Protease nexin-1 deficiency increases mouse hindlimb neovascularisation following ischemia and accelerates femoral artery perfusion.
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ABSTRACT:
SUBMITTER: Selbonne S
PROVIDER: S-EPMC8238971 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Peripheral ischemia is associated with higher degree of endothelial dysfunction and a worse prognosis after percutaneous coronary interventions (PCI). However, the role of peripheral ischemia on vascular remodeling in remote districts remains poorly understood. Here we show that the presence of hindlimb ischemia significantly enhances neointima formation and impairs endothelial recovery in balloon-injured carotid arteries. Endothelial-derived microRNAs are involved in the modulation of these processes. Indeed, endothelial miR-16 is remarkably upregulated after vascular injury in the presences of hindlimb ischemia and exerts a negative effect on endothelial repair through the inhibition of RhoGDI? and nitric oxide (NO) production. We showed that the repression of RhoGDI? by means of miR-16 induces RhoA, with consequent reduction of NO bioavailability. Thus, hindlimb ischemia affects negative carotid remodeling increasing neointima formation after injury, while systemic antagonizzation of miR-16 is able to prevent these negative effects.
| S-EPMC5768880 | biostudies-other
Project description:Critical limb ischemia is a devastating manifestation of peripheral arterial disease with no effective strategies for improving morbidity and mortality outcomes. We tested the hypothesis that cellular mitochondrial function is a key component of limb pathology and that improving mitochondrial function represents a novel paradigm for therapy. BALB/c mice were treated with a therapeutic mitochondrial-targeting peptide (MTP-131) and subjected to limb ischemia (HLI). Compared to vehicle control, MTP-131 rescued limb muscle capillary density and blood flow (64.7±11% of contralateral vs. 39.9±4%), and improved muscle regeneration. MTP-131 also increased electron transport system flux across all conditions at HLI day-7. In vitro, primary muscle cells exposed to experimental ischemia demonstrated markedly reduced (~75%) cellular respiration, which was rescued by MTP-131 during a recovery period. Compared to muscle cells, endothelial cell (HUVEC) respiration was inherently protected from ischemia (~30% reduction), but was also enhanced by MTP-131. These findings demonstrate an important link between ischemic tissue bioenergetics and limb blood flow and indicate that the mitochondria may be a pharmaceutical target for therapeutic intervention during critical limb ischemia.
| S-EPMC5002368 | biostudies-literature
Project description:ObjectiveThis study aimed to examine acellular extracellular matrix based hydrogels as potential therapies for treating peripheral artery disease (PAD). We tested the efficacy of using a tissue specific injectable hydrogel, derived from decellularized porcine skeletal muscle (SKM), compared to a new human umbilical cord derived matrix (hUC) hydrogel, which could have greater potential for tissue regeneration because of its young tissue source age.BackgroundThe prevalence of PAD is increasing and can lead to critical limb ischemia (CLI) with potential limb amputation. Currently there are no therapies for PAD that effectively treat all of the underlying pathologies, including reduced tissue perfusion and muscle atrophy.MethodsIn a rodent hindlimb ischemia model both hydrogels were injected 1-week post-surgery and perfusion was regularly monitored with laser speckle contrast analysis (LASCA) to 35 days post-injection. Histology and immunohistochemistry were used to assess neovascularization and muscle health. Whole transcriptome analysis was further conducted on SKM injected animals on 3 and 10 days post-injection.ResultsSignificant improvements in hindlimb tissue perfusion and perfusion kinetics were observed with both biomaterials. End point histology indicated this was a result of arteriogenesis, rather than angiogenesis, and that the materials were biocompatible. Skeletal muscle fiber morphology analysis indicated that the muscle treated with the tissue specific, SKM hydrogel more closely matched healthy tissue morphology. Short term histology also indicated arteriogenesis rather than angiogenesis, as well as improved recruitment of skeletal muscle progenitors. Whole transcriptome analysis indicated that the SKM hydrogel caused a shift in the inflammatory response, decreased cell death, and increased blood vessel and muscle development.ConclusionThese results show the efficacy of an injectable ECM hydrogel alone as a potential therapy for treating patients with PAD. Our results indicate that the SKM hydrogel improved functional outcomes through stimulation of arteriogenesis and muscle progenitor cell recruitment.
| S-EPMC4834896 | biostudies-literature
Project description:Effective treatments for chronic limb-threatening ischemia are lacking. (Pre)clinical studies on administration of bone marrow (BM) mononuclear cells (MNCs) and BM-derived mesenchymal stromal cells (MSCs) have shown variable results and no studies have directly compared administration of human BM MNCs and BM MSCs in in vivo models. We studied the effect of intramuscular administration of human BM-derived MNCs and MSCs on limb perfusion in the murine hindlimb ischemia (HLI) model. Human BM MNCs and MSCs were obtained from healthy consenting donors. Both cell types were cryopreserved before use. Twenty-four hours after induction of HLI, nude NMRI mice were randomized to receive intramuscular administration of human BM MNCs (n = 13), or BM MSCs (n = 14), or vehicle control (n = 19) in various doses. Limb perfusion was measured using laser Doppler imaging on day 0, 1, 4, 7, 10, and 14. Intramuscular injection of human BM MNCs did not improve limb perfusion as compared with vehicle over the 2 weeks after cell administration (P = 0.88, mean relative perfusion for vehicle 0.56 ± 0.04 and 0.53 ± 0.04 for BM MNCs at day 14). Administration of human BM MSCs significantly improved limb perfusion as compared with both BM MNCs and vehicle (P ≤ 0.001, mean relative perfusion at day 14 0.79 ± 0.06). Our data suggest that BM MNCs are less suitable than BM MSCs for cell-based therapy that aims to restore perfusion.
| S-EPMC9057881 | biostudies-literature
Project description:Surgical hindlimb ischemia (HLI) in mice has become a valuable preclinical model to study peripheral arterial disease. We previously identified that the different phenotypic outcomes after HLI across inbred mouse strains is related to a region on the short arm of mouse chromosome 7. The gene coding the interleukin-21 receptor (IL-21R) lies at the peak of association in this region.With quantitative real-time polymerase chain reaction, we found that a mouse strain with a greater ability to upregulate IL-21R after HLI had better perfusion recovery than a strain with no upregulation after HLI. Immunofluorescent staining of ischemic hindlimb tissue showed IL-21R expression on endothelial cells (ECs) from C57BL/6 mice. An EC-enriched fraction isolated from ischemic hindlimb muscle showed higher Il-21R levels than an EC-enriched fraction from nonischemic limbs. In vitro, human umbilical vein ECs showed elevated IL-21R expression after hypoxia and serum starvation. Under these conditions, IL-21 treatment increased cell viability, decreased cell apoptosis, and augmented tube formation. In vivo, either knockout Il21r or blocking IL-21 signaling by treating with IL-21R-Fc (fusion protein that blocks IL-21 binding to its receptor) in C57BL/6 mice resulted in less perfusion recovery after HLI. Both in vitro and in vivo modulation of the IL-21/IL-21R axis under hypoxic conditions resulted in increased signal transducer and activator of transcription 3 phosphorylation and a subsequent increase in the B-cell lymphoma leukemia-2/BCL-2-associated X protein ratio.Our data indicate that IL-21R upregulation and ligand activation in hypoxic ECs may help perfusion recovery by limiting/preventing apoptosis and favoring cell survival and angiogenesis through the signal transducer and activator of transcription 3 pathway.
| S-EPMC4865891 | biostudies-literature
Project description:Humans have fewer cardiovascular events and improved outcomes after cardiovascular events when living at low and moderate altitudes (<3000 m) above sea level. We have previously shown that low-altitude simulation using reductions in barometric pressure enhances vasodilation ex vivo in arterial segments and reduces systemic vascular resistance in vivo and can also improve left ventricular function after a myocardial infarction. We hypothesize that low-altitude simulation could also improve hindlimb ischemia, a model of peripheral artery disease in humans. We performed femoral artery ligation to generate hindlimb ischemia in 3-month-old C57BL6 mice. Control group mice (n = 10) recovered at 754 mmHg (control) for 14 days. Treatment group mice (n = 15) were placed in a low-altitude simulation chamber (at 714 mmHg) to recover from surgery for 3-hours daily for 14 days. Hindlimb perfusion imaging using a laser Doppler line scanner was performed for all mice prior to the surgery, and then on days 1, 3, 7, and 14 post-surgery. At 2 weeks, ischemic reserve was significantly higher in the treatment group mice (0.50 ± 0.13 vs. 0.20 ± 0.06; p = 0.01). Treatment mice had higher functional scores and were able to walk better at two weeks. There was approximately three times less HIF1α found via western blotting and a small but statistically significant improvement of lectin perfusion in calf tissue of treatment mice. We conclude that low-altitude simulation improves blood perfusion in murine hindlimb ischemia. This approach may have therapeutic implications for humans with peripheral artery disease.
| S-EPMC7839326 | biostudies-literature
Project description:During incomplete skeletal muscle recovery from ischemia, such as that occurs with critical limb ischemia, the temporal relationship between recovery of muscle capillary perfusion and contractile function is poorly defined. We examined this relationship in BALB/cJ mice (N = 24) following unilateral hindlimb ischemia (HLI), which pre-clinically mimics the myopathy observed in critical limb ischemia patients. Specifically, we examined this relationship in two phenotypically distinct muscles (i.e., "oxidative" soleus - Sol and "glycolytic" extensor digitorum longus - EDL) 14- or 56-days after HLI. Although overall limb blood flow (LDPI) reached its' recovery peak (48% of control) by HLI d14, the capillary networks in both the Sol and EDL (whole mount confocal imaging) were disrupted and competent muscle capillary perfusion (perfused lectin+μm2/muscle μm2) remained reduced. Interestingly, both Sol and EDL muscles recovered their distinct capillary structures and perfusion (Con Sol; 0.056 ± 0.02 lectin+μm2/muscle μm2, and Con EDL; 0.039 ± 0.005 lectin+μm2/muscle μm2) by HLI d56 (Sol; 0.062 ± 0.011 lectin+μm2/muscle μm2 and EDL; 0.0035 ± 0.005 lectin+μm2/muscle μm2), despite no further improvement in limb blood flow (LDPI). Both muscles suffered severe myopathy, indicated by loss of dystrophin positive immunostaining and the absence of stimulation induced isometric force production at HLI d14. Dystrophin immunofluorescence returned at HLI d56, although neither myofiber CSA (μm2) nor isometric force production (58 and 28% sustained deficits, Sol and EDL, respectively) recovered completely in either muscle. In summary, we reveal that the temporal relationship between the restoration of muscle capillary perfusion and functional ischemic skeletal muscle regeneration favors competent muscle capillary perfusion recovery in BALB/c mice in a phenotypically non-distinct manner.
| S-EPMC6611152 | biostudies-literature
Project description:BackgroundA better understanding of the blood supply of the femoral head is essential to guide therapeutic strategies for patients with femoral neck fractures. However, because of the limitations of conventional techniques, the precise distribution and characteristics of intraosseous arteries of the femoral head are not well displayed.Questions/purposesTo explore the characteristics and interconnections of the intraosseous vessel system between different areas of the femoral head and the possible blood supply compensatory mechanism after femoral neck fracture.MethodsThe three-dimensional (3-D) structures of the intraosseous blood supply in 30 uninjured normal human femoral heads were reconstructed using angiography methods and microCT scans. The data were imported in the AMIRA® and MIMICS® software programs to reconstruct and quantify the extra- and intraosseous arteries (diameter, length). In a separate experiment, we evaluated the residual blood supply of femoral heads in 27 patients with femoral neck fractures before surgery by analyzing digital subtraction angiography data; during the study period, this was performed on all patients in whom hip-preserving surgery was planned, rather than arthroplasty. The number of affected and unaffected subjects included in the three groups (superior, inferior, and anterior retinacular arteries) with different types of fractures (Garden Types I-IV) were recorded and analyzed (Fisher's exact test) to reflect the affected degrees of these three groups of retinacular arteries in patients after femoral neck fractures.ResultsThe main results of our cadaver study were: (1) the main blood supply sources of the femoral head were connected by three main network structures as a whole, and the epiphyseal arterial network is the most widely distributed and the primary network structure in the femoral head; (2) the main stems of the epiphyseal arteries which were located on the periphery of the intraosseous vascular system have fewer anastomoses than the network located in the central region; (3) compared with the round ligament artery and anterior retinacular artery, the inferior retinacular artery has a relatively large caliber. Digital subtraction angiography of the 27 patients with hip fractures indicated that the inferior retinacular arterial system had a high likelihood of being unaffected after femoral neck fracture (100% [14 of 14] in nondisplaced fractures and 60% [six of 10] in Garden Type III fractures).ConclusionsThe epiphyseal arterial network and inferior retinacular arterial system appear to be two important structures for maintaining the femoral head blood supply after femoral neck fracture. Increased efforts to protect these key structures during surgery, such as drilling and placing internal implants closer to the central region of the femoral head, might be helpful to reduce the effect of iatrogenic injury of the intraosseous vascular system.Clinical relevance3-D anatomic evidence of intraosseous arterial distribution of the femoral head and the high frequency with which the inferior retinacular arteries remained patent after femoral neck fracture lead us to consider the necessity of drilling and placing internal implants closer to the central region of the femoral head during surgery. Future controlled studies might evaluate this proposition.
| S-EPMC5498378 | biostudies-literature
Project description:The first three exons and the promoter of rat glia-derived nexin, also called protease nexin-1 (GDN/PN-1), have been identified through analysis of rat genomic clones. A 1.6 kilobase (kb) fragment containing 105 base pairs of the first exon and 5'-flanking sequences was sequenced. The 5'-flanking sequence and the first exon were found to be GC-rich, indicating that the 5' region of the rat GDN/PN-1 gene resides within a CpG island. A TATA box-like sequence, but no CAAT box, was found. The rat GDN/PN-1 promoter contains five SP1 consensus sites, four consensus sites for the MyoD1 transcription factor, and one binding site for the transcription factors NGFI-A, NGFI-C, Krox-20, and Wilms tumor factor. The presence of these consensus sequences is consistent with the known expression pattern of GDN/PN-1. Primer extension and RNase protection assays identified one transcriptional start site. The 1.6 kb promoter fragment cloned in a reporter plasmid was found to induce firefly luciferase expression in a cell-specific manner. A positive regulatory element is localized in the region -1545 to -389. In vitro CpG methylation blocked transcription from the GDN/PN-1 promoter in rat hepatoma cells but not in C6 rat glioma cells.
| S-EPMC6081634 | biostudies-literature
Project description:MicroRNAs are key regulators of gene expression in response to injury, but there is limited knowledge of their role in ischemia-induced angiogenesis, such as in peripheral arterial disease. Here, we used an unbiased strategy and took advantage of different phenotypic outcomes that follow surgically induced hindlimb ischemia between inbred mouse strains to identify key microRNAs involved in perfusion recovery from hindlimb ischemia.From comparative microRNA profiling between inbred mouse strains that display profound differences in their extent of perfusion recovery after hindlimb ischemia, we found that the mouse strain with higher levels of microRNA-93 (miR-93) in hindlimb muscle before ischemia and the greater ability to upregulate miR-93 in response to ischemia had better perfusion recovery. In vitro, overexpression of miR-93 attenuated hypoxia-induced apoptosis in both endothelial and skeletal muscle cells and enhanced proliferation in both cell types. In addition, miR-93 overexpression enhanced endothelial cell tube formation. In vivo, miR-93 overexpression enhanced capillary density and perfusion recovery from hindlimb ischemia, and antagomirs to miR-93 attenuated perfusion recovery. Both in vitro and in vivo modulation of miR-93 resulted in alterations in the expression of >1 cell cycle pathway gene in 2 different cell types.Our data indicate that miR-93 enhances perfusion recovery from hindlimb ischemia by modulation of multiple genes that coordinate the functional pathways of cell proliferation and apoptosis. Thus, miR-93 is a strong potential target for pharmacological modulation to promote angiogenesis in ischemic tissue.
| S-EPMC3856290 | biostudies-literature