ASC and SVF Cells Synergistically Induce Neovascularization in Ischemic Hindlimb Following Cotransplantation.
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ABSTRACT:
SUBMITTER: Zhang HZ
PROVIDER: S-EPMC8745515 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The angiopoietins (ANGPT) are ligands for the endothelial cell (EC) receptor tyrosine kinase, Tie2. Angpt-1 is a Tie2 agonist that promotes vascular maturation and stabilization, whereas Angpt-2 is a partial agonist/antagonist involved in the initiation of postnatal angiogenesis. Therefore, we hypothesized that overexpression of Angpt-2 would be more effective than Angpt-1 for enhancing the perfusion recovery in the ischemic hindlimb. Perfusion recovery was markedly impaired in Tie2-deficient animals at day 35 in a model of chronic hindlimb ischemia. Injections of Angpt-2 or VEGFA plasmid at 7 days post femoral artery resection enhanced recovery and improved arteriogenesis as assessed by angiographic scores, whereas Angpt-1 or null plasmid had no effect. In addition, Angpt-2 together with VEGF resulted in greater improvement in perfusion and collateral vessel formation than VEGF alone. Similarly, conditional overexpression of Angpt-2 in mice improved ischemic limb blood flow recovery, while Angpt-1 overexpression was ineffective. These data from Tie2 heterozygote deficient mice demonstrate, for the first time, the importance of the Tie2 pathway in spontaneous neovascularization in response to chronic hindlimb ischemia. Moreover, they show that overexpression of the partial agonist, Angpt-2, but not Angpt-1, enhanced ischemic hind limb perfusion recovery and collateralization, suggesting that a coordinated sequence antagonist and agonist activity is required for effective therapeutic revascularization.
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Project description:BackgroundHuman tissue kallikrein (hTK) plays an essential role in the physiological and pathological mechanisms of blood vessels. This study aimed to determine whether angiogenesis induced by endothelial progenitor cells (EPCs) transduced with the adenovirus-mediated hTK gene could improve blood flow in rat hindlimb ischemia in vivo and to establish a promising mechanism in vitro.MethodsEPCs transduced with adenovirus encoding hTK-162 (i.e., Ad/hTK-transduced EPCs or Ad/GFP-transduced EPCs) were administered to Wister rats with hindlimb ischemia through therapeutic neovascularization. Muscular capillary density (MCD), blood flow (BF), and the number of myofibers were measured at days 7, 14, and 21 after treatment. Expressions of integrin ?v?3 and endothelial nitric oxide synthase (eNOS) were detected on the surface of EPCs.ResultsMCD, BF, and the number of myofibers in rats with Ad/hTK-transduced EPCs remarkably increased at day 21 after treatment compared with rats with Ad/GFP-transduced EPCs or the control group (P<0.01). Expressions of integrin ?v?3 and eNOS protein on the surface of EPCs also increased in rats with Ad/hTK-transduced EPCs. The levels of integrin ?v?3 expression were reduced by PI3K and eNOS blockade, and the inhibitor of integrin ?v?3 abrogated the migration and adhesion of hTK-transduced EPCs (P<0.05).ConclusionhTK gene delivery in vivo improves the natural angiogenic response to ischemia. The ability of hTK gene-transduced EPCs can be enhanced in vitro, in which integrin ?v?3 plays a role in the process.
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Project description:Collateral vessel formation can functionally compensate for obstructive vascular lesions in patients with atherosclerosis. Neovascularization processes are triggered by fluid shear stress, hypoxia, growth factors, chemokines, proteases, and inflammation, as well as reactive oxygen species, in response to ischemia. Polymerase ?-interacting protein 2 (Poldip2) is a multifunctional protein that regulates focal adhesion turnover and vascular smooth muscle cell migration and modifies extracellular matrix composition. We, therefore, tested the hypothesis that loss of Poldip2 impairs collateral formation.The mouse hindlimb ischemia model has been used to understand mechanisms involved in postnatal blood vessel formation. Poldip2(+/-) mice were subjected to femoral artery excision, and functional and morphological analysis of blood vessel formation was performed after injury. Heterozygous deletion of Poldip2 decreased the blood flow recovery and spontaneous running activity at 21 days after injury. H2O2 production, as well as the activity of matrix metalloproteinases-2 and -9, was reduced in these animals compared with Poldip2(+/+) mice. Infiltration of macrophages in the peri-injury muscle was also decreased; however, macrophage phenotype was similar between genotypes. In addition, the formation of capillaries and arterioles was impaired, as was angiogenesis, in agreement with a decrease in proliferation observed in endothelial cells treated with small interfering RNA against Poldip2. Finally, regression of newly formed vessels and apoptosis was more pronounced in Poldip2(+/-) mice.Together, these results suggest that Poldip2 promotes ischemia-induced collateral vessel formation via multiple mechanisms that likely involve reactive oxygen species-dependent activation of matrix metalloproteinase activity, as well as enhanced vascular cell growth and survival.
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Project description:As one of the most severe manifestations of diabetes, vascular complications are the main causes of diabetes-related morbidity and mortality. Hyperglycemia induces systemic abnormalities, including impaired angiogenesis, causing diabetic patients to be highly susceptible in suffering hindlimb ischemia (HLI). Despite its severe prognosis, there is currently no effective treatment for diabetic HLI. Skeletal muscle cells secrete multiple angiogenic factors, hence, recently are reported to be critical for angiogenesis; however, hyperglycemia disrupted the paracrine function in skeletal muscle cells, leading to the impaired angiogenesis potential observed in diabetic patients. The present study showed that tyrosol, a phenylethanoid compound, suppresses accumulation of intracellular reactive oxygen species (ROS) caused by hyperglycemia, most plausibly by promoting heme oxygenase-1 (HO-1) expression in skeletal muscle cells. Consequently, tyrosol exerts cytoprotective function against hyperglycemia-induced oxidative stress in skeletal muscle cells, increases their proliferation vigorously, and simultaneously suppresses apoptosis. Furthermore, tyrosol grossly increases the secretion of vascular endothelial growth factor-A (VEGF-A) and platelet-derived growth factor-BB (PDGF-BB) from skeletal muscle cells. This leads to enhanced proliferation and migration capabilities of vascular endothelial and smooth muscle cells, two types of cells that are responsible in forming blood vessels, through cell-cell communication. Finally, in vivo experiment using the diabetic HLI mouse model showed that tyrosol injection into the gastrocnemius muscle of the ischemic hindlimb significantly enhances the formation of functional blood vessels and subsequently leads to significant recovery of blood perfusion. Overall, our findings highlight the potential of the pharmacological application of tyrosol as a small molecule drug for therapeutic angiogenesis in diabetic HLI patients.
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Project description:Recently, it was demonstrated that arteriogenesis is enhanced in mice deficient in regulatory T cells (CD4(+) CD25(+) FoxP3(+) T cell), which can suppress effector T cell responses. The present study investigates the effects of these regulatory T cells on arteriogenesis in more detail by either specific expanding or depleting regulatory T cells. Hind limb ischemia was induced by electro-coagulation of the femoral artery in mice. Regulatory T cells were either expanded by injecting mice with a complex of interleukin (IL)-2 with the IL-2 monoclonal antibody JES6-1, or depleted by anti-CD25 antibody or diphtheria toxin injections in DEREG mice (depletion of regulatory T cells). Blood flow restoration was monitored using laser Doppler perfusion imaging. Collateral arteries were visualized by immunohistochemistry. Regulatory T cell expansion led to a moderate though significant suppression of blood flow restoration after ischemia induction. Surprisingly, depletion of regulatory T cells resulted in minor increase on blood flow recovery. However, collateral and capillary densities in the post-ischemic skeletal muscle were significantly increased in DEREG mice depleted for regulatory T cells. The presence of regulatory T cells after ischemia induction when analysed in non-depleted DEREG mice could be demonstrated by green fluorescent protein staining only in lymph nodes in the ischemic area, and not in the ischemic muscle tissue. The current study demonstrates that, even under conditions of major changes in regulatory T cell content, the contribution of regulatory T cells to the regulation of the arteriogenic response is only moderate.
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Project description:Dysfunction or loss of blood vessel causes several ischemic diseases. Although endothelial progenitor cells (EPCs) are a promising source for cell-based therapy, ischemia-induced pathophysiological condition limits the recovery rate by causing drastic cell death. To overcome this issue, we attempted to develop a cell-targeted peptide delivery and priming system to enhance EPC-based neovascularization using an engineered M13 bacteriophage harboring nanofibrous tubes displaying ~2700 multiple functional motifs. The M13 nanofiber was modified by displaying RGD, which is an integrin-docking peptide, on the minor coat protein, and by mutilayering SDKP motifs, which are the key active sites for thymosin ?4, on the major coat protein. The engineered M13 nanofiber dramatically enhanced ischemic neovascularization by activating intracellular and extracellular processes such as proliferation, migration, and tube formation in the EPCs. Furthermore, transplantation of the primed EPCs with the M13 nanofiber harboring RGD and SDKP facilitated functional recovery and neovascularization in a murine hindlimb ischemia model. Overall, this study demonstrates the effectiveness of the M13 nanofiber-based novel peptide delivery and priming strategy in promoting EPC bioactivity and neovessel regeneration. To our knowledge, this is first report on M13 nanofibers harboring dual functional motifs, the use of which might be a novel strategy for stem and progenitor cell therapy against cardiovascular ischemic diseases.
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Project description:Unilateral femoral artery occlusion (right side) and a sham operation on the contralateral (left) side was performed in C57BL/6J mice under anesthesia by double ligation of the superficial femoral artery proximal to the deep femoral artery and distal femoral artery. Animal numbers are stated with the different experimental results. Total RNA was isolated from the distal adductor muscles by phenol-chloroform isolation (TRIzol, Invitrogen, Carlsbad, CA) at baseline and at 5 time points after femoral artery ligation (6h, 24h, 3 days, 7 days, 14 days) from 5 mice per time point. RNA was pooled in equal amounts, and microarray analysis for all identified murine miRNAs (miRBase 9.0) was performed by a service provider (LC Sciences, Houston, TX). 11 condition experiment. Biological replicates: 5 per condition, 5 pooled RNA replicates per array.
2015-06-12 | E-GEOD-63212 | biostudies-arrayexpress
Project description:AimsStudies in global androgen receptor knockout (G-ARKO) and orchidectomised mice suggest that androgen accelerates reperfusion of the ischaemic hindlimb by stimulating angiogenesis. This investigation used novel, vascular cell-specific ARKO mice to address the hypothesis that the impaired hindlimb reperfusion in G-ARKO mice was due to loss of AR from cells in the vascular wall.Methods and resultsMice with selective deletion of AR (ARKO) from vascular smooth muscle cells (SM-ARKO), endothelial cells (VE-ARKO), or both (SM/VE-ARKO) were compared with wild type (WT) controls. Hindlimb ischaemia was induced in these mice by ligation and removal of the femoral artery. Post-operative reperfusion was reduced in SM-ARKO and SM/VE-ARKO mice. Immunohistochemistry indicated that this was accompanied by a reduced density of smooth muscle actin-positive vessels but no change in the density of isolectin B4-positive vessels in the gastrocnemius muscle. Deletion of AR from the endothelium (VE-ARKO) did not alter post-operative reperfusion or vessel density. In an ex vivo (aortic ring culture) model of angiogenesis, AR was not detected in vascular outgrowths and angiogenesis was not altered by vascular ARKO or by exposure to dihydrotestosterone (DHT 10(-10)-10(-7)M; 6 days).ConclusionThese results suggest that loss of AR from vascular smooth muscle, but not from the endothelium, contributes to impaired reperfusion in the ischaemic hindlimb of G-ARKO. Impaired reperfusion was associated with reduced collateral formation rather than reduced angiogenesis.
| S-EPMC4861284 | biostudies-literature
Project description:Unilateral femoral artery occlusion (right side) and a sham operation on the contralateral (left) side was performed in C57BL/6J mice under anesthesia by double ligation of the superficial femoral artery proximal to the deep femoral artery and distal femoral artery. Animal numbers are stated with the different experimental results. Total RNA was isolated from the distal adductor muscles by phenol-chloroform isolation (TRIzol, Invitrogen, Carlsbad, CA) at baseline and at 5 time points after femoral artery ligation (6h, 24h, 3 days, 7 days, 14 days) from 5 mice per time point. RNA was pooled in equal amounts, and microarray analysis for all identified murine miRNAs (miRBase 9.0) was performed by a service provider (LC Sciences, Houston, TX).
2015-06-12 | GSE63212 | GEO
Project description:We examined the effect of delivery modality on the survival, localization, and functional effects of exogenously administered embryonic stem cells (ESCs) or endothelial cells derived from them (ESC-ECs) in the ischemic hindlimb.Murine ESCs or ESC-ECs were stably transduced with a construct for bioluminescence imaging (BLI) and fluorescent detection. In a syngeneic murine model of limb ischemia, ESCs or ESC-ECs were delivered by intramuscular (IM), intrafemoral artery (IA), or intrafemoral vein injections (n=5 in each group). For 2 weeks, cell survival and localization were tracked by BLI and confirmed by immunohistochemistry, and functional improvement was assessed by laser Doppler perfusion. BLI showed that ESCs localized to the ischemic limb after IM or IA, but not after intrafemoral vein administration. Regardless of the route of administration, ESCs were detected outside the hindlimb circulation in the spleen or lungs. ESCs did not improve limb perfusion and generated teratomas. In contrast, ESC-ECs delivered by all 3 modalities localized to the ischemic limb, as assessed by BLI. Most surprisingly, ESC-EC injected intrafemoral vein eventually localized to the ischemic limb after initially lodging in the pulmonary circulation. Immunohistochemical studies confirmed the engraftment of ESC-ECs into the limb vasculature after 2 weeks. Notably, ESC-ECs were not detected in the spleen or lungs after 2 weeks, regardless of route of administration. Furthermore, ESC-ECs significantly improved limb perfusion and neovascularization compared with the parental ESCs or the vehicle control group.In contrast to parental ESCs, ESC-ECs preferentially localized in the ischemic hindlimb by IA, IM, and intrafemoral vein delivery. ESC-ECs engrafted into the ischemic microvasculature, enhanced neovascularization, and improved limb perfusion.
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