Regenerative medicine for skeletal muscle loss: a review of current tissue engineering approaches.

Skeletal muscle is capable of regeneration following minor damage, more significant volumetric muscle loss (VML) however results in permanent functional impairment. Current multimodal treatment methodologies yield variable functional recovery, with reconstructive surgical approaches restricted by limited donor tissue and significant donor morbidity. Tissue-engineered skeletal muscle constructs promise the potential to revolutionise the treatment of VML through the regeneration of functional skeletal muscle. Herein, we review the current status of tissue engineering approaches to VML; firstly the design of biocompatible tissue scaffolds, including recent developments with electroconductive materials. Secondly, we review the progenitor cell populations used to seed scaffolds and their relative merits. Thirdly we review in vitro methods of scaffold functional maturation including the use of three-dimensional bioprinting and bioreactors. Finally, we discuss the technical, regulatory and ethical barriers to clinical translation of this technology. Despite significant advances in areas, such as electroactive scaffolds and three-dimensional bioprinting, along with several promising in vivo studies, there remain multiple technical hurdles before translation into clinically impactful therapies can be achieved. Novel strategies for graft vascularisation, and in vitro functional maturation will be of particular importance in order to develop tissue-engineered constructs capable of significant clinical impact.

Optimizing the decellularization process of an upper limb skeletal muscle; implications for muscle tissue engineering.

Upper limb muscle reconstruction is required following cancer resection, trauma, and congenital deformities. Current surgical reconstruction of the muscle involves local, regional and free flaps. However, muscle reconstruction is not always possible due to the size of the defect and functional donor site morbidity. These challenges could be addressed with the production of scaffolds composed of an extracellular matrix (ECM) derived from decellularized human skeletal muscle. This study aimed to find an optimal technique to decellularize a flexor digitorum superficialis muscle. The first two protocols were based on a detergent only (DOT) and a detergent-enzymatic protocol (DET). The third protocol avoided the use of detergents and proteolytic enzymes (NDNET). The decellularized scaffolds were characterized using qualitative techniques including histological and immunofluorescent staining and quantitative techniques assessing deoxyribonucleic acid (DNA), glycosaminoglycan (GAG), and collagen content. The DOT protocol consisting of 2% SDS for 4 hours was successful at decellularizing human FDS, as shown by DNA content assay and nuclei immunofluorescence staining. The DOT protocol maintained the microstructure of the scaffolds as shown by Masson's trichrome staining and collagen and GAG content. DET and NDNET protocols maintained the ECM, but were unsuccessful in removing all DNA content after two cycles of decellularization. Decellularization of skeletal muscle is a viable option for muscle reconstruction using a detergent only technique for upper limb defects. Further testing in vivo will assess the effectiveness of decellularized scaffolds for upper limb muscle skeletal tissue engineering.

Fat Grafting Improves Fibrosis and Scarring in Vulvar Lichen Sclerosus: Results From a Prospective Cohort Study.

OBJECTIVE: The aim of the study was to evaluate the effect of lipotransfer in women presenting with fibrosis and scarring due to lichen sclerosus. MATERIALS AND METHODS: This prospective cohort study included 33 women attending the vulvar clinic of a public hospital. Patients received one lipotransfer treatment. Validated measures were used prospectively to assess the sexual function (Female Sexual Function Index, Female Sexual Distress Scale); symptoms (visual analog scale for itching, burning, soreness), pain (Pain Anxiety Symptoms Scale 20); psychological status and quality of life (Hospital Anxiety and Depression Scale, Relationship Assessment Scale, Wound Management Questionnaire Revised); physician-based disease signs (Vulvar Architecture Severity Scale). Data were analyzed using paired t test with nonparametric Wilcoxon matched-pairs signed rank test and unpaired t test with nonparametric Mann-Whitney test (Prism6 Software). RESULTS: The mean (SD) follow-up was 12.9 (3.5) months. Sexual function improved after treatment (p < .001), as well as the distress associated with sexuality (p < .0001). A significant improvement was reported in itching (p < .001), burning (p < .05), soreness (p < .001), and pain (p < .0001). Patients reported a significant improvement in romantic relationship (p < .05), anxiety (p < .0001), and depression (p < .0001). Improvement was not significant in the self-care associated with self-disgust assessment (p = .42). The clinical physician-based score showed an overall improvement in all the treated areas to lesser or greater extent. CONCLUSIONS: The use of fat grafting in lichen sclerosus is promising. Further studies are required to rule out a potential placebo effect and to better understand the underlying molecular mechanism of action.

Measuring Differential Volume Using the Subtraction Tool for Three-Dimensional Breast Volumetry: A Proof of Concept Study.

Background. Three-dimensional (3D) photography provides a promising means of breast volumetry. Sources of error using a single-captured surface to calculate breast volume include inaccurate designation of breast boundaries and prediction of the invisible chest wall generated by computer software. An alternative approach is to measure differential volume using subtraction of 2 captured surfaces. Objectives. To explore 3D breast volumetry using the subtraction of superimposed images to calculate differential volume. To assess optimal patient positioning for accurate volumetric assessment. Methods. Known volumes of breast enhancers simulated volumetric changes to the breast (n = 12). 3D photographs were taken (3dMDtorso) with the subject positioned upright at 90° and posteriorly inclined at 30°. Patient position, breathing, distance and camera calibration were standardised. Volumetric analysis was performed using 3dMDvultus software. Results. A statistically significant difference was found between actual volume and measured volumes with subjects positioned at 90° (P < .05). No statistical difference was found at 30° (P = .078), but subsequent Bland-Altman analysis showed evidence of proportional bias (P < .05). There was good correlation between measured and actual volumes in both positions (r = .77 and r = .85, respectively). Univariate analyses showed breast enhancer volumes of 195 mL and 295 mL to incur bias. The coefficient of variation was 5.76% for single observer analysis. Conclusion. Positioning the subject at a 30° posterior incline provides more accurate results from better exposure of the inferior breast. The subtraction tool is a novel method of measuring differential volume. Future studies should explore methodology for application into the clinical setting.

Development and Validation of a Clinical Grading Scale to Assess the Vulvar Region: The Vulvar Architecture Severity Scale.

BACKGROUND: The vulva is composed of aesthetic units that can be affected differently by vulvar conditions. A reliable, comprehensive, and quick-to-use clinical scoring system is required to assess the disease extent in the vulvar area. OBJECTIVES: The aim of this study was to develop and validate a grading scale based on the aesthetic unit principle to evaluate the extent of vulvar lichen sclerosus (VLS). METHODS: After reviewing photographs of 100 patients affected by VLS, the authors targeted the aesthetic units most frequently affected. The disease signs were recorded and graded in 4 levels of severity (none, mild, moderate, severe) taking into account the vulvar architecture and skin involvement. To validate the scale, 14 observers were asked to apply it to photographs of 25 VLS patients on 2 different occasions. Intra- and inter-observer reliabilities were determined employing Pearson's and intraclass correlation coefficients. RESULTS: A 6-region, 4-point grading system was designed and identified as the Vulvar Architecture Severity Scale (VASS). In all 6 areas, the Pearson's r was greater than 0.9 (mean, 0.994; 95% confidence interval [CI] = 0.992), indicating that the intra-observer reliability of the VASS was consistent over time (P < 0.001). Intraclass correlation at time 1 was 0.928 (95% CI = 0.910, 0.943) and at time 2 was 0.944 (95% CI = 0.931, 0.996), indicating a high reliability level among different observers. CONCLUSIONS: The VASS is a reliable scale to assess the severity of VLS, and it might be considered as an outcome measure in future VLS trials.

Fat Hypertrophy as a Complication of Fat Transfer for Hemifacial Atrophy.

Fat hypertrophy is a less commonly known complication of autologous fat transfer. We present a 32-year-old female with left hemifacial atrophy associated with systemic sclerosis, who was treated with 7 fat transfer procedures to correct the facial asymmetry. A total of 236.5 mL of fat was injected to the hemiface over a 4-year period to achieve good symmetry. A progressively enlarging, painless, soft mass over the left parotid region was noted at 3 months after the final fat transfer procedure. Magnetic resonance imaging showed a markedly enlarged bulk of subcutaneous fat over the left cheek with no evidence of necrosis, edema, or pathologic enhancement. Concurrent weight gain was noted secondary to additional nutritional input. The patient's aesthetic, symptomatic, and functional concerns led to the subsequent removal of 115 mL fat by liposuction.

Use of Lipotransfer in Scleroderma.

Lipotransfer for soft tissue filling is a well-established plastic and aesthetic surgical technique. Elective fat grafting is considered a safe and well-tolerated procedure. Coleman and others have reported that fat grafting may have tissue regenerative properties and not only serve as a soft tissue filler. There have been reports from our group and others that it may improve fibrosis secondary to many different pathological aetiologies including scleroderma, burn injury, lichen sclerosis, graft vs host disease, and radiation. The mechanism of action remains unclear but has been postulated that is adipose derived stem cells (ADSCs) related. Lipoaspirate has been characterised and shown to contain several cell populations including fibroblasts, endothelial cells, and ADSCs. The ADSCs have shown to secrete angiogenic, immunodulatory, and antiapoptotic factors as well as proliferate and differentiate into different cell types similarly to other stem cell sources. We have shown that ADSCs are functionally different in scleroderma patients but despite this lipotransfer produces a significant reversal in the effects of fibrosis in these patients. The advantage of lipoaspirate containing a valuable source of regenerative properties, ease of access, isolation, and processing may serve a significant future role in the treatment of fibrotic conditions.

Development of mechano-responsive polymeric scaffolds using functionalized silica nano-fillers for the control of cellular functions.

We demonstrate an efficient method to produce mechano-responsive polymeric scaffolds which can alter cellular functions using two different functionalized (OH and NH2) silica nano-fillers. Fumed silica-hydroxyl and fumed silica-amine nano-fillers were mixed with a biocompatible polymer (POSS-PCU) at various wt% to produce scaffolds. XPS and mechanical testing demonstrate that bulk mechanical properties are modified without changing the scaffold's surface chemistry. Mechanical testing showed significant change in bulk properties of POSS-PCU scaffolds with an addition of silica nanofillers as low as 1% (P<0.01). Scaffolds modified with NH2 silica showed significantly higher bulk mechanical properties compared to the one modified with the OH group. Enhanced cell adhesion, proliferation and collagen production over 14days were observed on scaffolds with higher bulk mechanical properties (NH2) compared to those with lower ones (unmodified and OH modified) (P<0.05) during in vitro analysis. This study provides an effective method of manufacturing mechano-responsive polymeric scaffolds, which can help to customize cellular responses for biomaterial applications.

A polyhedral oligomeric silsesquioxane-based bilayered dermal scaffold seeded with adipose tissue-derived stem cells: in vitro assessment of biomechanical properties.

BACKGROUND: Although commercial skin substitutes are widely available, its use remains challenging at surgery and postoperatively. The high cost is also prohibitive. We designed and characterized a scaffold for dermal replacement, using advanced nanocomposite materials, which are known to have unique nanoscale features that enhance cellular behavior. METHODS: A bilayered scaffold was developed using the nanocomposite, polyhedral oligomeric silsesquioxane, incorporated into poly(caprolactone-urea)urethane, resulting in a mechanically robust bioabsorbable polymer; forming the inner layer, which was designed with a range of porosities. The removable outer layer contained nanosilver. Tensile testing, surface tension, permeability, and scanning electron microscopy were performed. Optimal pore morphology for cellular proliferation was elucidated through adipose tissue-derived stem cell culture and a cell viability assay. All tests were repeated on Integra Dermal Regeneration Template. RESULTS: The physical construct was easy to handle and clinically applicable. Macroporosity and permeability of scaffolds was demonstrated, confirmed by scanning electron microscopy. Both tensile strength and surface tension were comparable with skin; outer layer demonstrated hydrophobicity and inner layer showed hydrophilicity. Cell assay confirmed cellular proliferation onto the scaffold, comparable with Integra. CONCLUSIONS: We demonstrate that a porous bilayered dermal scaffold could form the basis of a new generation of skin substitute that is both mechanically robust and harbors the ability for enhancing cell regeneration.

Stable mixed hematopoietic chimerism permits tolerance of vascularized composite allografts across a full major histocompatibility mismatch in swine.

This study tested the hypothesis that vascularized composite allografts (VCA) could be accepted in a robust model of hematopoietic chimerism by injecting allogeneic bone marrow cells (BMC) into swine fetuses. Outbred Yorkshire sows and boars were screened to ensure the absence of the major histocompatibility (MHC) allele SLA(cc) of inbred MGH miniature swine and then mated. Bone marrow harvested from an SLA(cc) swine donor was T-cell depleted and injected intravenously into the fetuses between days 50-55 of gestation. After birth, the piglets were studied with flow cytometry to detect donor cells and mixed lymphocyte reactions (MLR) and cell-mediated lympholysis (CML) assays to assess their response to donor. Donor-matched VCAs from SLA(cc) donors were performed on four chimeric and two nonchimeric swine. The results showed donor cell engraftment and multilineage macrochimerism after the in utero transplantation of adult BMC, and chimeric animals were unresponsive to donor antigens in vitro. Both control VCAs were rejected by 21 days and were alloreactive. Chimeric animals accepted the VCAs and never developed antidonor antibodies or alloreactivity to donor. These results confirm that the intravascular, in utero transplantation of adult BMC leads to donor cell chimerism and donor-specific tolerance of VCAs across a full MHC barrier in this animal model.

The emerging role of robotics in plastic and reconstructive surgery: a systematic review and meta-analysis.

The role of robotics has grown exponentially. There is an active interest amongst practitioners in the transferability of the potential benefits into plastic and reconstructive surgery; however, many plastic surgeons report lack of widespread implementation, training, or clinical exposure. We report the current evidence base, and surgical opportunities, alongside key barriers, and limitations to overcome, to develop the use of robotics within the field. This systematic review of PubMed, Medline, and Embase has been conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PROSPERO (ID: CRD42024524237). Preclinical, educational, and clinical articles were included, within the scope of plastic and reconstructive surgery. 2, 181, articles were screened; 176 articles met the inclusion criteria across lymph node dissection, flap and microsurgery, vaginoplasty, craniofacial reconstruction, abdominal wall reconstruction and transoral robotic surgery (TOR). A number of benefits have been reported including technical advantages such as better visualisation, improved precision and accuracy, and tremor reduction. Patient benefits include lower rate of complications and quicker recovery; however, there is a longer operative duration in some categories. Cost presents a significant barrier to implementation. Robotic surgery presents an exciting opportunity to improve patient outcomes and surgical ease of use, with feasibility for many subspecialities demonstrated in this review. However, further higher quality comparative research with careful case selection, which is adequately powered, as well as the inclusion of cost-analysis, is necessary to fully understand the true benefit for patient care, and justification for resource utilisation.

Exploring Anti-Fibrotic Effects of Adipose-Derived Stem Cells: Transcriptome Analysis upon Fibrotic, Inflammatory, and Hypoxic Conditioning.

Autologous fat transfers show promise in treating fibrotic skin diseases, reversing scarring and stiffness, and improving quality of life. Adipose-derived stem cells (ADSCs) within these grafts are believed to be crucial for this effect, particularly their secreted factors, though the specific mechanisms remain unclear. This study investigates transcriptomic changes in ADSCs after in vitro fibrotic, inflammatory, and hypoxic conditioning. High-throughput gene expression assays were conducted on ADSCs exposed to IL1-ß, TGF-ß1, and hypoxia and in media with fetal bovine serum (FBS). Flow cytometry characterized the ADSCs. RNA-Seq analysis revealed distinct gene expression patterns between the conditions. FBS upregulated pathways were related to the cell cycle, replication, wound healing, and ossification. IL1-ß induced immunomodulatory pathways, including granulocyte chemotaxis and cytokine production. TGF-ß1 treatment upregulated wound healing and muscle tissue development pathways. Hypoxia led to the downregulation of mitochondria and cellular activity.