 transfected endothelial progenitor cells mediated by lentivirus alleviated ALI in rats

To investigate the protective effects of vascular endothelial growth factor-165 (VEGF165) transfected the endothelial progenitor cells (EPCs) mediated by lentivirus on acute lung injury (ALI) in rats. The mononuclear cells from the male Sprague-Dawley (SD) rats were isolated and cultured to get the EPCs for study. The lentivirus vector carrying the human VEGF165 gene was constructed. According to the random number table method, 90 male SD rats were divided into ALI model group, phosphate buffer solution (PBS) group, EPCs treatment group, none transfected EPCs treatment group and VEGF165 transfected EPCs treatment group, and the rats in each group were subdivided into 4, 12 and 48 hours subgroups, with 6 rats in each subgroup. The rat model of ALI was reproduced by intravenous injection of oleic acid (0.15 μL/g). Then each treatment group was given PBS, EPCs, none transfected EPCs and VEGF165 transfected EPCs respectively with the same volume of 0.2 mL. For the groups with cells, about 1×106 cells were contained. Abdominal aortic blood and lung tissue were harvested at 4, 12 and 48 hours. Arterial blood gas analysis was performed. The lung wet/dry weight ratio (W/D) was calculated. The expressions of induced nitric oxide synthase (iNOS), endothelin-1 (ET-1) and VEGF165 were determined by enzyme-linked immunosorbent assay (ELISA). After dyed with hematoxylin-eosin (HE), the lung tissue pathology was observed and the lung injury score was performed. Compared with the ALI model group, the arterial partial pressure of oxygen (PaO2) in EPCs, none transfected EPCs and VEGF165 transfected EPCs treatment groups was significantly increased from 4 hours, and lung W/D, expressions of iNOS and ET-1 were significantly decreased, and VEGF165 expression was significantly increased. Compared with the EPCs treatment group, the increase in PaO2, the decrease in lung W/D and expressions of iNOS and ET-1, and the increase in VEGF165 expression in VEGF165 transfected EPCs treatment group were more significant [4 hours: PaO2 (mmHg, 1 mmHg = 0.133 kPa) was 82.84±10.69 vs. 72.34±9.36, lung W/D ratio was 4.83±0.23 vs. 5.55±0.37, iNOS (ng/mg) was 8.77±1.10 vs. 14.84±1.34, ET-1 (ng/mg) was 103.41±5.66 vs. 153.08±5.12, VEGF165 (ng/mg) was 130.56±12.16 vs. 83.03±5.95; 12 hours: PaO2 (mmHg) was 91.67±6.81 vs. 78.5±8.81, lung W/D ratio was 4.44±0.35 vs. 5.32±0.25, iNOS (ng/mg) was 7.23±0.24 vs. 14.04±1.18, ET-1 (ng/mg) was 91.98±3.52 vs. 125.99±7.55, VEGF165 (ng/mg) was 164.49±5.71 vs. 96.61±6.12]; individual parameters reached valley value or peak value at 48 hours [lung W/D ratio was 4.26±0.30 vs. 4.89±0.15, iNOS (ng/mg) was 5.79±0.85 vs. 12.72±1.10, ET-1 (ng/mg) was 74.53±7.10 vs. 108.33±5.84, VEGF165 (ng/mg) was 237.43±10.79 vs. 134.24±11.99, all P < 0.05]. Over time, lung tissue injury in each group was gradually increased, and the lung injury score was gradually increased. The lung injury score at 48 hours in the EPCs, none transfected EPCs and VEGF165 transfected EPCs treatment groups were lower than that in the ALI model group. Compared with the EPCs treatment group, the VEGF165 transfected EPCs treatment group had a lower score at 48 hours (8.50±1.05 vs. 10.50±1.05, P < 0.05). The transplantation of EPCs which were transfected with VEGF165 mediated by lentivirus could obviously improve the oxygen pressure, reduce the lung water seepage, decrease the iNOS and ET-1 expressions in lung tissue, and had obvious protective effects on ALI.

Intratracheal transplantation of endothelial progenitor cells attenuates smoking-induced COPD in mice.

BackgroundEndothelial progenitor cells (EPCs) might play a protective role in COPD. The aim of this study was to investigate whether intratracheal allogeneic transplantation of bone-marrow-derived EPCs would attenuate the development of smoking-induced COPD in mice.MethodsIsolated mononuclear cells from the bone marrow of C57BL/6J mice were cultured in endothelial cell growth medium-2 for 10 days, yielding EPCs. A murine model of COPD was established by passive 90-day exposure of cigarette smoke. On day 30, EPCs or phosphate-buffered saline alone was administered into the trachea. On day 90, EPCs or 30 μL phosphate-buffered saline alone was administered into the trachea, and on day 120, inflammatory cells, antioxidant activity, apoptosis, matrix metalloproteinase (MMP)-2, and MMP-9 were measured.ResultsAfter EPC treatment, the lung function of the mice had improved compared with the untreated mice. Mean linear intercept and destructive index were reduced in the EPCs-treated group compared with the untreated group. In addition, the EPCs-treated mice exhibited less antioxidant activity in bronchoalveolar lavage fluid compared with the untreated mice. Moreover, decreased activities of MMP-2, MMP-9, and TUNEL-positive cells in lung tissues were detected in EPCs-treated mice.ConclusionIntratracheal transplantation of EPCs attenuated the development of pulmonary emphysema and lung function disorder probably by alleviating inflammatory infiltration, decelerating apoptosis, inhibiting proteolytic enzyme activity, and improving antioxidant activity.

Constitutive Expression of Adiponectin in Endothelial Progenitor Cells Protects a Rat Model of Cerebral Ischemia

Endothelial progenitor cells (EPCs), as precursors to endothelial cells, play a significant part in the process of endogenous blood vessel repair and maintenance of endothelial integrity. Adiponectin (APN) is an adipocyte-specific adipocytokine. In this study, we aim to test whether we transplant a combined graft of EPCs transfected with the adiponectin gene into a rat model of cerebral ischemia could improve functional recovery after middle cerebral artery occlusion (MCAO). Sprague-Dawley (SD) rats were randomly divided into a MCAO control group, a MCAO EPC treatment group, and a MCAO LV-APN-EPC treatment group. A focal cerebral ischemia and reperfusion model was induced by the intraluminal suture method. After 2 h of reperfusion, EPCs were transplanted by injection through the tail vein. A rotarod test was conducted to assess behavioral function before MCAO and on days 1, 7, and 14 after MCAO. After 14 d, TTC staining, CD31 immunofluorescence, and TUNEL staining were used to evaluate infarct volume, microvessel density, and cell apoptosis. Results revealed that behavioral function, infarct area percentage, microvessel density, and cell apoptosis rates were more favorable in the LV-APN-EPC treatment group than in the EPC treatment group. These data suggested that gene-modified cell therapy may be a useful approach for the treatment of ischemic stroke.

Human Endothelial Progenitor Cells as a Novel Therapeutic Option to Treat Severe Erectile Dysfunction in Obese Type 2 Diabetic Rats

Erectile dysfunction (ED) is a consequence of a number of medical conditions, with a high prevalence among men with type II diabetes. We investigated the effect of intracavernous injection of human umbilical cord blood derived endothelial progenitor cells (EPCs) on ED in a type II diabetic rat model. Erectile function was first assessed by cavernous nerve electrostimulation in Zucker Diabetic Fat (ZDF) rats aged 24 weeks to establish the baseline intracavernous pressure (ICP). At the age of 28 weeks, severe ED was confirmed. ZDF rats at age 29 weeks were randomised into 3 groups: treatment with 1million and 2million EPCs, and control, PBS. The injected animals were maintained with daily cyclosporine (10mg/kg) for 4 weeks, followed by the evaluation of erectile function and immunohistochemistry of penile tissue. The maximum (mean±SD) ICP observed for EPCs treated animals were significantly higher, 24.7±7.4 and 42.1±5.2 mmHg for 1million and 2million EPCs respectively in comparison to 4.4±1.6 mmHg in the age-matched PBS treated animals (p&lt;0.001). EPCs treatment also improved the smooth muscle content of the cavernosum (p&lt;0.001) and the expression of eNOS, RECA-1 and nNOS (p&lt;0.05) compared to the PBS control. In conclusion, we showed for the first time that human cord blood derived EPCs significantly improved severe ED in ZDF rats in a dose-dependent manner through improvement of the nerve and endothelium function and also through preservation of smooth muscle cells in the cavernosum.

Transplanted Endothelial Progenitor Cells Improve Ischemia Muscle Regeneration in Mice by Diffusion Tensor MR Imaging

Endothelial progenitor cells (EPCs) play an important role in repairing ischemia tissues. Diffusion tensor imaging (DTI) was applied to detect the architectural organization of skeletal muscle. This study investigated the feasibility and accuracy of using the DTI to evaluate effectiveness of EPCs treatment. Mouse bone marrow-derived EPCs were isolated, cultured, characterized, and transplanted to hindlimb ischemia mice model. DTI was performed on the hindlimb at postischemia time points. The edema regions of diffusion restriction (high signal in diffusion weighted imaging) were decreased in the ischemic muscle of EPCs treated mice after 14 days compared with the controls. These results from DTI show the lower apparent diffusion coefficient and eigenvalues (λ1, λ2, and λ3) and the higher fractional anisotropy and fiber counts of ischemic muscle on 7 and 14 days after EPCs treatment compared to the controls. There was a significant correlation between fiber counts calculated by DTI and survival fibers evaluated by histological section (r = 0.873, P < 0.01). Our study demonstrated that the time frame for muscle fiber regeneration after EPCs transplantation was significantly shortened in vivo. DTI could be a useful tool for noninvasive evaluation of muscle tissue damage and repair in animal models and patient with ischemic diseases.

Restoration of erectile function with intracavernous injections of endothelial progenitor cells after bilateral cavernous nerve injury inrats.

Endothelial progenitor cells (EPCs) are bone marrow-derived endothelial cells capable of circulating, proliferating, and differentiating into mature endothelial cells. Circulating EPCs can be directly recruited to some extent at sites of injury, and their administration could accelerate repair or endothelialization of the damaged tissue. We investigated the effects of intracavernous injections of EPCs into the corpora cavernosa of rats with erectile dysfunction (ED) caused by bilateral cavernous nerve (CN) injury. Overall, 24 male Sprague-Dawley rats were randomized into three groups: sham surgery, vehicle-only, or EPC treatment. Rats in the EPC treatment and vehicle-only groups were subjected to bilateral CN injury before injection of EPCs or vehicle, respectively, into the corpora cavernosa. Four weeks after surgery, erectile function was assessed by measuring maximum intracavernosal pressure (ICP), change in ICP, area under the ICP curve, and ratio of change in ICP and mean arterial pressure (MAP; ΔICP/MAP). Penile tissue was histomorphometrically analyzed for the expression of neural nitric oxide synthase (nNOS), neurofilament-1 (NF-1), von Willebrand factor (vWF), endothelial NOS (eNOS), and smooth muscle cell content. Maximum ICP and all other functional parameters of erectile function were significantly reduced in the vehicle-only group vs. the sham and EPC treatment groups (all p<0.001). Smooth muscle cell content was decreased in the vehicle-only vs. the sham and EPC treatment groups (both p<0.01). Expressions of vWF and eNOS in the dorsal artery were significantly higher in the EPC treatment than the vehicle-only group (p<0.05). In conclusion, EPC treatment restored erectile function in a rat model of bilateral CN injury through recruitment of EPCs toward the dorsal artery and preservation of smooth muscle cells in the corpus cavernosum. These findings elucidate the therapeutic potential of EPCs for treating ED in humans.

AB035. Restoration of erectile function with intracavernous injection of endothelial or smooth muscle progenitor cells after bilateral cavernous nerve injury in rats.

Injury to the cavernous nerve (CN) is a significant concern for patients with prostate cancer after radical prostatectomy because erectile dysfunction is secondary to denervation of the penis. Loss of innervation causes severe and irreversible morphologic changes in the penis, including induction of endothelial and smooth muscle cells apoptosis, collagen induction or fibrosis in the corpus cavernosum. Currently, cell-based therapy offers a promising strategy for neuroprotection or tissue-protection after bilateral CN injury. Bone marrow-derived progenitor cells (BMPCs) such as endothelial progenitor cells (EPCs) and smooth muscle progenitor cells (SPCs) can be derived from peripheral blood and be differentiated more easily into vascular cells during arterial remodeling than stem cells. In addition, administration of EPCs can increase vascular repair in animal models and has demonstrated some therapeutic benefits for cardiovascular conditions in clinical trials. A tight cooperation between endothelial cells and smooth muscle cells is important to regulate vessel maturation and stability. Therefore, we investigate the therapeutic effects of intracavernous injections of EPCs and SPCs into the corpora cavernosa of rats with erectile dysfunction (ED) caused by bilateral CN injury. The results show maximum ICP and all other functional parameters of erectile function are significantly reduced in the vehicle-only group versus the sham, EPC treatment and SPC treatment groups (all P<0.001). Smooth muscle cell content decreases in the vehicle-only versus the sham and EPCs treatment and SPCs treatment groups (both p<0.01). Expressions of von Willebrand factor (vWF) and endothelial nitric oxide synthase (eNOS) in the dorsal artery are significantly higher in the EPC treatment than the vehicle-only group (P<0.05). In conclusion, EPCs and SPCs treatment restores erectile function in a rat model of bilateral CN injury through recruitment of EPCs toward the dorsal artery and preservation of smooth muscle cells in the corpus cavernosum, respectively. These findings elucidate the therapeutic potential of EPCs and SPCs for treating ED in humans.

Diverse contribution of bone marrow-derived late-outgrowth endothelial progenitor cells to vascular repair under pulmonary arterial hypertension and arterial neointimal formation

AimsIt is still controversial whether bone marrow (BM)-derived endothelial progenitor cells (EPCs) can contribute to vascular repair and prevent the progression of vascular diseases. We aimed to characterize BM-derived EPC subpopulations and to evaluate their therapeutic efficacies to repair injured vascular endothelium of systemic and pulmonary arteries. Methods and resultsBM mononuclear cells of Fisher-344 rats were cultured under endothelial cell-conditions. Early EPCs appeared on days 3–6. Late-outgrowth and very late-outgrowth EPCs (LOCs and VLOCs) were defined as cells forming cobblestone colonies on days 9–14 and 17–21, respectively. Among EPC subpopulations, LOCs showed the highest angiogenic capability with enhanced proliferation potential and secretion of proangiogenic proteins. To investigate the therapeutic effects of these EPCs, Fisher-344 rats underwent wire-mediated endovascular injury in femoral artery (FA) and were concurrently injected intraperitoneally with 60mg/kg monocrotaline (MCT). Injured rats were then treated with six injections of one of three EPCs (1×106 per time). After 4weeks, transplanted LOCs, but not early EPCs or VLOCs, significantly attenuated neointimal lesion formation in injured FAs. Some of CD31+ LOCs directly replaced the injured FA endothelium (replacement ratio: 11.7±7.0%). In contrast, any EPC treatment could neither replace MCT-injured endothelium of pulmonary arterioles nor prevent the progression of pulmonary arterial hypertension (PAH). LOCs modified protectively the expression profile of angiogenic and inflammatory genes in injured FAs, but not in MCT-injured lungs. ConclusionBM-derived LOCs can contribute to vascular repair of injured systemic artery; however, even they cannot rescue injured pulmonary vasculature under MCT-induced PAH.

CCR5 facilitates endothelial progenitor cell recruitment and promotes the stabilization of atherosclerotic plaques in ApoE-/- mice.

IntroductionUnstable atherosclerotic plaques are prone to rupture, which leads to atherothrombosis. Endothelial progenitor cells (EPCs) are bone marrow-derived precursor cells that may repair vascular injury in atherosclerosis. Chemokine (C-C motif) receptor 5 (CCR5) promotes mobilization of EPCs. In this study, we investigated the therapeutic potential of CCR5-overexpressing EPCs on plaque stabilization in an apolipoprotein E (ApoE)−/− mouse model.MethodsThe expression of CCR5 and its cognate ligand chemokine (C-C motif) ligand 5 (CCL5) was examined in atherosclerotic aortas of humans and mice by immunohistochemistry. Splenectomized ApoE−/− C57BL/6 J mice fed a high-fat diet for 24 weeks were intravenously injected with EPCs transfected with CCR5 overexpression lentivirus. The recruitment of EPCs over the atherosclerotic plaques was evaluated by immunofluorescence. The content of lipid, smooth muscle cells, monocytes/macrophages, and endothelial cells in atherosclerotic plaques was assayed by specific immunostaining. The serum levels of atherosclerosis-related inflammatory factors in ApoE−/− mice were measured by mouse atherosclerosis antibody array I.ResultsCCR5 and CCL5 are highly expressed in atherosclerotic plaques in both humans and mice. The ApoE−/− mice with CCR5-overexpressing EPC treatment demonstrated a more stable plaque formation with enhanced recruitment of EPC, reduced lipid, and macrophage content in the atherosclerotic plaques. CCR5-overexpressing EPC treatment also increased the content of endothelial cells and nitric oxide production in the plaques. In addition, the serum levels of interleukin-3 (IL-3), IL-5, IL-6, IL-13, CD40, and tumor necrosis factor-alpha and the plaque contents of IL-6 and matrix metalloproteinase-9 were reduced in mice with CCR5-overexpressing EPC treatment.ConclusionsThese findings suggest that CCR5 is a novel therapeutic target in EPC treatment for stabilization of atherosclerotic plaques.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-015-0026-0) contains supplementary material, which is available to authorized users.