Orlando Canizares

Human fat grafting alleviates radiation skin damage in a murine model.

Background: Autogenous fat grafting has been observed to alleviate the sequelae of chronic radiodermatitis. To date, no study has replicated this finding in an animal model. Methods: The dorsa of adult wild-type FVB mice were shaved and depilated. The dorsal skin was then distracted away from the body and irradiated (45 Gy). Four weeks after irradiation, 1.5-cc fat or sham grafts were placed in the dorsal subcutaneous space. Gross results were analyzed photometrically. The animals were euthanized at 4 and 8 weeks after fat or sham grafting and their dorsal skin was processed for histologic analysis. Results: Hyperpigmentation and ulceration were grossly improved in fat-grafted mice compared with sham-grafted controls. This improvement manifested histologically in a number of ways. For example, epidermal thickness measurements demonstrated decreased thickness in fat-grafted animals at both time points (20.6 ± 1.5 &mgr;m versus 55.2 ± 5.6 &mgr;m, p = 0.004; 17.6 ± 1.1 &mgr;m versus 36.3 ± 6.1 &mgr;m, p = 0.039). Picrosirius red staining demonstrated a diminished scar index in fat-treated animals at both time points as well (0.54 ± 0.05 versus 0.74 ± 0.07, p = 0.034; and 0.55 ± 0.06 versus 0.93 ± 0.07, p = 0.001). Conclusion: Fat grafting attenuates inflammation in acute radiodermatitis and slows the progression of fibrosis in chronic radiodermatitis.

Grading lipoaspirate: is there an optimal density for fat grafting?

Background: Clinical results of fat grafting have been unpredictable. In this article, the authors hypothesize that centrifugation creates “graded densities” of fat with varying characteristics that influence lipoaspirate persistence and quality. Methods: Aliquots of human female lipoaspirate (10 cc) were centrifuged for 3 minutes at 1200 g. The bloody and oil fractions were discarded. Subsequently, 1.0 cc of the highest density and lowest density fat was separated for lipoinfiltration or analysis. Highest density or lowest density fat grafted into adult FVB mice was harvested at 2 and 10 weeks to quantify short- and long-term persistence, respectively. Progenitor cell number and expression of vascular endothelial growth factor, stromal cell–derived factor-1&agr;, platelet-derived growth factor, and adiponectin were analyzed by flow cytometry and enzyme-linked immunosorbent assay, respectively. Results: Greater percentages of highest density fat grafts remain at 2 and 10 weeks after injection compared with lowest density fat grafts (85.4 ± 1.9 percent versus 62.3 ± 0.1 percent, p = 0.05; and 60.8 ± 4.9 versus 42.2 ± 3.9, p < 0.05, respectively). Highest density fractions contain more progenitor cells per gram than lowest density fractions (2.0 ± 0.2-fold increase, p < 0.01). Furthermore, concentrations of vascular endothelial growth factor, stromal vascular fraction, platelet-derived growth factor, and adiponectin are all elevated in highest density compared with lowest density fractions (34.4 percent, p < 0.01; 34.6 percent, p < 0.05; 52.2 percent, p < 0.01; and 45.7 percent, p < 0.05, respectively). Conclusions: Greater percentages of highest density fractions of lipoaspirate persist over time compared with lowest density fractions. A vasculogenic mechanism appears to contribute significantly, as highest density fractions contain more progenitor cells and increased concentrations of several vasculogenic mediators than lowest density fractions.

Flow perfusion maintains ex vivo bone viability: a novel model for bone biology research

Encased in lacunae, osteocytes receive nutrition and biomechanical signals through the lacunocanalicular system. We have developed a novel flow‐perfusion bioreactor designed to support lacunocanalicular fluid flow. We hypothesize that ex vivo fluid flow can maintain endochondral bone viability and, ultimately, serve as a novel model to study bone biology in vitro. Sprague–Dawley rat femurs were harvested, stripped of soft tissue, loaded into a custom‐designed bioreactor and perfused with osteogenic culture medium. After 14 days of flow‐perfusion or static culture, the bones were harvested, fixed, decalcified, embedded, sectioned and stained with haematoxylin and eosin. Fresh long bone samples were similarly processed for comparison. Osteocyte viability and function were also evaluated, using thiazolyl blue tetrazolium bromide (MTT), fluorospectrophotometric DNA quantification, alkaline phosphatase (ALP) colorimetric assay and fluorochrome labelling of mineralizing surfaces. All samples remained free of infection throughout the study period. After 14 days of flow perfusion, histological analysis showed normal‐appearing bony architecture, with 72% of lacunae being osteocyte‐filled compared with 93% in freshly harvested samples and only 36% in static samples. MTT staining and assay confirmed osteocyte viability in the flow‐perfusion samples as well as in fresh samples. DNA quantification demonstrated DNA to be preserved in flow‐perfused samples when compared with freshly harvested samples. ALP activity in flow‐perfusion explants was upregulated compared with fresh and static samples. Fluorochrome‐labelled mineralizing surfaces were seen throughout the explanted flow‐perfused samples. This is the first demonstration that flow perfusion provides adequate chemotransportation to explanted murine endochondal bones. Copyright

Augmenting neovascularization accelerates distraction osteogenesis.

Background: Distraction osteogenesis has revolutionized the treatment of craniofacial deformities, but it is limited by lengthy consolidation periods and tenuous healing in certain clinical settings, such as irradiated tissue. In this study, the authors aim to investigate whether increasing neovascularization by progenitor cell mobilization accelerates bone formation during distraction. Methods: Sprague-Dawley rats aged 8 weeks (n = 36) were subjected to unilateral mandibular distraction with 3-day latency, 7-day activation (0.25 mm twice daily), and 21-day consolidation periods. From the beginning of the consolidation period, animals received daily injections of either AMD3100 (bone marrow progenitor cell mobilizing agent) or sterile saline. Animals were euthanized on postoperative day 31; mandibles were harvested; and bone regeneration was assessed using micro–computed tomography, immunohistochemistry, bone morphogenetic protein-2 enzyme-linked immunosorbent assay, and mechanical testing. Results: Immunohistochemistry demonstrated that AMD3100 treatment increased vascular density and bone formation. Micro–computed tomography and dual-emission x-ray absorptiometry demonstrated that AMD3100-treated animals had improved bone generation compared with sham-treated controls. Greater force was required on three-point testing to break AMD3100-treated bone. Bone morphogenetic protein-2 expression was up-regulated with AMD3100. Interestingly, the nondistracted contralateral hemimandibles treated with AMD3100 were also stronger than sham-treated counterparts. Conclusions: Progenitor cell mobilization improves bone regeneration in a rat distraction model. Furthermore, because this effect is seen in healthy bone and in ischemic bone healing during distraction, the mechanism is not merely related to oxygenation, but could be a phenomenon of fluid flow.

Optimizing Efficiency in Deep Inferior Epigastric Perforator Flap Breast Reconstruction.

BackgroundThe process of harvesting and performing microsurgical anastomosis in a deep inferior epigastric perforator (DIEP) flap for breast reconstruction can be a lengthy procedure, which could affect outcomes and patient safety. We hypothesize that the implementation of a high volume center, preoperative planning, and the adaptation of key intraoperative components will optimize the efficiency of perforator flap surgery for breast reconstruction. MethodsA retrospective review of 68 consecutive patients who underwent 104 DIEP flaps for immediate or delayed breast reconstruction was performed. Standardized preoperative planning, including computed tomography/magnetic resonance imaging angiogram, operating room setup, and operative technique, was followed. The times of flap harvest, internal mammary vessel harvest, microsurgical anastomosis, flap inset, wound closure, and total length of procedure were reviewed as well as patient outcomes. ResultsThe average length of surgery for a unilateral DIEP was 3 hours and 21 minutes and for a bilateral DIEP was 5 hours and 46 minutes. In bilateral DIEP flaps, a significantly longer operative time was noted in immediate (363 ± 14 minutes) compared to delayed (296 ± 17 minutes) (P < 0.05) reconstruction and between procedures performed by 1 surgeon (400 ± 29 minutes) versus 2 surgeons (326 ± 11 minutes) (P < 0.05). Interestingly, no significant difference in operative time was seen in DIEP flaps performed on patients with a body mass index (BMI) less than 30 (193 ± 7.6, 352 ± 17 minutes) versus a BMI greater than 30 (213 ± 11, 333 ± 14 minutes) in both unilateral and bilateral procedures, respectively. ConclusionsEfficiency is optimized by preoperative planning with computed tomography/magnetic resonance imaging angiogram, a dedicated operating room team, including 2 microsurgeons and a systematic approach for surgery. The BMI may not significantly affect the duration of surgery.

Expanding the Applications of the Profunda Artery Perforator Flap.

Background: The profunda artery perforator free flap has not gained traction for nonbreast reconstruction, likely because of the presence of a proven workhorse in the anterolateral thigh flap. The authors believe that the profunda artery perforator flap offers similar coverage characteristics with the benefits of a medial donor site, a more consistent anatomy, and relatively easy dissection. The authors review their indications, technique, and outcomes in seven patients requiring eight free flap reconstructions. Methods: The authors applied the use of the vertically oriented profunda artery perforator flap to both lower extremity and head and neck reconstructions in which an anterolateral thigh flap would normally have been used. Details reviewed include soft-tissue defect, perforator location, flap size, recipient vessel, and complications. Results: Eight soft-tissue defects were covered with a vertically oriented profunda artery perforator flap in seven patients. Six reconstructions were for distal lower extremity and two were for head and neck reconstruction, both trauma and oncologic reconstructions. Flap sizes ranged from 40 to 92 cm2. The pedicle length ranged from 7 to 10 cm. There were no partial or complete flap losses. One complication of seroma at the donor site requiring washout and closure was encountered. Conclusions: The profunda artery perforator flap is a safe and effective option for perforator-based free flap reconstruction with relative ease of harvest and an inconspicuous donor site. This flap offers an excellent alternative to the anterolateral thigh flap. In certain patient demographics, the profunda artery perforator flap should be considered as a primary option. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, V.

Shaken Not Stirred? The Effect of Processing Techniques on Fat Graft Survival

36 CONCLUSION: Although HD fractions of centrifuged lipoaspirate contain fewer functional adipocytes than LD fractions, they survive to a greater extent following grafting. The mechanism responsible may be vasculogenic in nature, as baseline analysis found HD fractions to contain more progenitor cells/g as well as increased concentrations of several vasculogenic mediators (VEGF, SDF) compared to LD fractions. This study is the first of its kind, identifying graded densities of lipoaspirate with unique characteristics that directly influence fat graft survival. Shaken Not Stirred? The Effect of Processing Techniques on Fat Graft Survival

1: LACUNOCANALICULAR FLOW AND DISTRACTION OSTEOGENESIS: THEORY AND THERAPEUTICS

Introduction: Our hypothesis is that the tension stress of activation increases lacunocanalicular flow and upregulates the mechanotransductive osteogenic pathway. Furthermore, we hypothesize improving vascularization by endothelial progenitor cell (EPC) mobilization enhances lacunocanalicular flow in the consolidation period, maintaining upregulation of the mechanotransductive pathway to improve and accelerate osteogenesis.

112B: CHEWING THE FAT: THE IMPORTANCE OF PROCESSING TECHNIQUES ON FAT GRAFT SURVIVAL

Background: Lipoaspirate processing methods including centrifugation and Telfa®-rolling have been advocated without direct comparison of their efficacy. Adipocytes secrete proteins that are active participants in regulating angiogenesis and apoptosis. We hypothesize that processing adipose tissue on Telfa gauze creates a lipoaspirate with a more functional adipokine profile and comparable fat graft survival to centrifugation.

The Effects of Graded Densities of Lipoaspirate on Fat Graft Survival

106 CONCLUSION: This series of studies represents a major advance in the development of a practical, deployable, integrated, low-impedance, high fidelity neural interface. Nonmetallic, biosynthetic hybrid PNS interfaces both sense and stimulate physiologically relevant efferent and afferent action potentials in vivo, are biocompatible and electrically relevant. Current work is addressing signal delivery to electronics and scaling of the device for multiple input and output channels.