Mesenchymal stem cells and myoblast differentiation under HGF and IGF-1 stimulation for 3D skeletal muscle tissue engineering.

BACKGROUND: Volumetric muscle loss caused by trauma or after tumour surgery exceeds the natural regeneration capacity of skeletal muscle. Hence, the future goal of tissue engineering (TE) is the replacement and repair of lost muscle tissue by newly generating skeletal muscle combining different cell sources, such as myoblasts and mesenchymal stem cells (MSCs), within a three-dimensional matrix. Latest research showed that seeding skeletal muscle cells on aligned constructs enhance the formation of myotubes as well as cell alignment and may provide a further step towards the clinical application of engineered skeletal muscle. In this study the myogenic differentiation potential of MSCs upon co-cultivation with myoblasts and under stimulation with hepatocyte growth factor (HGF) and insulin-like growth factor-1 (IGF-1) was evaluated. We further analysed the behaviour of MSC-myoblast co-cultures in different 3D matrices. RESULTS: Primary rat myoblasts and rat MSCs were mono- and co-cultivated for 2, 7 or 14 days. The effect of different concentrations of HGF and IGF-1 alone, as well as in combination, on myogenic differentiation was analysed using microscopy, multicolour flow cytometry and real-time PCR. Furthermore, the influence of different three-dimensional culture models, such as fibrin, fibrin-collagen-I gels and parallel aligned electrospun poly-ε-caprolacton collagen-I nanofibers, on myogenic differentiation was analysed. MSCs could be successfully differentiated into the myogenic lineage both in mono- and in co-cultures independent of HGF and IGF-1 stimulation by expressing desmin, myocyte enhancer factor 2, myosin heavy chain 2 and alpha-sarcomeric actinin. An increased expression of different myogenic key markers could be observed under HGF and IGF-1 stimulation. Even though, stimulation with HGF/IGF-1 does not seem essential for sufficient myogenic differentiation. Three-dimensional cultivation in fibrin-collagen-I gels induced higher levels of myogenic differentiation compared with two-dimensional experiments. Cultivation on poly-ε-caprolacton-collagen-I nanofibers induced parallel alignment of cells and positive expression of desmin. CONCLUSIONS: In this study, we were able to myogenically differentiate MSC upon mono- and co-cultivation with myoblasts. The addition of HGF/IGF-1 might not be essential for achieving successful myogenic differentiation. Furthermore, with the development of a biocompatible nanofiber scaffold we established the basis for further experiments aiming at the generation of functional muscle tissue.

De novo generation of an axially vascularized processed bovine cancellous-bone substitute in the sheep arteriovenous-loop model.

BACKGROUND/AIMS: The aim of this study was to generate an axially vascularized bone substitute. The arteriovenous (AV)-loop approach in a large-animal model was applied in order to induce axial vascularization in a clinically approved processed bovine cancellous bone (PBCB) matrix of significant volume with primary mechanical stability and to assess the course of increasing axial vascularization. METHODS: PBCB constructs were implanted into 13 merino sheep together with a microsurgically created AV loop in an isolation chamber. The vascularization process was monitored by sequential magnetic resonance imaging (MRI) scans. Explants were subjected to micro-computed tomography (micro-CT) analysis, histomorphometry and immunohistochemistry for CD31 and CD45. RESULTS: Increasing axial vascularization in PBCB constructs was quantified by histomorphometry and visualized by micro-CT scans. Intravital sequential MRI scans demonstrated a significant progressive increase in perfused volume within the matrices. Immunohistochemistry confirmed endothelial lining of newly formed vessels. CONCLUSION: This study demonstrates successful axial vascularization of a clinically approved, mechanically stable bone substitute with a significant volume by a microsurgical AV loop in a large-animal model. Thus microsurgical transplantation of a tissue-engineered, axially vascularized and mechanically stable bone substitute with clinically relevant dimensions may become clinically feasible in the future.

[Establishing and evaluating a multinational Science Academy: A new tool for fostering scientific young talents in microsurgical research - a Consensus Statement of the DAM]. / Etablierung und Evaluation einer multinationalen Wissenschaftsakademie: Ein neues Modell zur Stärkung des wissenschaftlichen Nachwuchses in der mikrochirurgischen Forschung ­ Ein Konsensus Papier der DAM.

BACKGROUND: The formation of professional networks and cooperations - in addition to any qualified good education - seems fundamental for a successful career. In a number of disciplines, various symposia or conferences exist. In the field of microsurgery, however, a specific, guided and designated opportunity for junior scientists to network with one another has been missing so far. METHODS: In 2017, a science academy was initiated for the first time by the German-speaking Association for Nerves and Vessels (DAM) with the goal of bringing together and networking microsurgically researching young physicians and scientists. This was intended to happen on a small scale once a year in order to develop synergies for joint research projects. For this purpose, motivated junior researchers were individually selected by their mentors and sent to the academy by the boards of research institutions that are organized in the DAM. After getting to know each other in a relaxed atmosphere, the participants were given the opportunity to present their respective research project within the framework of thematic blocks and moderated by experienced mentors. Each presentation was followed by a round table discussion and small group work, in which knowledge and methods were exchanged and points of contact for possible later cooperation were identified. RESULTS: In the past 3 years, the DAM Science Academy proved to be an optimal format to initiate and promote networks of young researchers comprising microsurgically interested physicians and scientists. There were many lively and in-depth discussions, which were mainly due to the open working atmosphere and the obligation to confidentiality. Most of the synergies were shown i. a. in the field of angiogenesis, bioreactor, carcinoma-ADSC interactions, stem cells, AV loop model, ischemia/reperfusion, and nerve regeneration. The participants consistently gave a very positive feedback in the final evaluation with the wish to continue this academy. CONCLUSION: The DAM Science Academy can be considered a highly suitable complemental platform to the existing networking opportunities among microsurgical researchers. Experience so far suggests that this will hopefully result in long-term cooperations and a permanent transfer of knowledge among the participants.

Scalp reconstruction: A 10-year retrospective study.

Scalp reconstruction is a challenging task for the reconstructive surgeon. In consideration of the anatomical and cosmetic characteristics, the defect depth and size, an armamentarium of reconstructive procedures ranging from skin grafts over local flaps to free tissue transfer has been described. In this 10-year retrospective study, 85 operative procedures for scalp reconstruction were performed at our department. The underlying entity, defect size/depth, reconstructive procedure, complications, and mean hospital stay were analyzed. In most cases, scalp reconstruction was necessary after oncologic resection (67%) or radiation therapy (16%). A total of 85 operative procedures were performed for scalp reconstruction including local flaps (n = 50), free tissue transfer (n = 18), and skin grafts (n = 17). Regarding the complication rate, we could detect an overall major complication rate of 16.5% with one free flap loss. Briefly, local flaps are an adequate and safe procedure for limited scalp defects. In the case of extensive scalp defects affecting the calvarium, prior multiple surgical interventions and/or radiation, we prefer free tissue transfer.

[Establishment and Evaluation of a Microsurgery Course for Medical Students]. / Etablierung und Evaluation eines Mikrochirurgie - Kurses für Medizinstudierende.

BACKGROUND: Very few microsurgical courses have been offered for medical students in Germany to date. To raise early interest in this technique, which is essential for plastic and reconstructive surgery, and to guide eligible medical students to choose plastic surgery as their specialist field, the Department of Plastic and Hand Surgery, supported by the Faculty of Medicine of the Friedrich-Alexander-University of Erlangen-Nuremberg, implemented a microsurgical course for students in 2011. This study describes the implementation of that course and evaluates its impact on the subsequent choice of the participants' specialist fields. MATERIAL AND METHODS: Since the summer of 2011, the microsurgery course for medical students has taken place regularly 3 times per term. It is free of charge for participants and is guided by senior physicians of the Department of Plastic and Hand Surgery together with student tutors from the Faculty of Medicine. The arterial end-to-end anastomosis in the fresh chicken leg is used as a training model. Based on a questionnaire survey the participants were evaluated and statistically analysed regarding their course satisfaction, self-assessment of their own eligibility before and after the course, the anticipated future choice of their medical specialist field and how their choice was influenced by this course. RESULTS: After the successful implementation of the microsurgical course in 2011, a significant number of students were interested in microsurgery. According to the questionnaire, the level of enthusiasm was high among all participants. The self-assessment of microsurgical skills improved significantly after the course compared with the pre-course assessment. In 82% of the participants, the course had a strong positive influence on the future choice of their specialist field. CONCLUSIONS: The regular implementation of a microsurgical course for students in the form described here is practicable and possible without undue personnel and cost of materials. The ongoing interest among students in such an offer is enormous and the satisfaction of the participants is very high. This might be a way to recruit future plastic surgeons by raising early enthusiasm for microsurgery. These future plastic surgeons, in turn, would be given the chance to experience a very fascinating aspect of plastic surgery, which might help them to decide on their specialisation within that field at a later point in their career.

[Skeletal muscle tissue engineering--current concepts and future perspectives]. / Tissue Engineering von Skelettmuskelgewebe--Stand und Perspektiven.

Tissue engineering of skeletal muscle could have great advantages in every clinical setting in need of neurovascular muscle transfer, e. g., facial palsy or Volkmann's contracture. There are 2 great obstacles for the clinical application of engineered muscle tissue at the moment: firstly, finding a three-dimensional matrix that matches the demands concerning biocompatibility, stability and elasticity; secondly, the insufficient differentiation of implanted myoblasts, since myoblast differentiation in vivo is barely controllable and subject to a variety of influences. Furthermore axial vascularisation and neurotisation of such tissue-engineered skeletal muscle constructs play a pivotal role for any later application. An overview of the current status of skeletal muscle tissue engineering technologies and concepts for future perspective in this emerging field is presented in this article.

[Bone tissue engineering for bone defect therapy]. / Knochen Tissue Engineering zur Therapie von Knochendefekten.

In critical size bone defects resulting from failed fracture healing or pseudarthrosis surgery is usually required. In this context, autologous bone grafts and callus distraction represent the gold standard, while sometimes even vascularised bone transfer is mandatory including microsurgical techniques. The availability of donor sites for such procedures is limited and the resulting morbidity significant. Therefore, synthetic bone grafts have been developed as an alternative. They consist of a broad range of different materials such as natural and synthetic polymers, ceramic and compound materials, aiming to mimic the three-dimensional character of autografts. In addition, they may act as a delivery vehicle for growth factors, antibiotics or cells. Their main limitation has been the lack of an intrinsic blood supply, limiting the potential for transplantation. This review provides an overview of matrices, cells and other therapeutic substances in the field of bone tissue engineering.

[Skin tissue engineering--from split skin to engineered skin grafts?]. / Tissue Engineering von Haut--von der Spalthaut zum gezüchteten Hauttransplantat?

Today split or full skin grafts are still the gold standard in the treatment of substance defects of the skin. Such results can be seen, for example, in the therapy for burn patients. However, in patients with more than 50% burned skin area, donor sites are limited. Likewise in chronic wound patients inferior take rates of skin grafts as compared to burn wounds are observed. This may be attributed, for example, to accompanying or underlying chronic diseases or a higher rate of local infections. These phenomena also lead to a lack of availability of transplantable skin grafts. Hence the need for cost effective and user friendly synthetic or engineered skin grafts, which can serve for acute and chronic wounds and which can be also used in critically ill patients, is at hand. During the last 30 years a huge number of biological and synthetic skin graft materials and products based on the patient's own cells were launched on the market. Researchers and clinicians are constantly working on further improvements. One possibility is the engineering of skin grafts in vitro, which have to be integrated into the wound bed after transplantation. Another approach is the fabrication of biocompatible and bioresorbable matrices, which can attract host cells and stimulate a wound-healing process without scars. However, the skin graft materials available today cannot yet replace split or full skin grafts completely because of their inherent limitations such as insufficient take rates and/or the lack of mechanical stability and differentiated structures of the grafted artificial skin. Thus researchers in the field of skin tissue engineering are still working on the final goal of developing a skin graft which has all the features of healthy human skin and is capable of replacing human skin completely. This article gives on overview of the currently available solutions and products in the field of skin tissue engineering.