Melinda Costa

Live imaging of Smad2/3 signaling in mouse skin wound healing

Biophotonics and real‐time imaging are novel technologies that can greatly enhance the study of complex biological processes. We applied this technology in a transgenic mouse with a luciferase reporter gene fused to a transforming growth factor‐β (TGF‐β) responsive Smad2/3‐binding element to study bioluminescence after skin wounding. Two dorsal midline excisional skin wounds were made using a biopsy punch. One wound was randomized to suture closure and the other allowed to heal by secondary intention (n=8 each wound). Bioluminescence was measured at fixed time points following surgery. Phospho‐Smad2/3 immunohistochemistry was performed to localize expression in skin wound samples. In vivo bioluminescence increased following skin wounding. Peak activity occurred on day 17 and was fourfold that of baseline (p<0.05). Subgroup analysis of primary and secondary healing showed that primarily sutured wounds had peak activities earlier than those with secondary healing, although this did not reach statistical significance. Intense phospho‐Smad2/3 staining was found in the hair follicles. In vivo bioluminescence tracks Smad2/3‐dependent TGF‐β signaling in the in vivo wound healing process. Our findings suggest that signaling increases after wound healing, which contrasts with other studies that show raised TGF‐β signaling in the initial days following wounding.

In Vitro Analysis of Transforming Growth Factor-β1 Inhibition in Novel Transgenic Sbe-luciferase Mice

Background: Transforming growth factor β1 (TGF-β1) expression correlates with scarring. A novel transgenic mouse model with a Smad2/3-responsive luciferase reporter construct (SBE-luc) has been developed. We hypothesized that bioluminescence in SBE-luc dermal fibroblasts could be measured to assess TGF-β1 inhibition. Materials and Methods: Cultured dermal fibroblasts from SBE-luc mice were treated simultaneously with TGF-β1 and increasing doses of either neutralizing antibody to TGF-β (NA-TGFβ) or SB-431542, a novel TGF-β receptor kinase inhibitor. Fibroblasts were measured for luciferase activity. SBE-luc fibroblasts underwent Western blot analysis for collagen type I production. Results: TGF-β1 produced maximal luciferase activity in SBE-luc fibroblasts at 0.1 ng/mL (P < 0.05). NA-TGFβ and SB-431542 inhibited luciferase activity in a dose-dependent fashion, with complete inhibition achieved by 0.1 μg/mL and 1 μM, respectively (P < 0.05). NA-TGFβ and SB-431542 inhibited collagen type I production. Conclusions: Our in vitro results provide validation for further in vivo real-time imaging studies using the SBE-luc mouse as a novel wound-healing model.

Spring-assisted cranial vault expansion in the setting of multisutural craniosynostosis and anomalous venous drainage: case report

Patients with multisutural craniosynostosis can develop anomalous venous connections between the intracranial sinuses and cutaneous venous system through enlarged emissary veins. Cranial vault remodeling in this subset of patients carries the risk of massive intraoperative blood loss and/or occlusion of collateral draining veins leading to intracranial venous hypertension and raised intracranial pressure, increasing the morbidity of cranial expansion. The authors report the use of spring-mediated expansion as a technique for cranial reconstruction in which the collateral intracranial venous drainage system can be preserved. A patient with bilateral lambdoid, sagittal, and unicoronal synostosis presented for cranial reconstruction. A tracheostomy and ventriculoperitoneal shunt were placed prior to intervention. At the time of reconstruction, a Luckenschadel skull abnormality and Chiari malformation Type I were present. A preoperative CT venogram demonstrated large collateral superficial occipital veins, small bilateral internal jugular veins, and hypoplastic jugular foramina. Collateral flow from the transverse and sigmoid sinuses through large occipital emissary veins was seen. Spring-mediated cranial vault expansion was performed with care to preserve the large collateral veins at the occipital midline. Four springs were placed at each lambdoid and the posterior and anterior sagittal sutures following 1-cm strip suturectomies. Removal of the springs was performed 2 months postoperatively. Cranial vault expansion was performed without disturbing the aberrant intracranial/extracranial venous collateral system. Estimated blood loss was 150 ml. A CT scan obtained 3 months postoperatively showed resolution of the Luckenschadel deformity and a 40% volumetric increase in the skull compared with the preoperative CT. Patients with anomalous venous drainage patterns and multisutural synostosis can undergo spring-mediated cranial vault expansion while preserving the major emissary veins draining the intracranial sinuses. Risks of blood loss, intracranial venous hypertension, and increased intracranial pressure may be decreased compared with traditional techniques of repair.

Successful Neonatal Mandibular Distraction Osteogenesis in Patients with Concomitant Laryngomalacia

8 CONCLUSIONS: Mandibular distraction is an effi cacious treatment modality for neonates with PRS, and should be considered the fi rst line intervention to avoid tracheostomy. Patient variables including birth weight, syndromic status, neurologic impairment and multi-level airway obstruction do not preclude the utilization of MD; however, predicting the future necessity of tracheostomy and the ability to decannulate following MD of these patients is less reliable.

Incorporation of Fresh Tissue Surgical Simulation into Plastic Surgery Education: 5-Year Evolution of an Essential Educational Experience

Text: Background: As interest in surgical simulation gains momentum, plastic surgical educators are pressed to provide realistic surgical experience outside of the operating suite. High fidelity models of plastic surgery procedures are still incomparable to dissection of fresh tissue. We have evolved a fresh tissue dissection and simulation program with emphasis on surgical technique and simulation of clinical surgery. We hypothesized that resident education, confidence, and ultimately quality of surgical performance could be improved by adding fresh tissue dissection and simulation to our resident educational curriculum. Methods: Over a 5 year period Fresh Tissue Dissection (FTD) using fresh cadavers was incorporated into resident curricula. Participants performed biweekly dissections and simulations of procedures performed on each rotation with structured emphasis on anatomical detail, surgical technique and maneuvers, and rehearsal of operative sequence, according to skill level and clinical need. Surgical simulation was performed with standard fresh tissue dissection. Vascular perfusion was also added to simulation to maximize realistic operative simulation. Resident performance was evaluated using retrospective pre and post tests in addition to a web based survey that evaluated resident perception of technique. All answers were ranked according to a 5 point scale: 1-not confident to 5very confident. Results: The curriculum evolved, became increasingly relevant, and ultimately was incorporated directly into most resident rotations. A total of 79 dissection days occurred, and a total of 199 procedures were reported, including 81 anatomical dissections, 54 direct surgical simulations, and 26 preoperative surgical rehearsals. Data regarding confidence pre and post participation were gathered for 189 procedures and for 26 surgical rehearsals. Overall, Resident predissection confidence was 1.93±1.04 and post dissection was 4.21±0.93 (p<.01). Survey data demonstrated that participants believed that FTD improved their operative abilities. All residents felt that their technique (4.6±.75), speed (4.3±.56), safety (4.57±0.5) and knowledge of anatomy (4.7±0.5) improved. Conclusions: Given adequate access to fresh tissue, a plastic surgery experience with FTD can improve resident confidence and perception of performance. High fidelity models were created that simulated live surgery. We believe FTD provides adequate extraclinical experience to improve the abilities of our residents. While initial data suggests that operative performance is subjectively improved, additional evaluation is needed to establish objective evidence that patient outcomes and surgical quality can be improved.

81: Continuous Live Imaging of TGF-Beta Activity Using a Novel Transgenic Mouse Wound-Healing Model

iNTRODUCTiON: Scar formation is a fundamental plastic surgery problem. The pathogenesis of scarring is unclear, and the treatment options are time-consuming, expensive, and consistently unsuccessful ( , 2). Increased transforming growth factor beta (TGF-Beta) expression correlates with scarring and excess collagen deposition (3, 4, 5); however, the spatial and temporal expression patterns of TGF-Beta during normal and pathological wound repair remain unresolved. We used a novel transgenic mouse system that allows real-time continuous in vivo tracking of TGF-Beta1 activity by measuring bioluminescence after skin wounding.