Enhanced Arteriogenesis and Wound Repair in Dystrophin-Deficient <i>mdx</i> Mice

Background— The absence of functional dystrophin in Duchenne muscular dystrophy (DMD) patients and in mdx mice results in progressive muscle degeneration associated with necrosis, fibrosis, and inflammation. Because vascular supply plays a key role in tissue repair, we examined whether new blood vessel development was altered in mdx mice. Methods and Results— In a model of hindlimb ischemia on femoral artery dissection, hindlimb perfusion, measured by laser Doppler imaging, was higher in mdx mice (0.67±0.26) than in wild-type (WT) mice (0.33±0.18, P <0.03). In keeping with these data, a significant increase in arteriole length density was found in mdx mice (13.6±8.4 mm/mm 3 ) compared with WT mice (7.8±4.6 mm/mm 3 , P <0.03). Conversely, no difference was observed in capillary density between mice of the 2 genotypes. The enhanced regenerative response was not limited to ischemic skeletal muscle, because in a wound-healing assay, mdx mice showed an accelerated wound closure rate compared with WT mice. Moreover, a vascularization assay in Matrigel plugs containing basic fibroblast growth factor injected subcutaneously revealed an increased length density of arterioles in mdx (46.9±14.7 mm/mm 3 ) versus WT mice (19.5±5.8 mm/mm 3 , P <0.001). Finally, serum derived from mdx mice sustained formation of endothelium-derived tubular structures in vitro more efficiently than WT serum. Conclusions— These results demonstrate that arteriogenesis is enhanced in mdx mice both after ischemia and skin wounding and in response to growth factors

Characterization of the Pall Celeris system as a point-of-care device for therapeutic angiogenesis

Abstract Background aims The Pall Celeris system is a filtration-based point-of-care device designed to obtain a high concentrate of peripheral blood total nucleated cells (PB-TNCs). We have characterized the Pall Celeris–derived TNCs for their in vitro and in vivo angiogenic potency. Methods PB-TNCs isolated from healthy donors were characterized through the use of flow cytometry and functional assays, aiming to assess migratory capacity, ability to form capillary-like structures, endothelial trans-differentiation and paracrine factor secretion. In a hind limb ischemia mouse model, we evaluated perfusion immediately and 7 days after surgery, along with capillary, arteriole and regenerative fiber density and local bio-distribution. Results Human PB-TNCs isolated by use of the Pall Celeris filtration system were shown to secrete a panel of angiogenic factors and migrate in response to vascular endothelial growth factor and stromal-derived factor-1 stimuli. Moreover, after injection in a mouse model of hind limb ischemia, PB-TNCs induced neovascularization by increasing capillary, arteriole and regenerative fiber numbers, with human cells detected in murine tissue up to 7 days after ischemia. Conclusions The Pall Celeris system may represent a novel, effective and reliable point-of-care device to obtain a PB-derived cell product with adequate potency for therapeutic angiogenesis

Cyclophilin A modulates bone marrow-derived CD117+ cells and enhances ischemia-induced angiogenesis via the SDF-1/CXCR4 axis

Abstract Background Critical limb ischemia (CLI) is a major health problem with no adequate treatment. Since CLI is characterized by insufficient tissue vascularization, efforts have focused on the discovery of novel angiogenic factors. Cyclophilin A (CyPA) is an immunophilin that has been shown to promote angiogenesis in vitro and to enhance bone marrow (BM) cell mobilization in vivo . However, its potential as an angiogenic factor in CLI is still unknown. Thus, this study aimed to evaluate whether CyPA might induce neo-angiogenesis in ischemic tissues. Methods and results Wild-type C57Bl/6j mice underwent acute hind-limb ischemia (HLI) and received a single intramuscular administration of recombinant CyPA or saline. Limb perfusion, capillary density and arteriole number in adductor muscles were significantly increased after CyPA treatment. Interestingly, BM-derived CD117 + cell recruitment was significantly higher in ischemic adductor tissue of mice treated with CyPA versus saline. Therefore, the effect of CyPA on isolated BM-derived CD117 + cells in vitro was evaluated. Low concentrations of CyPA stimulated CD117 + cell proliferation while high concentrations promoted cell death. Moreover, CyPA enhanced CD117 + cell adhesion and migration in a dose-dependent manner. Mechanistic studies revealed that CyPA up-regulated CXCR4 in CD117 + cells and in adductor muscles after ischemia. Additionally, SDF-1/CXCR4 axis inhibition by the CXCR4 antagonist AMD3100 decreased CyPA-mediated CD117 + cell recruitment in the ischemic limb. Conclusion CyPA induces neo-angiogenesis by recruiting BM-derived CD117 + cell into ischemic tissues, at least in part, through SDF-1/CXCR4 axis

Implantable Therapeutic Reservoir Systems for Diverse Clinical Applications in Large Animal Models

Regenerative medicine approaches, specifically stem cell technologies, have demonstrated significant potential to treat a diverse array of pathologies. However, such approaches have resulted in a modest clinical benefit, which may be attributed to poor cell retention/survival at the disease site. A delivery system that facilitates regional and repeated delivery to target tissues can provide enhanced clinical efficacy of cell therapies when localized delivery of high doses of cells is required. In this study, a new regenerative reservoir platform (Regenervoir) is described for use in large animal models, with relevance to cardiac, abdominal, and soft tissue pathologies. Regenervoir incorporates multiple novel design features essential for clinical translation, with a focus on scalability, mechanism of delivery, fixation to target tissue, and filling/refilling with a therapeutic cargo, and is demonstrated in an array of clinical applications that are easily translated to human studies. Regenervoir consists of a porous reservoir fabricated from a single material, a flexible thermoplastic polymer, capable of delivering cargo via fill lines to target tissues. A radiopaque shear thinning hydrogel can be delivered to the therapy reservoir and multiple fixation methods (laparoscopic tacks and cyanoacrylate bioadhesive) can be used to secure Regenervoir to target tissues through a minimally invasive approach.In this study, a new regenerative reservoir platform (Regenervoir) is described for use in large animal models that are easily translated to human studies, with relevance to cardiac, abdominal, and soft tissue pathologies. Regenervoir incorporates multiple novel design features essential for clinical translation, with a focus on scalability, mechanism of delivery, fixation, and filling/refilling with a therapeutic cargo.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/155890/1/adhm202000305.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/155890/2/adhm202000305_am.pd

High-Mobility Group Box 1 Protein in Human and Murine Skin: Involvement in Wound Healing

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Enhancement of lysine acetylation accelerates wound repair

In physiopathological conditions, such as diabetes, wound healing is significantly compromised and chronic complications, including ulcers, may occur. In a mouse model of skin repair, we recently reported that wound treatment with Sirtuin activators and class I HDAC inhibitors induced keratinocyte proliferation and enhanced healing via a nitric oxide (NO) dependent mechanism. We observed an increase in total protein acetylation in the wound area, as determined by acetylation of α-tubulin and histone H3 Lysine 9. We reasoned that this process activated cell function as well as regulated gene expression to foster tissue repair. We report here that the direct activation of P300/CBP-associated factor (PCAF) by the histone acetylase activator pentadecylidenemalonate 1b (SPV-106) induced Lysine acetylation in the wound area. This intervention was sufficient to enhance repair process by a NO-independent mechanism. Hence, an impairment of PCAF and/or other GCN5 family acetylases may delay skin repair in physiopathological conditions

Adenovirus-mediated human tissue kallikrein gene delivery induces angiogenesis in normoperfused skeletal muscle

We investigated whether local delivery of the tissue kallikrein gene induces angiogenesis in normoperfused mouse hindlimb muscles. Intramuscular injection of adenovirus containing the human tissue kallikrein gene under the control of a cytomegalovirus enhancer/promoter sequence resulted in local production and release of recombinant human tissue kallikrein, whereas transgene expression was absent in muscles of the contralateral hindlimb. Angiogenesis in infected muscles was documented by histological evidence of increased capillary density. In contrast, no angiogenic effect was seen either in the ipsilateral gastrocnemius or contralateral hindlimb muscles. Neovascularization was associated with a transient increase in muscular blood flow as determined by laser Doppler flowmetry. We also investigated the mechanisms of kallikrein-induced angiogenesis. We found that the angiogenic response to kallikrein was abolished by chronic blockade of the kinin B1 or B2 receptor or by inhibition of nitric oxide synthase. In addition, inhibition of cyclooxygenase-2 by nimesulide significantly reduced kallikrein-induced effects. These results indicate that (1) human tissue kallikrein acts as an angiogenic factor in normoperfused skeletal muscle and (2) nitric oxide and prostacyclin are essential mediators of kallikrein-induced angiogenesis. Our findings provide new insights into the role of the tissue kallikrein-kinin system in vascular biology