Surviving an Extensive Burn Injury Using Advanced Skin Replacement Technologies

There have been significant improvements in the technology available for treating extensive burns in the past decade. This case presents two unique, skin replacement technologies that were used to treat an 86% surface area flame burn in a pediatric patient. A temporary dermal replacement, known as “Novosorb™ Biodegradable Temporizing Matrix” was first used to stabilize the burn injury and remained in place for approximately three months. Given the large burn size and lack of available donor skin for grafting, a permanent skin replacement product known as “Self-Assembled Skin Substitute (SASS)” was then utilized to cover the burns. SASS is a novel technology that was developed to replace skin as an autologous skin graft and is currently available in Canada through a clinical trial for major burns. Ultimately, the concurrent use of these two technologies allowed for the unprecedented survival of a child following an extensive and life-threatening burn injury.

Identification and functional characterisation of a rare MTTP variant underlying hereditary non-alcoholic fatty liver disease.

Background and aims: Non-alcoholic fatty liver disease (NAFLD) is a complex trait that has a global prevalence estimated as 25%. We aimed to identify the genetic variant underlying a four-generation family with progressive NAFLD leading to cirrhosis, decompensation and development of hepatocellular carcinoma in the absence of common risk factors such as obesity and type 2 diabetes. Methods: Exome sequencing and genome comparisons were used to identify the likely causal variant. We extensively characterised the clinical phenotype and post-prandial metabolic responses of family members with the identified novel variant in comparison to healthy non-carriers and wild type patients with NAFLD. Variant-expressing hepatocyte-like cells (HLCs) were derived from human induced pluripotent stem cells generated from homozygous donor skin fibroblasts. The phenotype was assessed using imaging, targeted RNA analysis and molecular expression arrays. Results: We identified a rare causal variant in MTTP, c.1691T>C p.I564T (rs745447480) encoding microsomal triglyceride transfer protein (MTP) associated with progressive non-alcoholic fatty liver disease, unrelated to metabolic syndrome. Although other described mutations in MTTP cause abetalipoproteinemia, neither homozygotes nor heterozygotes exhibited characteristic manifestations of this severe disease. HLCs derived from a homozygote donor had lower lipoprotein ApoB secretion, compared to wild type cells. Cytoplasmic triglyceride accumulation in HLCs triggered endoplasmic reticulum stress, secretion of pro-inflammatory mediators and production of reactive oxygen species. Conclusion: We have identified and characterized a rare causal variant in MTTP and homozygosity for MTTP p.I564T is associated with progressive NAFLD without any other manifestations of abetalipoproteinemia.

Pengaruh Platelet Rich Fibrin Pada Proses Epitelisasi Luka Donor Skin Graft: Studi Meta Analisis

Pendahuluan. Kehilangan kulit yang terlalu luas perlu jaringan penutup untuk mengatasinya, salah satu pilihan untuk menutup luka tersebut dengan melakukan tindakan skin grafting. Berdasarkan data IRJ Bedah Plastik Rekonstruksi dan Estetik RSUD Dr. Soetomo dalam 1 tahun (2017-2018) didapatkan pada 26 dari 50 kasus donor skin graft mengalami penyembuhan yang lebih dari waktu penyembuhan normal. Metode penyembuhan luka telah mengalami perkembangan beberapa tahun terakhir, salah satunya mulai dikenal peran platelet-rich fibrin (PRF).Metode. Penelitian ini merupakan penelitian dengan desain meta-analisis yang bersifat kuantitatif. Sumber data dari penelitian ini melalui penelusuran literatur di pencarian elektronik dengan menggunakan keyword pencarian literature. Database yang digunakan adalah Medline dan Pubmed antara tahun 2005-2020.Hasil. Seleksi literatur didapatkan 5 studi, dengan 3 studi subjek menggunakan donor split thickness skin graft dan 2 studi menggunakan donor free gingival graft. Dalam 3 studi menyebutkan pemberian platelet-rich fibrin (PRF) dapat mempercepat waktu penyembuhan dan epitelisasi. Hasil meta analisis menunjukkan tidak ada perbedaan antara kelompok pemberian platelet-rich fibrin (PRF) dan kelompok kontrol dalam proses epitelisasi pada donor skin graft (summary effect 1.30, 95% CI -0.42 – 3.02). Kesimpulan. Bukti - bukti preklinik berdasarkan studi meta-analisis ini menunjukkan bahwa tidak ada pengaruh signifikan terhadap pemberian platelet-rich fibrin (PRF) pada donor skin graft dalam kecepatan epitelisasi

537 Autologous Skin Cell Suspension for the Treatment of Small (≤10% TBSA) Mixed-Depth Burns

Introduction Split-thickness skin grafts (STSGs) have been the standard of care for many decades. Despite their widespread use, STSGs frequently fail. Autologous skin cell suspension (ASCS) is an FDA approved point of care regenerative medicine technology that reduces donor skin requirements without compromising clinical outcomes. ASCS allows for early treatment and less donor skin harvested that may be useful for hard-to-treat anatomical locations, in compromised patients that have risks for impaired wound healing, or elderly patients with thinner skin. We examined ASCS treatment as an adjunct to meshed autografts in adults with small mixed-depth/full-thickness burns. Methods We obtained IRB-approval for a prospective, multi-center, uncontrolled observational study that allowed continued access to ASCS before FDA approval (ClinicalTrials.gov Identifiers: NCT03333941). Subjects with mixed-depth/full-thickness injuries that required skin grafting with a minimum treatment area of 320 cm2 and burns ranging from 5–50% TBSA were eligible for study enrollment. Our analyses included only patients >18 years of age and ≤10% TBSA mixed-depth/full-thickness injuries that had completed the trial. All subjects had ≥1 burn wounds treated with meshed autografts (2:1–4:1) in combination with ASCS. Healing outcomes were accessed following ASCS treatment by direct visualization of each individual wound and included healing, scar outcomes, and safety data. Results Analyses included 20 subjects older than 18 years of age with ≤10% TBSA mixed-depth/full-thickness injuries. Of these, compromised wound healing was seen in 50.0% of subjects. Burn wounds with ≥90% re-epithelialization increased over time, with 62%, 80%, and 100% of wounds achieving closure at Weeks 1, 2, and 8, respectively. Similar results were seen in subjects with comorbidities known to affect wound healing and in elderly subjects despite their risks for impaired healing. Total POSAS patient (37.8 and 35.4) and observer scores were comparable (23.3 and 18.4) at Weeks 12 and 24. Safety events were typical for this patient population, and no serious adverse events occurred for any of the wounds. Conclusions This analysis provides additional information supporting the use of ASCS for the treatment of small, mixed-depth/full-thickness acute thermal burn injuries in adults, notably those with risk factors for impaired wound healing.

Dual mTOR/DNA-PK Inhibitor CC-115 Induces Cell Death in Melanoma Cells and Has Radiosensitizing Potential

CC-115 is a dual inhibitor of the mechanistic target of rapamycin (mTOR) kinase and the DNA-dependent protein kinase (DNA-PK) that is currently being studied in phase I/II clinical trials. DNA-PK is essential for the repair of DNA-double strand breaks (DSB). Radiotherapy is frequently used in the palliative treatment of metastatic melanoma patients and induces DSBs. Melanoma cell lines and healthy-donor skin fibroblast cell lines were treated with CC-115 and ionizing irradiation (IR). Apoptosis, necrosis, and cell cycle distribution were analyzed. Colony forming assays were conducted to study radiosensitizing effects. Immunofluorescence microscopy was performed to determine the activity of homologous recombination (HR). In most of the malign cell lines, an increasing concentration of CC-115 resulted in increased cell death. Furthermore, strong cytotoxic effects were only observed in malignant cell lines. Regarding clonogenicity, all cell lines displayed decreased survival fractions during combined inhibitor and IR treatment and supra-additive effects of the combination were observable in 5 out of 9 melanoma cell lines. CC-115 showed radiosensitizing potential in 7 out of 9 melanoma cell lines, but not in healthy skin fibroblasts. Based on our data CC-115 treatment could be a promising approach for patients with metastatic melanoma, particularly in the combination with radiotherapy.

The Reconstructed Human Epidermis in vitro — a Model for Basic and Applied Research of Human Skin

Background. The reconstructed human epidermis (RE) is an in vitro tissue-engineering construct similar to the native epidermis. Objective. To develop a full-layer RE. Describe its structure: determine the presence of all layers of the epidermal component, including basal, spinous and granular layers and stratum corneum of the epidermis; detect the basement membrane, the border between the epidermal and mesenchymal component. Materials and methods. Isolation of keratinocytes and fibroblasts from human donor skin. Cultivation of keratinocytes and fibroblasts in vitro under 2D conditions, cell subculturing and 3D modeling of RE, obtaining cryosections, histological staining, immunohistochemical (IHC) study with antibodies to cytokeratins 14 and 10, Ki67 protein, loricrin, laminin 5 and plectin. Results. A technique was developed for the formation of RE. Histological examination showed that the stratification of keratinocyte layers occurs during the formation of RE. Layers are formed including basal, spinous and granular layers and stratum corneum. The IHC study has shown the proliferative activity of keratinocytes of the basal layer and has detected the presence of marker proteins of keratinocytes at different stages of differentiation. RE basal keratinocytes, like native ones, form hemidesmosomes and synthesize basement membrane proteins. Conclusions. A full-layer human RE was obtained in vitro. RE meets all the characteristics of the native epidermis and it is suitable for basic and practical research in the field of skin biology, dermatology, and cosmetology.

Fistula formation after two staged Aivar Bracka’s repair for hypospadias.

Objectives: To evaluate the frequency of fistula formation after Bracka’s repair for hypospadias. Study Design: Retrospective Case study. Setting: Department of Plastic and Reconstructive Surgery, Shaikh Zayed hospital Lahore and Bahawal Victoria Hospital Bahawalpur. Period: 05 years (2014 -18). Material & Methods: Sixty patients of primary hypospadias underwent two staged Aivar Bracka’s repair. Age of patients, type of hypospadias, presence and extent of chordee, donor skin (preputial, post auricular) used, complications and fistula formation were recorded. Leakge of urine from repair site (Primary/Secondary) was noted as fistula formation after 03 weeks postoperatively. Results: A total of 60 patients underwent two staged Bracka’s repair. There were 32 patients (53.33%) with distal hypospadias, 16 patients (26.67%) with mid penile and 12 Patients (20%) with proximal hypospadias. Chordee was present in 33 Patients (55%). Prepucial skin was used in 46 Patients (76.66%) and post auricular skin in 14 Patients (23.34%) as a donor graft. Four patients (6.66%) developed fistula. 03 patients (5.00%) developed fistula at primary site and 01 patient (1.66%) developed at secondary site. Post auricular skin was used as a donor graft in all 04 patients developing fistula. Conclusion: Two staged Bracka’s repair is a versatile technique and reliable procedure for hypospadias repair with minimal complications and is applicable to all types of hypospadias due to excellent functional and cosmetic outcome.

652 Topical Delivery of Fidgetin-Like 2 siRNA to Enhance Cell Migration for Burn Wound Healing in a Swine Model

Introduction Donor site skin is a precious commodity for burn wounds and graft-sparing procedures are critical for larger total body surface area burns. Currently, large ratio skin grafts have been found to preserve donor skin but at the expense of delayed reepithelialization and increased scar formation at the recipient site. In this study, we propose to utilize a topical gene therapy treatment as an adjunct to higher meshing ratio (4:1) split thickness skin grafts (mSTSG) to improve re-epithelialization and reduce donor skin requirements. Administration of small interfering RNA (siRNA) to suppress the expression of one microtubule regulatory protein, Fidgetin-Like 2 (FL2), has been shown to increase cell migration rate both in vitro and in vivo. We hypothesize that topical application of nanoparticle (NP) encapsulated FL2 siRNA will increase cell migration resulting in expedited reepithelialization in large mesh ratio mSTSG in a full-thickness porcine burn model. A pilot study was conducted to down-select siRNA dose by measuring the rate of reepithelialization of full-thickness grafted burns. Methods Ten full-thickness (FT) 5x5cm burn wounds were created on the dorsum of anesthetized Yorkshire pigs. Four days later, the wounds were surgically debrided to a bleeding wound bed and grafted using a 4:1 mSTSG. Treatments groups (n= 2 per pig) consisted of vehicle control (PBS), scrambled sequence NP siRNA (non-targeting siRNA; negative control), and either 5, 10, or 20uM concentration of NP FL2 siRNA were applied topically to the mSTSG. Rechecks were performed twice a week for four week during which time digital images, non-invasive measurements, punch biopsies were acquired, and treatment was reapplied. Quantitative measurements include rate of reepithelialization and laser speckle imaging. Histopathology was assessed by a blinded pathologist. Results All wounds reepithelialized within 28 days post-grafting without infection. There was a positive trend with increasing concentration of NP FL2 siRNA on the burn wound healing. LSI data showed no statistical differences to vehicle control. Pathology analysis is ongoing. Conclusions Topical application of NP FL2 siRNA did increase the rate of reepithelialization of large ratio mSTSG treated full-thickness injuries. Additional pilot animals are currently ongoing to down-select siRNA dose for future studies comparing the experimental treatment to the current clinical gold standard 1:1.5 mesh ratio. Applicability of Research to Practice A topical gene therapy treatment combined with a higher meshing ratio to improve current treatment modalities by increasing reepithelialization and decreasing multiple donor site harvesting.

56 Reduced Length of Stay with Autologous Skin Cell Suspension Reduces Burn Injuries

Introduction Burn injuries remain a surgical challenge with few recent innovations. Grafting with split-thickness skin grafts (STSGs) has been the standard of care for decades. Although shown to have mortality benefits, STSGs are associated with significant morbidity in the form of pain and additional open wounds. For years, surgeons have looked for ways to decrease this associated morbidity. To that end, autologous skin cell suspension (ASCS) is a recently FDA-approved point of care regenerative medicine technology that reduces donor skin requirements without compromising clinical outcomes. Our study evaluated the cost and length of stay comparing STSG alone versus ASCS. Methods We obtained IRB-approval for single institution, retrospective chart review of patients age >14 years admitted with burn injuries from March 2018 – September 2018. Primary outcome was length of stay/%TBSA for patients undergoing STSG alone as compared to patients undergoing ASCS. The 2016 American Burn Association National Burn Repository (NBR) was used to benchmark LOS/%TBSA. Age, percentage burn injury (TBSA), LOS, mortality, and number of surgeries were reviewed. Student’s t-test was used to assess statistical significance of intragroup analysis. Results 36 patients were treated with ASCS in combination with meshed autografts for full-thickness acute burn injuries. 37 patients were treated with STSGs at our center. Mean age and %TBSA was 45.2 years and 6.6% for the STSG group and 46.0 years and 18.6% for the ASCS group. The LOS/%TBSA for the STSG was 1.72 versus 1.19 for the ASCS patients (p-value=0.02). The NBR predicts a LOS/%TBSA of 3.38 and 3.42 for the STSG and ASCS groups. Patients in the STSG group and ASCS group had statistically similar surgeries and mortalities. Conclusions Burn injured patients treated with ASCS had a decreased LOS/%TBSA when compared to both the STSGs and NBR predictions. ASCS is a novel technology allowing for point-of-care treatment that may decrease LOS for burn injured patients and should be considered as an adjunct to traditional techniques for burn patients. Applicability of Research to Practice Reduced length of stay compared to traditional burn care.