Fat Graft Volume Retention Research Articles

Early activation of macrophage-2 with IL-4 in stromal vascular fraction increases VEGF levels and adipocyte count and maintains volume of fat graft in Wistar rats (Rattus norvegicus)

Several previous studies have demonstrated the benefits of early macrophage 2 activation fat grafts supplemented with macrophage culture. However, this approach is considered impractical in clinical settings because of intraperitoneal induction use. The aim of this study was to investigate the effect of early stromal vascular fraction (SVF) macrophage-2 activation with IL-4 on fat graft survival compared to SVF alone using an animal model for better fat graft viability. This experimental study included inguinal fat harvesting, isolated with collagenases to retrieve the SVF, and then injected with a combination of fat graft and SVF (0.3 mL) into the scalp region. The intervention group received an IL-4 intralesional injection on the third day, and the fat grafts were biopsied on days 7, 14, and 30. The primary outcomes were the final volume of the fat graft, vascular endothelial growth factor (VEGF) expression, and the adipocyte cell count using perilipin staining on immunohistochemistry examination. The group receiving IL-4 exhibited significantly higher VEGF on days 7, 14, and 30 (p=0.009, 0.009, and 0.021, respectively). Similarly, the IL-4 treatment significantly increased the perilipin concentration on days 7, 14, and 30 (p=0.008, 0.008, and 0.029, respectively). In this group, VEGF concentration was significantly increased on day 14 as compared to day 7 (p=0.009), while no significant difference was observed in the control group (p=0.090). Additionally, the IL-4 group displayed significantly less reduction of fat graft volume than the control group, as observed on days 7, 14, and 30 (p=0.009, 0.009, and 0.021, respectively). Overall, the study underscores the potential benefits of early M2 polarization in fat grafting, as well as providing practical advantages for improving fat graft volume retention.

Delayed Supplementation Strategy of Extracellular Vesicles from Adipose-Derived Mesenchymal Stromal Cells with Improved Proregenerative Efficiency in a Fat Transplantation Model.

Background Mesenchymal stromal cells (MSCs) and their secreted extracellular vesicles (MSC-EVs) possess similar proregenerative effects when injected into defects immediately following trauma. However, MSC-EVs are superior to MSCs in terms of storage and rejection reflection, while immediate administration of MSC-EVs is related to several target cells for most donor cells die within few weeks. Besides, the inflammatory cascade is incited, providing an unfavorable environment for target cells. We hypothesized that delayed injection of MSC-EVs might have priority on tissue regeneration than instant injection. Method Extracellular vesicles isolated from adipose-derived mesenchymal stromal cells (ADSC-EVs) were administered into human umbilical vein endothelial cells (HUVECs) in vitro at different doses. The migration of HUVECs was assessed using the scratch wound healing assay, whereas the length of tubes and number of vessel-like structures formed by HUVECs were determined using tube formation assay. Next, 24 BALB/c nude mice were randomly divided into three groups (n = 8). For the EV-delayed group, ADSC-EVs were injected into transplanted fat a week later than the EV-immediate group. The volume and weight of grafts were measured at 3 months after fat transplantation. Further, the number of CD31-possitive vessels and CD206-possitive cells in the fat grafts was quantified. Results Compared with the EV-immediate group, the EV-delayed group had a higher fat tissue retention volume (0.11 ± 0.02 mL versus 0.08 ± 0.01 mL), more neovessels (31.00 ± 4.60 versus 24.20 ± 3.97), and fewer cysts. Furthermore, there were more Ki67-positive cells (25.40 ± 7.14 versus 16.20 ± 4.17) and CD206-positive M2 macrophages cells (23.60 ± 3.44 versus 14.00 ± 3.85) in the EV-delayed group than in the EV-immediate group. Conclusion Delayed injection of ADSC-EVs promotes fat graft volume retention by stimulating angiogenesis. These findings suggest that delayed supplementation might be a more effective strategy for the application of MSC-EVs in tissue regeneration.

109. Vitamin E Treated Fat Grafts Maintain Volume to Improve Grafting Effects on Radiation-induced Fibrosis

Introduction: Fat grafts and vitamin E administration have both previously been described to mitigate radiation-induced skin fibrosis. Fat graft retention continues to hinder its long-term functionality affected mainly by two factors: initial inflammatory response and chronic necrosis secondary to hypovascularity. Thus, we aimed to simultaneously improve fat graft retention along with radiation-induced fibrosis. Methods: Ten adult CD-1 nude male mice at 6-8 weeks of age underwent scalp irradiation with 5 Gy every other day for 12 days, totaling in 30 Gy. Mice recovered for four weeks to allow chronic fibrosis to develop. After recovery, mice received 1000μL of donor human fat graft to the scalp. Mice were separated into 4 conditions: ungrafted irradiated, untreated fat graft, fat graft treated with pentoxifylline (PTX), fat graft treated with vitamin E (VE). Fat graft volume retention was monitored in-vivo using microCT scans with 3-dimensional reconstruction and analysis at weeks 0, 1, 2, 4, 6, and 8 after grafting. Fat graft and overlying skin were harvested and underwent histological and cytokine analysis. Results: Histological analysis demonstrated improvement in radiation-induced fibrosis, CD31+ expression, and 8-isoprostane expression in the vitamin E group. Cytokine analysis revealed decreased cell differentiation markers and inflammatory markers. Fat graft volume retention significantly improved as early as 1 week post-grafting with local vitamin E administration compared to control. Conclusion: Radiation-induced fibrosis and fat graft volume retention are both simultaneously improved with novel, local administration of vitamin E, improving both cosmesis and functionality of fat grafting in chronically irradiated fields.

Fat Grafting With Expanded Adipose-Derived Stromal Cells for Breast Augmentation: A Randomized Controlled Trial.

The main challenge with fat grafting is loss of some of the graft to postsurgery resorption. Previous studies suggest that adipose-derived stromal cells (ASCs) can improve the volume retention of fat grafts but there is a lack of randomized trials to support the use of ASCs in clinical practice. This trial aimed to investigate whether ASCs improve fat graft volume retention in patients undergoing breast augmentation with lipofilling. This was a double-blind, randomized controlled trial of breast augmentation with ASC-enriched fat grafting. Healthy women aged 30 to 45 years were enrolled. First, the participants underwent liposuction to obtain fat for culture expansion of ASCs. Then, the participants were randomly assigned to undergo a 300- to 350-mL breast augmentation with ASC-enriched fat grafting (10 × 106 ASCs/mL fat graft) to 1 of their breasts and placebo-enriched fat grafting of identical volume to the contralateral breast. The primary outcome was fat graft volume retention after a 1-year follow-up measured with MRI. The trial is registered at www.clinicaltrialsregister.eu (EudraCT-2014-000510-59). Ten participants were included in the trial; all completed the treatment and follow-up. No serious adverse events occurred. Fat graft volume retention after 1 year was 54.0% (95% CI, 30.4%-77.6%) in the breasts treated with ASC-enriched fat grafting (n = 10) and 55.9% (95% CI, 28.9%-82.9%) in the contralateral breasts treated with placebo-enriched fat grafting (n = 10) (P = 0.566). The findings of this trial do not support that ASC-enriched fat grafting is superior to standard fat grafting for breast augmentation.

Platelet-Rich Fibrin in Fat Grafts for Facial Lipofilling: A Randomized, Controlled Split-Face Clinical Trial.

ObjectivePrevious studies have reported that platelet-rich fibrin (PRF) may enhance the efficacy of fat grafts in facial lipofilling. However, these studies either lacked objective data or were not randomized, controlled trials. Thus, we aimed to objectively evaluate the efficacy of PRF in facial lipofilling.MethodsA controlled, split-face, randomized trial (January 2018 to May 2019) based on 18 patients who underwent fat grafts for bilateral temple lipofilling was performed. Each patient received a combination of an autologous fat graft and PRF on one side and a fat graft combined with an equal volume of saline on the other side. The effects of PRF were evaluated by comparing the remaining bilateral fat graft volumes through a digital three-dimensional reconstruction technique. Improvements in the appearance and recovery time of each temple were assessed by both a surgeon and patients who were blinded to the treatment assignment. Complications were also recorded.ResultsBilateral temple lipofilling showed no evidence of fat embolism, vascular/nerve injury, infection, massive edema, or prolonged bruising. Three-dimensional reconstruction data and the assessments from both the surgeon and patients revealed no significant differences in fat graft retention volume between the PRF-positive and PRF-negative lipofilling groups. However, recovery time in the PRF-positive lipofilling sites was significantly shortened compared with that of the PRF-negative lipofilling sites.ConclusionFacial filling with autologous fat grafts is effective and safe. Our results show that PRF does not markedly improve fat graft volume retention in the temple but significantly reduces postoperative recovery time.Trial Registration NumberChiCTR2100053663.

Fat Grafts Augmented With Vitamin E Improve Volume Retention and Radiation-Induced Fibrosis.

Treatments for radiation-induced fibrosis range from vitamin E (VE) and pentoxifylline (PTX) systemically to deferoxamine and fat grafting locally. Regarding fat grafting, volume retention hinders its long-term functionality and is affected by 2 factors: inflammation and necrosis secondary to hypovascularity. The authors aimed to simultaneously improve fat graft retention and radiation-induced fibrosis by integrating VE and PTX into fat grafts locally. Forty adult CD-1 nude male mice, 6 weeks old, underwent scalp irradiation and recovered for 4 weeks to allow for development of fibrosis. Mice received 200 μL of donor human fat graft to the scalp. Mice were separated into 4 conditions: no grafting, fat graft without treatment, graft treated with PTX, and graft treated with VE. Fat graft volume retention was monitored in vivo with micro-computed tomography scans at weeks 0, 1, 2, 4, 6, and 8 after grafting. Histological and cytokine analysis of the scalp skin and fat grafts were performed. VE-treated grafts had significant improvement in dermal thickness and collagen density of overlying skin compared with all other groups. VE decreased 8-isoprostane and increased CD31+ staining compared with the other grafted groups. Cytokine analysis revealed decreased inflammatory and increased angiogenic markers in both the fat graft and overlying skin of the VE group. Fat graft volume retention was significantly improved in the VE group starting at 1 week post grafting. Radiation-induced fibrosis and fat graft volume retention are both simultaneously improved with local administration of VE.

QS8: Fat Grafting Depletes Profibrotic Prrx1-positive Fibroblasts In Irradiated Skin And Mitigates Radiation-induced Groin Contracture

Purpose:Contracture is a long-term complication of radiotherapy (RT) that results from pathologic fibrosis of soft tissue in the radiation field. Fat grafting can regenerate fibrotic soft tissue by decreasing collagen density and reorganizing collagen fiber networks. Fibroblasts are the predominant cell involved in extracellular matrix (ECM) synthesis, and dermal fibroblasts possess distinct embryonic origins. We recently identified a profibrotic fibroblast in the mouse ventral dermis marked by embryonic expression of paired related homeobox 1 (Prrx1). Here we sought to investigate how fat grafting alters Prrx1-positive subpopulation distribution as well as functional contractures.Methods:Adult Prrx-1Cre;R26mTmG reporter mice (n=9) underwent whole body lethal irradiation with 9 Gy for hematopoietic depletion. Mice were immediately reconstituted with 2 million nucleated bone-derived cells from donor NSG (NOD.CB17-Prkdcsscid/J) mice via retro-orbital sinus injection. The success of reconstitution and immunodepletion was assessed by fluorescence-activated cell sorting (FACS) analysis of peripheral blood. After 4 weeks, 30 Gy was delivered to the right hindlimb in five fractionated doses to generate limb contracture. The irradiated, contracted limb was then grafted with 200 μl fresh human lipoaspirate and limb extension was measured over the subsequent 8 weeks, at which point skin was harvested for assessment of fibroblast subtypes for FACS and immunofluorescence. A group of mice with radiation-induced groin contracture did not undergo fat grafting and served as the control group.Results:FACS analysis indicated successful immunodepletion and engraftment by 3 weeks post bone marrow transplantation. At one month following groin irradiation, mice had developed significant right hind limb contracture with significantly reduced limb extension (****p≤0.0001). Histologically this was paralleled by thickening of the dermis, and substantial expansion of the fibrogenic Prrx-1-positive fibroblast subpopulation. While human fat graft volume retention was reduced over 8 weeks following implantation, this was associated with significantly improved in limb extension. The skin overlying the grafted fat showed reduced collagen density, as indicated by trichrome staining, as well as a reduction in the fibrogenic Prrx-1-positive fibroblast subpopulation by immunofluorescence imaging, as compared to the control mice.Conclusion:Here we show that fat grafting improves the extensibility of irradiated and contracted hind limbs and reverses radiation-induced skin fibrosis by both reducing the collagen content and by altering the composition of dermal fibroblast subpopulations. Specifically, fat grafting results in a depletion of the Prrx-1-positive fibroblast subpopulation. Further elucidating how this profibrotic fibroblast subpopulation is involved in ventral surface soft tissue fibrosis will facilitate development of novel strategies to treat/prevent debilitating late side-effect of radiotherapy.

New Validated Method for Measuring Fat Graft Retention in the Breast with MRI.

Summary:In this study, we present a new method for measuring fat graft volume retention in the breast based on magnetic resonance imaging scans and a validation study to assess its accuracy and precision. The method was validated by 4 observers using the magnetic resonance imaging scans of 14 patients undergoing breast augmentation with fat grafting. The method was translated into software and was used to measure the change in breast volume from a preoperative scan to a postoperative scan recorded within 3 hours after the surgery, which was compared with the injected fat graft volume. The new method measured the injected fat graft volumes with an average systematic overestimation of 6.3% (SD, 10.5). The median interobserver variation was <7%. We propose that this new method can be a good alternative to previous techniques for clinical research purposes. The software can be made available upon request free of charge for use on the MeVisLab platform.

Abstract 38: Fat Grafting Rescues Radiation-induced Groin Contracture And Results In Diminished Numbers Of Profibrotic Prrx1-positive Dermal Fibroblasts

Purpose: Contracture is a long-term complication of radiotherapy (RT) that results from pathologic fibrosis of soft tissue in the radiation field. Fat grafting can regenerate fibrotic soft tissue by decreasing collagen density and reorganizing collagen fiber networks. Fibroblasts are the predominant cell involved in extracellular matrix (ECM) synthesis, and dermal fibroblasts possess distinct embryonic origins. We recently identified a profibrotic fibroblast in the mouse ventral dermis marked by embryonic expression of paired related homeobox 1 (Prrx1). Here we sought to investigate how fat grafting alters Prrx1-positive subpopulation distribution as well as functional contractures. Methods: Adult Prrx-1Cre;R26mTmG reporter mice (n=9) underwent whole body lethal irradiation with 9 Gy for hematopoietic depletion. Mice were immediately reconstituted with 2 million nucleated bone-derived cells from donor NSG (NOD.CB17-Prkdcsscid/J) mice via retro-orbital sinus injection. The success of reconstitution and immunodepletion was assessed by fluorescence-activated cell sorting (FACS) analysis of peripheral blood. After 4 weeks, 30 Gy was delivered to the right hindlimb in five fractionated doses to generate limb contracture. The irradiated, contracted limb was then grafted with 200 μl fresh human lipoaspirate and limb extension was measured over the subsequent 8 weeks, at which point skin was harvested for assessment of fibroblast subtypes for FACS and immunofluorescence. A group of mice with radiation-induced groin contracture did not undergo fat grafting and served as the control group. Results: FACS analysis indicated successful immunodepletion and engraftment by 3 weeks post bone marrow transplantation. At one month following groin irradiation, mice had developed significant right hind limb contracture with significantly reduced limb extension (****p≤0.0001). Histologically this was paralleled by thickening of the dermis, and substantial expansion of the fibrogenic Prrx-1-positive fibroblast subpopulation. While human fat graft volume retention was reduced over 8 weeks following implantation, this was associated with significantly improved in limb extension. The skin overlying the grafted fat showed reduced collagen density, as indicated by trichrome staining, as well as a reduction in the fibrogenic Prrx-1-positive fibroblast subpopulation by immunofluorescence imaging, as compared to the control mice. Conclusion: Here we show that fat grafting improves the extensibility of irradiated and contracted hind limbs and reverses radiation-induced skin fibrosis by both reducing the collagen content and by altering the composition of dermal fibroblast subpopulations. Specifically, fat grafting results in a depletion of the Prrx-1-positive fibroblast subpopulation. Further elucidating how this profibrotic fibroblast subpopulation is involved in ventral surface soft tissue fibrosis will facilitate development of novel strategies to treat/prevent debilitating late side-effect of radiotherapy.

Increasing Fat Graft Retention in Irradiated Tissue after Preconditioning with External Volume Expansion.

Fat grafting is an adjuvant that may improve the quality of radiation-damaged tissue. However, fat grafting for volume restoration in irradiated sites may be less effective because of a poorly vascularized fibrotic recipient bed. External volume expansion has emerged as a potential technique to prepare the recipient sites for improved survival of grafted fat. The authors previously demonstrated increased vasculature with external volume expansion stimulation of irradiated tissues. The authors now hypothesize that external volume expansion's improvements in recipient-site vascularity will increase the volume retention and quality of fat grafts in fibrotic irradiated sites. Athymic mice were irradiated until development of chronic radiation injury. Then, the irradiated site was stimulated by external volume expansion (external volume expansion group), followed by subcutaneous fat grafting. Grafts in an irradiated site without external volume expansion stimulation (irradiated control group) and grafts in a healthy nonirradiated (nonirradiated control group) site were used as controls. All grafts were monitored for 8 weeks and evaluated both histologically and by micro-computed tomography for analysis of volume retention. Hyperspectral imaging confirmed a 25 percent decrease in vascularity of irradiated tissue (irradiated control group) compared with nonirradiated tissue (nonirradiated control group). Grafts in the irradiated control group retained 11 percent less volume than grafts in the nonirradiated control group. The experimental external volume expansion group achieved a 20 percent (p = 0.01) increase in retained graft volume compared with the irradiated control group. External volume expansion stimulation can mitigate the effects of irradiation at the recipient site and in turn help preserve fat graft volume retention. Possible mechanisms include increased vascularity, adipogenic conversion, and increased compliance of a fibrotic recipient site.

Recipient-Site Preconditioning with Deferoxamine Increases Fat Graft Survival by Inducing VEGF and Neovascularization in a Rat Model.

The authors hypothesize that ischemic preconditioning of the recipient site with deferoxamine will increase fat graft survival by enhancing angiogenesis in a rat model. Cell viability, tube formation, and mRNA expression were measured in human umbilical vein endothelial cells treated with deferoxamine. A total of 36 rats were then used for an in vivo study. A dose of 100 mg/kg of deferoxamine was injected subcutaneously into the rat scalp every other day for five treatments. On the day after the final injection, the scalp skin was harvested from half the animals to evaluate the effects of deferoxamine on the recipient site. In the remaining animals, inguinal fat tissue was transplanted to the scalp. Eight weeks after transplantation, the grafts were harvested to evaluate the effects of deferoxamine preconditioning on fat graft survival. In human umbilical vein endothelial cells, treatment with a deferoxamine concentration higher than 400 μM decreased cell viability compared with the control (p = 0.002). Treatment with 100 and 200 μM deferoxamine increased endothelial tube formation (p = 0.001) and mRNA levels of angiogenesis-related factors (p = 0.02). Rat scalps treated with deferoxamine exhibited increased capillary neoformation (p = 0.001) and vascular endothelial growth factor protein expression (p = 0.024) compared with controls. Fat graft volume retention, capillary density (p < 0.001), and adipocyte viability (p < 0.001) in the grafted fat increased when the recipient site was preconditioned with deferoxamine. This study demonstrated that recipient site preconditioning with deferoxamine increases fat graft survival by inducing vascular endothelial growth factor and neovascularization.

The current gold standard breast volumetry technique seems to overestimate fat graft volume retention in the breast: A validation study

MRI is generally considered as the gold standard for measuring breast volume because of its high accuracy of the modality. Many techniques used to measure total breast volume have been validated, but none of these techniques have been validated for their ability to measure the volume retention of fat grafts in the breast. In this study, the authors investigated the accuracy of the most common MRI technique used to measure fat graft retention in the breast by measuring the volume changes after breast augmentation. Patients undergoing breast augmentation with either breast implants or fat grafting underwent MRI scans before and after surgery. Blinded observers measured the change in breast volume from the MRI scans. The difference between the measured change in breast volume and the volume of the breast augmentation was used to determine the accuracy of the MRI technique. Twenty-eight patients with a total of 56 breasts were included. In total, 168 measurements of change in breast volume were performed by the observers. The MRI measurements of change in breast volume overestimated the true volumes of the breast augmentations by an average of 50.8%, and only 8 of the 168 individual measurements had measurement errors below 50 mL. The MRI technique, which is considered as the gold standard for the quantification of fat graft volume retention, was associated with a significant measurement error. These findings have potential implications for the interpretation of previously published results of studies based on this technique.

Fat Grafting into Younger Recipients Improves Volume Retention in an Animal Model.

Soft-tissue deficits associated with various craniofacial anomalies can be addressed by fat grafting, although outcomes remain unpredictable. Furthermore, consensus does not exist for timing of these procedures. Whereas some advocate approaching soft-tissue reconstruction after the underlying skeletal foundation has been corrected, other studies have suggested that earlier grafting may exploit a younger recipient niche that is more conducive to fat graft survival. As there is a dearth of research investigating effects of recipient age on fat graft volume retention, this study compared the effectiveness of fat grafting in younger versus older animals through a longitudinal, in vivo analysis. Human lipoaspirate from three healthy female donors was grafted subcutaneously over the calvaria of immunocompromised mice. Volume retention over 8 weeks was evaluated using micro-computed tomography at three experimental ages: 3 weeks, 6 months, and 1 year. Histologic examination was performed on explanted grafts to evaluate graft health and vascularity. Recipient-site vascularity was also evaluated by confocal microscopy. The greatest retention of fat graft volume was noted in the youngest group compared with both older groups (p < 0.05) at 6 and 8 weeks after grafting. Histologic and immunohistochemical analyses revealed that improved retention in younger groups was associated with greater fat graft integrity and more robust vascularization. The authors' study provides evidence that grafting fat into a younger recipient site correlates with improved volume retention over time, suggesting that beginning soft-tissue reconstruction with fat grafting in patients at an earlier age may be preferable to late correction.

Deferoxamine Preconditioning of Irradiated Tissue Improves Perfusion and Fat Graft Retention.

Radiation therapy is a mainstay in the treatment of many malignancies, but collateral damage to surrounding tissue, with resultant hypovascularity, fibrosis, and atrophy, can be difficult to reconstruct. Fat grafting has been shown to improve the quality of irradiated skin, but volume retention of the graft is significantly decreased. Deferoxamine is a U.S. Food and Drug Administration-approved iron-chelating medication for acute iron intoxication and chronic iron overload that has also been shown to increase angiogenesis. The present study evaluates the effects of deferoxamine treatment on irradiated skin and subsequent fat graft volume retention. Mice underwent irradiation to the scalp followed by treatment with deferoxamine or saline and perfusion and were analyzed using laser Doppler analysis. Human fat grafts were then placed beneath the scalp and retention was also followed up to 8 weeks radiographically. Finally, histologic evaluation of overlying skin was performed to evaluate the effects of deferoxamine preconditioning. Treatment with deferoxamine resulted in significantly increased perfusion, as demonstrated by laser Doppler analysis and CD31 immunofluorescent staining (p < 0.05). Increased dermal thickness and collagen content secondary to irradiation, however, were not affected by deferoxamine (p > 0.05). Importantly, fat graft volume retention was significantly increased when the irradiated recipient site was preconditioned with deferoxamine (p < 0.05). The authors' results demonstrated increased perfusion with deferoxamine treatment, which was also associated with improved fat graft volume retention. Preconditioning with deferoxamine may thus enhance fat graft outcomes for soft-tissue reconstruction following radiation therapy.

Melatonin promotes survival of nonvascularized fat grafts and enhances the viability and migration of human adipose-derived stem cells via down-regulation of acute inflammatory cytokines.

Nonvascularized fat grafting is a valuable technique for soft tissue reconstruction but poor survival of fat in the host environment remains a problem. A process known as cell-assisted transfer is used to enhance fat graft retention by adding stromal vascular fraction, an adipose-derived stem cell (ASC) rich content to lipoaspirate. We have recently shown that the use of melatonin, a reactive oxygen species scavenger, protects human ASCs from hydrogen peroxide-induced oxidative stress and cell death in vitro but its role as a pharmacological adjunct in clinical fat grafting has not been studied. Herein, the effect of melatonin was examined on human ASCs in vitro using survival and functional assays including the MTT assay, CellTox Green assay, monolayer scratch assay as well as a human cytokine chemoluminescence, and tumour necrosis factor-α assay. Further, the effect of melatonin-treated fat grafts was tested in vivo with a murine model. Haematoxylin and eosin staining, perilipin and CD31 immunostaining were performed with morphometric analysis of adipose tissue. The results demonstrate that, in vitro, the addition of melatonin to ASCs significantly improved their cell-viability, promoted cell migration and preserved membrane integrity as compared to controls. In addition, it induced a potent anti-inflammatory response by downregulating acute inflammatory cytokines particularly tumour necrosis factor-α. For the first time, it is demonstrated in vivo that melatonin enhances fat graft volume retention by reducing inflammation and increasing the percentage of adipose volume within fat grafts with comparable volumes to that of cell-assisted lipotransfer. Based on these novel findings, melatonin may be a useful pharmacological adjunct in clinical fat grafting.

Abstract 68: Deferoxamine Preconditioning of Irradiated Tissue Increases Fat Graft Volume Retention

PURPOSE: Hypovascularity due to irradiation therapy causes the skin to become fibrotic and causes soft tissue to atrophy. Fat grafting has been shown to improve the quality of irradiated skin, but volume retention of the graft is significantly decreased. Deferoxamine is an FDA-approved iron-chelating medication, that has been shown to increase angiogenesis. Preconditioning of irradiated sites with this compound may reduce radiation induced ischemia and enhance fat graft survival. METHODS: Immunocompromised nude-mice underwent external beam irradiation of the scalp. Five weeks later, mice either received seven deferoxamine treatments (1mg in 100ul) or saline subcutaneously to the irradiated area every other day. Laser Doppler analysis (LDA) was recorded prior to irradiation, following irradiation, and 24 hours following each treatment. Human fat grafts were then injected in the subcutaneous plane of the scalp and volume retention measured by CT scan over 8 weeks. Finally, skin and fat samples were evaluated histologically for vasculature, dermal thickness, and fat graft quality. RESULTS: After 4 treatments with deferoxamine, a significant increase in microvasculature was observed using LDA.There was also significance with the development of microvasculature in the fat graft with LDA. Using microCT, we observed a significant increase in fat graft volume retention with the deferoxamine treated group compared to the saline treated group, and this was paralleled by improved histologic staining of skin and fat grafts. CONCLUSION: Our results show increased microvasculature and increased fat graft volume retention with deferoxamine treatment. Deferoxamine treatment may also promote beneficial effects in dermal thickness and in quality scoring of the fat grafts, thus leading to a potential clinical application in radiation damaged soft tissue.

Cell-Assisted Lipotransfer Improves Volume Retention in Irradiated Recipient Sites and Rescues Radiation-Induced Skin Changes.

Radiation therapy is not only a mainstay in the treatment of many malignancies but also results in collateral obliteration of microvasculature and dermal/subcutaneous fibrosis. Soft tissue reconstruction of hypovascular, irradiated recipient sites through fat grafting remains challenging; however, a coincident improvement in surrounding skin quality has been noted. Cell-assisted lipotransfer (CAL), the enrichment of fat with additional adipose-derived stem cells (ASCs) from the stromal vascular fraction, has been shown to improve fat volume retention, and enhanced outcomes may also be achieved with CAL at irradiated sites. Supplementing fat grafts with additional ASCs may also augment the regenerative effect on radiation-damaged skin. In this study, we demonstrate the ability for CAL to enhance fat graft volume retention when placed beneath the irradiated scalps of immunocompromised mice. Histologic metrics of fat graft survival were also appreciated, with improved structural qualities and vascularity. Finally, rehabilitation of radiation-induced soft tissue changes were also noted, as enhanced amelioration of dermal thickness, collagen content, skin vascularity, and biomechanical measures were all observed with CAL compared to unsupplemented fat grafts. Supplementation of fat grafts with ASCs therefore shows promise for reconstruction of complex soft tissue defects following adjuvant radiotherapy.

Studies in fat grafting: Part IV. Adipose-derived stromal cell gene expression in cell-assisted lipotransfer.

Fat graft volume retention remains highly unpredictable, but addition of adipose-derived stromal cells to fat grafts has been shown to improve retention. The present study aimed to investigate the mechanisms involved in adipose-derived stromal cell enhancement of fat grafting. Adipose-derived stromal cells isolated from human lipoaspirate were labeled with green fluorescent protein and luciferase. Fat grafts enhanced with adipose-derived stromal cells were injected into the scalp and bioluminescent imaging was performed to follow retention of adipose-derived stromal cells within the fat graft. Fat grafts were also explanted at days 1, 5, and 10 after grafting for adipose-derived stromal cell extraction and single-cell gene analysis. Finally, CD31 immunohistochemical staining was performed on fat grafts enriched with adipose-derived stromal cells. Bioluminescent imaging demonstrated significant reduction in luciferase-positive adipose-derived stromal cells within fat grafts at 5 days after grafting. A similar reduction in viable green fluorescent protein-positive adipose-derived stromal cells retrieved from explanted grafts was also noted. Single-cell analysis revealed expression of multiple genes/markers related to cell survival and angiogenesis, including BMPR2, CD90, CD105, FGF2, CD248, TGFß1, and VEGFA. Genes involved in adipogenesis were not expressed by adipose-derived stromal cells. Finally, CD31 staining revealed significantly higher vascular density in fat grafts explanted at day 10 after grafting. Although adipose-derived stromal cell survival in the hypoxic graft environment decreases significantly over time, these cells provide multiple angiogenic growth factors. Therefore, improved fat graft volume retention with adipose-derived stromal cell enrichment may be attributable to improved graft vascularization.