VEGF released by deferoxamine preconditioned mesenchymal stem cells seeded on collagen-GAG substrates enhances neovascularization

Hypoxia preconditioning of mesenchymal stem cells (MSCs) has been shown to promote wound healing through HIF-1 alpha stabilization. Preconditioned MSCs can be applied to three-dimensional biomaterials to further enhance the regenerative properties. While environmentally induced hypoxia has proven difficult in clinical settings, this study compares the wound healing capabilities of adipose derived (Ad) MSCs seeded on a collagen-glycosaminoglycan (GAG) dermal substrate exposed to either environmental hypoxia or FDA approved deferoxamine mesylate (DFO) to stabilize HIF-1 alpha for wound healing. The release of hypoxia related reparative factors by the cells on the collagen-GAG substrate was evaluated to detect if DFO produces results comparable to environmentally induced hypoxia to facilitate optimal clinical settings. VEGF release increased in samples exposed to DFO. While the SDF-1 alpha release was lower in cells exposed to environmental hypoxia in comparison to cells cultured in DFO in vitro. The AdMSC seeded biomaterial was further evaluated in a murine model. The implants where harvested after 1 days for histological, inflammatory, and protein analysis. The application of DFO to the cells could mimic and enhance the wound healing capabilities of environmentally induced hypoxia through VEGF expression and promises a more viable option in clinical settings that is not merely restricted to the laboratory

Towards autotrophic tissue engineering: Photosynthetic gene therapy for regeneration

Artículo científicoThe use of artificial tissues in regenerative medicine is limited due to hypoxia. As a strategy to overcome this drawback, we have shown that photosynthetic biomaterials can produce and provide oxygen independently of blood perfusion by generating chimeric animal-plant tissues during dermal regeneration. In this work, we demonstrate the safety and efficacy of photosynthetic biomaterials in vivo after engraftment in a fully immunocompetent mouse skin defect model. Further, we show that it is also possible to genetically engineer such photosynthetic scaffolds to deliver other key molecules in addition to oxygen. As a proof-of-concept, biomaterials were loaded with gene modified microalgae expressing the angiogenic recombinant protein VEGF. Survival of the algae, growth factor delivery and regenerative potential were evaluated in vitro and in vivo. This work proposes the use of photosynthetic gene therapy in regenerative medicine and provides scientific evidence for the use of engineered microalgae as an alternative to deliver recombinant molecules for gene therapy

Occlusive dressing-induced secretomes influence the migration and proliferation of mesenchymal stem cells and fibroblasts differently

Background: Fingertip injuries treated with occlusive dressings (ODs) lead to nearly scar-free, functionally, and aesthetically pleasing results. We hypothesized that paracrine factors in the wound fluid (secretome) may influence migration and proliferation of mesenchymal stem cells (MSCs) and fibroblasts and modulate the wound-healing process. Methods: We could collect wound fluid samples from 4 fingertip injuries and 7 split skin donor sites at the 5th day during dressing change. Blood serum samples served as controls. The proliferation rate of MSCs and fibroblasts (HS27) was continuously measured through impedance analysis for 60h and by Alamarblue analysis after 72h. Cell migration was evaluated continuously for 15h and confirmed by the in vitro wound-healing assay. Results: Migration of MSCs under the influence of both wound fluids was significantly faster than controls from 4 to 6h after incubation and reversed after 9h. MSC proliferation in wound fluid groups showed a significant increase at 5 and 10h and was significantly decreased after 45h. Fibroblasts in wound fluid groups showed overall a significant increase in migration and a significant decrease in proliferation compared to controls. Conclusion: OD-induced secretomes influence MSCs and fibroblasts and thereby possibly modulate wound healing and scar tissue formation

Towards a Reliable and Rapid Automated Grading System in Facial Palsy Patients: Facial Palsy Surgery Meets Computer Science

Background: Reliable, time- and cost-effective, and clinician-friendly diagnostic tools are cornerstones in facial palsy (FP) patient management. Different automated FP grading systems have been developed but revealed persisting downsides such as insufficient accuracy and cost-intensive hardware. We aimed to overcome these barriers and programmed an automated grading system for FP patients utilizing the House and Brackmann scale (HBS). Methods: Image datasets of 86 patients seen at the Department of Plastic, Hand, and Reconstructive Surgery at the University Hospital Regensburg, Germany, between June 2017 and May 2021, were used to train the neural network and evaluate its accuracy. Nine facial poses per patient were analyzed by the algorithm. Results: The algorithm showed an accuracy of 100%. Oversampling did not result in altered outcomes, while the direct form displayed superior accuracy levels when compared to the modular classification form (n = 86; 100% vs. 99%). The Early Fusion technique was linked to improved accuracy outcomes in comparison to the Late Fusion and sequential method (n = 86; 100% vs. 96% vs. 97%). Conclusions: Our automated FP grading system combines high-level accuracy with cost- and time-effectiveness. Our algorithm may accelerate the grading process in FP patients and facilitate the FP surgeon’s workflow

A Randomized Controlled Trial: Regenerative Effects, Efficacy and Safety of Erythropoietin in Burn and Scalding Injuries

In adult’s burn injuries belong to the top 15 causes of injury. Annually more than a million patients receive specialized treatment. Improving burned patients’ outcomes is still a challenge. Effects of erythropoietin (EPO) are reported to be pro-angiogenic, pro-regenerative, anti-inflammatory, immunomodulatory and hypoxia/ischemia protective. Study objectives were to demonstrate cytoprotective and regenerative effects of EPO in burned patients in terms of improved wound healing, reduced morbidity and mortality. This was a prospective, placebo-controlled, randomized, double-blind trial. The trial was conducted in 13 specialized burn care centers in Germany. Adult Patients with 2b° or 3° burn injuries were included. Patients received state of the art burn care including obligatory split skin graft transplantation. Study medication was EPO or placebo every other day for 21 days. Between 12/08 and 06/14, 116 patients were randomized, 84 received study medication (EPO 45, Placebo 39). Primary endpoint analysis revealed inconclusive results, as only a minority of patients reached the primary endpoint [100% re-epithelialization: EPO: 23% (9/40); Placebo 30% (11/37)]. Several secondary endpoints such as SOFA score (morbidity), EPO level in blood and wound healing onset revealed clinical, and statistically significant results in favor of the EPO group. Adverse Events (AEs) and Severe Adverse Events (SAEs) were in expected ranges; AEs EPO: 80%, (36/45), Placebo: 77%, (30/39); SAEs EPO: 24%, (11/45), Placebo: 24%, (8/39). Out of 84 patients two died, one per group, thus mortality was lower than expected. Results (SOFA score) indicate a lower morbidity of the EPO group, suggesting pro-regenerative effects of EPO in burned patients. Higher EPO levels might influence the faster onset of re-epithelialization in the first 10 days of the treatment. Both effects could reveal new therapeutic options.Clinical Trial Registration: ISRCT Number: ISRCTN95777824 and EudraCT Number: 2006-002886-38, Protocol Number: 0506

Enhanced Collateral Growth by Double Transplantation of Gene-Nucleofected Fibroblasts in Ischemic Hindlimb of Rats

BACKGROUND: Induction of neovascularization by releasing therapeutic growth factors is a promising application of cell-based gene therapy to treat ischemia-related problems. In the present study, we have developed a new strategy based on nucleofection with alternative solution and cuvette to promote collateral growth and re-establishment of circulation in ischemic limbs using double transplantation of gene nucleofected primary cultures of fibroblasts, which were isolated from rat receiving such therapy. METHODS AND RESULTS: Rat dermal fibroblasts were nucleofected ex vivo to release bFGF or VEGF165 in a hindlimb ischemia model in vivo. After femoral artery ligation, gene-modified cells were injected intramuscularly. One week post injection, local confined plasmid expression and transient distributions of the plasmids in other organs were detected by quantitative PCR. Quantitative micro-CT analyses showed improvements of vascularization in the ischemic zone (No. of collateral vessels via micro CT: 6.8±2.3 vs. 10.1±2.6; p<0.05). Moreover, improved collateral proliferation (BrdU incorporation: 0.48±0.05 vs. 0.57±0.05; p<0.05) and increase in blood perfusion (microspheres ratio: gastrocnemius: 0.41±0.10 vs. 0.50±0.11; p<0.05; soleus ratio: soleus: 0.42±0.08 vs. 0.60±0.08; p<0.01) in the lower hindlimb were also observed. CONCLUSIONS: These results demonstrate the feasibility and effectiveness of double transplantation of gene nucleofected primary fibroblasts in producing growth factors and promoting the formation of collateral circulation in ischemic hindlimb, suggesting that isolation and preparation of gene nucleofected cells from individual accepting gene therapy may be an alternative strategy for treating limb ischemia related diseases