Role of Oxygen and Its Radicals in Peripheral Nerve Regeneration: From Hypoxia to Physoxia to Hyperoxia.

Oxygen is compulsory for mitochondrial function and energy supply, but it has numerous more nuanced roles. The different roles of oxygen in peripheral nerve regeneration range from energy supply, inflammation, phagocytosis, and oxidative cell destruction in the context of reperfusion injury to crucial redox signaling cascades that are necessary for effective axonal outgrowth. A fine balance between reactive oxygen species production and antioxidant activity draws the line between physiological and pathological nerve regeneration. There is compelling evidence that redox signaling mediated by the Nox family of nicotinamide adenine dinucleotide phosphate (NADPH) oxidases plays an important role in peripheral nerve regeneration. Further research is needed to better characterize the role of Nox in physiological and pathological circumstances, but the available data suggest that the modulation of Nox activity fosters great therapeutic potential. One of the promising approaches to enhance nerve regeneration by modulating the redox environment is hyperbaric oxygen therapy. In this review, we highlight the influence of various oxygenation states, i.e., hypoxia, physoxia, and hyperoxia, on peripheral nerve repair and regeneration. We summarize the currently available data and knowledge on the effectiveness of using hyperbaric oxygen therapy to treat nerve injuries and discuss future directions.

Therapeutic Potential and Challenges of Mesenchymal Stem Cell-Derived Exosomes for Peripheral Nerve Regeneration: A Systematic Review.

Gap injuries to the peripheral nervous system result in pain and loss of function, without any particularly effective therapeutic options. Within this context, mesenchymal stem cell (MSC)-derived exosomes have emerged as a potential therapeutic option. Thus, the focus of this study was to review currently available data on MSC-derived exosome-mounted scaffolds in peripheral nerve regeneration in order to identify the most promising scaffolds and exosome sources currently in the field of peripheral nerve regeneration. We conducted a systematic review following PRISMA 2020 guidelines. Exosome origins varied (adipose-derived MSCs, bone marrow MSCs, gingival MSC, induced pluripotent stem cells and a purified exosome product) similarly to the materials (Matrigel, alginate and silicone, acellular nerve graft [ANG], chitosan, chitin, hydrogel and fibrin glue). The compound muscle action potential (CMAP), sciatic functional index (SFI), gastrocnemius wet weight and histological analyses were used as main outcome measures. Overall, exosome-mounted scaffolds showed better regeneration than scaffolds alone. Functionally, both exosome-enriched chitin and ANG showed a significant improvement over time in the sciatica functional index, CMAP and wet weight. The best histological outcomes were found in the exosome-enriched ANG scaffold with a high increase in the axonal diameter and muscle cross-section area. Further studies are needed to confirm the efficacy of exosome-mounted scaffolds in peripheral nerve regeneration.

[Management of patients with complex ear deformities]. / Prise en charge des patients présentant des déformations complexes du pavillon de l'oreille.

Complex ear reconstruction requires specialized multidisciplinary care. Most patients present with microtia, often associated with hearing disorders. The management of these disorders is a priority, and reconstruction of the external ear remains optional. Nowadays, auricular reconstruction is based on the subcutaneous implantation of either autologous cartilage or an allogeneic implant. Autologous reconstruction requires highly specialized surgical expertise and involves harvesting rib cartilage but carries a lower risk of exposure compared to allogeneic implants. Both techniques yield good results with a high success rate and have a positive impact on the social functioning and daily life of patients.

La reconstruction complexe du pavillon auriculaire nécessite une prise en charge multidisciplinaire spécialisée. La majorité des patients nécessitant ce geste présentent une microtie, souvent associée à des troubles de l'audition. La prise en charge de ceux-ci est prioritaire et la reconstruction du pavillon reste facultative. Aujourd'hui, la reconstruction du pavillon se base sur l'implantation sous-cutanée d'une maquette de cartilage autologue ou d'un implant allogène. La reconstruction autologue demande une expertise chirurgicale hautement spécialisée et nécessite un prélèvement de cartilage costal mais présente un risque d'exposition inférieur par rapport à l'implant allogène. Les deux techniques permettent d'atteindre de bons résultats avec un taux de réussite élevé et un effet positif sur le fonctionnement social et le quotidien des patients.

Adipose-derived stem cell spheroids are superior to single-cell suspensions to improve fat autograft long-term survival.

Autologous fat transplantation is a widely used procedure for surgical reconstruction of tissues. The resorption rate of this transplantation remains high and unpredictable, reinforcing the need of adjuvant treatments that increase the long-term stability of grafts. Adipose-derived stem cells (ASC) introduced as single cells in fat has been shown clinically to reduce the resorption of fat grafts. On the other hand, the formulation of ASC into cell spheroids results in the enhancement of their regenerative potential. In this study, we developed a novel method to produce highly homogeneous ASC spheroids and characterized their features and efficacy on fat transplantation. Spheroids conserved ASC markers and multipotency. A regenerative gene expression profile was maintained, and genes linked to autophagy were upregulated whereas proliferation was decreased. Their secreted proteome was enriched in comparison with single-cell ASC suspension. Addition of spheroids to fat graft in an animal model of transplantation resulted in a better graft long-term stability when compared to single ASC suspension. In conclusion, we provide a novel method to manufacture homogenous ASC spheroids. These ASC spheroids are superior to ASC in single-cell suspension to improve the stability of fat transplants, reinforcing their potential in reconstructive surgery.

Reactive Oxygen Species and NOX Enzymes Are Emerging as Key Players in Cutaneous Wound Repair.

Our understanding of the role of oxygen in cell physiology has evolved from its long-recognized importance as an essential factor in oxidative metabolism to its recognition as an important player in cell signaling. With regard to the latter, oxygen is needed for the generation of reactive oxygen species (ROS), which regulate a number of different cellular functions including differentiation, proliferation, apoptosis, migration, and contraction. Data specifically concerning the role of ROS-dependent signaling in cutaneous wound repair are very limited, especially regarding wound contraction. In this review we provide an overview of the current literature on the role of molecular and reactive oxygen in the physiology of wound repair as well as in the pathophysiology and therapy of chronic wounds, especially under ischemic and hyperglycemic conditions.

Hyperbaric oxygen therapy promotes wound repair in ischemic and hyperglycemic conditions, increasing tissue perfusion and collagen deposition.

The treatment of chronic wounds remains inconsistent and empirical. Hyperbaric oxygen therapy (HBOT) is a promising method to improve wound repair but there is still a lack of understanding of its mechanisms of action and its indications are not yet clearly defined. We studied the effects of HBOT in four different wound conditions by inflicting bilateral wounds on the dorsal aspect of the feet of nonischemic or ischemic limbs in normoglycemic or hyperglycemic rats. To create an ischemic condition, arterial resection was performed unilaterally. Forty-four animals received HBOT five times a week until complete wound closure. Wound repair was compared with 44 rats receiving standard dressing only. HBOT increased blood flow and accelerated wound closure in ischemic and hyperglycemic wounds, most significantly when the two conditions were combined. Wound contraction and reepithelialization were similarly stimulated by HBOT. The acceleration of wound contraction was not associated with increased myofibroblasts expression, nor fibroblast recruitment or higher cell count in the granulation tissue. Of note, we observed a significant increase in collagen deposition in early time points in ischemic wounds receiving HBOT. This data emphasizes that an early application of HBOT might be crucial to its efficacy. We concluded that wounds where ischemia and hyperglycemia are combined, as it is often the case in diabetic patients, have the best chance to benefit from HBOT.

Adipose-Derived Stromal Cells within a Gelatin Matrix Acquire Enhanced Regenerative and Angiogenic Properties: A Pre-Clinical Study for Application to Chronic Wounds.

This study evaluates the influence of a gelatin sponge on adipose-derived stromal cells (ASC). Transcriptomic data revealed that, compared to ASC in a monolayer, a cross-linked porcine gelatin sponge strongly influences the transcriptome of ASC. Wound healing genes were massively regulated, notably with the inflammatory and angiogenic factors. Proteomics on conditioned media showed that gelatin also acted as a concentrator and reservoir of the regenerative ASC secretome. This secretome promoted fibroblast survival and epithelialization, and significantly increased the migration and tubular assembly of endothelial cells within fibronectin. ASC in gelatin on a chick chorioallantoic membrane were more connected to vessels than an empty sponge, confirming an increased angiogenesis in vivo. No tumor formation was observed in immunodeficient nude mice to which an ASC gelatin sponge was transplanted subcutaneously. Finally, ASC in a gelatin sponge prepared from outbred rats accelerated closure and re-vascularization of ischemic wounds in the footpads of rats. In conclusion, we provide here preclinical evidence that a cross-linked porcine gelatin sponge is an optimal carrier to concentrate and increase the regenerative activity of ASC, notably angiogenic. This formulation of ASC represents an optimal, convenient and clinically compliant option for the delivery of ASC on ischemic wounds.

NADPH oxidase 4 is dispensable for skin myofibroblast differentiation and wound healing.

Differentiation of fibroblasts to myofibroblasts is governed by the transforming growth factor beta (TGF-ß) through a mechanism involving redox signaling and generation of reactive oxygen species (ROS). Myofibroblasts synthesize proteins of the extracellular matrix (ECM) and display a contractile phenotype. Myofibroblasts are predominant contributors of wound healing and several pathological states, including fibrotic diseases and cancer. Inhibition of the ROS-generating enzyme NADPH oxidase 4 (NOX4) has been proposed to mitigate fibroblast to myofibroblast differentiation and to offer a therapeutic option for the treatment of fibrotic diseases. In this study, we addressed the role of NOX4 in physiological wound healing and in TGF-ß-induced myofibroblast differentiation. We explored the phenotypic changes induced by TGF-ß in primary skin fibroblasts isolated from Nox4-deficient mice by immunofluorescence, Western blotting and RNA sequencing. Mice deficient for Cyba, the gene coding for p22phox, a key subunit of NOX4 were used for confirmatory experiments as well as human primary skin fibroblasts. In vivo, the wound healing was similar in wild-type and Nox4-deficient mice. In vitro, despite a strong upregulation following TGF-ß treatment, Nox4 did not influence skin myofibroblast differentiation although a putative NOX4 inhibitor GKT137831 and a flavoprotein inhibitor diphenylene iodonium mitigated this mechanism. Transcriptomic analysis revealed upregulation of the mitochondrial protein Ucp2 and the stress-response protein Hddc3 in Nox4-deficient fibroblasts, which had however no impact on fibroblast bioenergetics. Altogether, we provide extensive evidence that NOX4 is dispensable for wound healing and skin fibroblast to myofibroblast differentiation, and suggest that another H2O2-generating flavoprotein drives this mechanism.

Operative Planning of Chest Wall Reconstructions Illustrated by a Large Defect in a Child.

Reconstruction of large chest wall defects is challenging. Here we discuss the process of decision-making in planning chest wall reconstruction, considering the requirements of tumor removal, stabilization of the chest wall, and soft tissue coverage, illustrated by a case of a hemi-chest wall defect in a child. Ewing sarcoma measuring 10 × 9 × 13 cm was resected in a 9-year-old boy, followed by stabilization using a Gore-Tex patch. Due to extension of the oncologic resection far into the superomedial quadrant of the chest, tension-free coverage with a classical latissimus-dorsi flap could not be achieved. Integrating the serratus-anterior muscle into the flap creating a chimeric latissimus-dorsi/serratus-anterior flap allowed for excellent soft tissue coverage of the foreign body. As the skin could be preserved, careful incision planning was necessary to allow for best possible exposure during oncologic resection and flap harvest, while ensuring skin vascularization impaired by underlying tumor resection. Two vertical skin incisions were chosen, one presternal and a second in the mid-axillary fold delineating a large bipedicled skin flap. Postoperative recovery was excellent. Solid skin vascularization and adequate soft tissue coverage of the alloplastic material allowed for the patient to receive two cycles of postoperative radiotherapy without developing wound dehiscence. Careful interdisciplinary planning of skin incisions allowed for good exposure for tumor resection and flap harvest while preserving skin vascularization. Choosing a chimeric latissimus-dorsi/serratus-anterior flap provided larger coverage than a classical latissimus-dorsi flap with minimal additional donor site morbidity. Taken together, we here present a pragmatic solution to a complex problem.

Fate of systemically and locally administered adipose-derived mesenchymal stromal cells and their effect on wound healing.

There is increasing interest in the use of adipose-derived mesenchymal stromal cells (ASCs) for wound repair. As the fate of administered cells is still poorly defined, we aimed to establish the location, survival, and effect of ASCs when administered either systemically or locally during wound repair under physiological conditions. To determine the behavior of ASCs, a rat model with wounds on the dorsal aspect of the hind paws was used and two treatment modes were assessed: ASCs administered systemically into the tail vein or locally around the wound. ASCs were transduced to express both firefly luciferase (Fluc) and green fluorescent protein to enable tracking by bioluminescence imaging and immunohistological analysis. Systemically administered ASCs were detected in the lungs 3 hours after injection with a decrease in luminescent signal at 48 hours and signal disappearance from 72 hours. No ASCs were detected in the wound. Locally administered ASCs remained strongly detectable for 7 days at the injection site and became distributed within the wound bed as early as 24 hours post injection with a significant increase observed at 72 hours. Systemically administered ASCs were filtered out in the lungs, whereas ASCs administered locally remained and survived not only at the injection site but were also detected within the wound bed. Both treatments led to enhanced wound closure. It appears that systemically administered ASCs have the potential to enhance wound repair distally from their site of entrapment in the lungs whereas locally administered ASCs enhanced wound repair as they became redistributed within the wound bed.

Does non-activated platelet-rich plasma (PRP) enhance fat graft outcome? An assessment with 3D CT-scan in mice.

BACKGROUND: The adjunction of autologous platelet-rich plasma (PRP) is emerging as a promising approach to enhance the long-term survival of fat grafting, but there are still insufficient data on its efficacy. The aim of this in vivo study was to assess the effect of the addition of non-activated PRP on fat graft outcome. METHODS: Human adipose tissue mixed with 20% of non-activated PRP was injected under the scalp skin of nude Balb/cAnNRj mice and compared to grafted fat mixed with 20% of saline. The fat graft volume was analyzed by a computed tomography scan until day 90 and immunohistochemistry was then performed to assess adipocyte viability and graft revascularization. RESULTS: At day 90, the volume of fat graft was not enhanced by PRP compared to the saline control group. However, immunohistochemistry showed that PRP significantly increased the fat graft area occupied by intact adipocytes compared to the saline group (72.66% vs. 60.78%, respectively; p < 0.05). Vascularity was also significantly higher in the PRP group compared to the control group (6695 vs. 4244 CD31+ cells/µm2, respectively; p < 0.05). CONCLUSION: The adjunction of non-activated-PRP to fat grafts significantly increased adipocyte viability and tissue vascularity. However, in contrast to other studies adding activated-PRP, non-activated-PRP did not increase residual fat graft volume until day 90. Further studies are therefore needed to understand whether PRP has a positive effect on fat graft volume. As 3D computed tomography scan is a reproducible and precise technique, it should be used to analyze fat graft volume changes over time.