Brown Adipose Tissue Promotes Autologous Fat Grafts Retention Possibly Through Inhibiting Wnt/ß-Catenin Pathway.

BACKGROUND: In plastic surgery, autologous fat grafts (AFG) play an important role because of their abundant supply, biocompatibility, and low rejection rate. However, the lower retention rate of fat grafts limits their widespread use. Brown adipose tissue (BAT) can promote angiogenesis and regulate the level of associated inflammation. This study explored whether BAT has a facilitative effect on fat graft retention. METHODS: We obtained white adipose tissue (WAT) from c57 mice and combined it with either BAT from c57 mice or phosphate-buffered saline (PBS) as a control. These mixtures were injected subcutaneously into the back of thymus-free nude mice. After 12 weeks, fat grafts were harvested, weighed, and analyzed. RESULTS: We found that the BAT-grafted group had higher mass retention, more mature adipocytes, and higher vascularity than the other group. Further analysis revealed that BAT inhibited M1 macrophages; down-regulated IL-6, IL-1ß, and TNF-ß; upregulated M2 macrophages and Vascular endothelial growth factor-A (VEGFA); and promoted adipocyte regeneration by inhibiting the Wnt/ß-catenin pathway, which together promoted adipose graft retention. CONCLUSION: The study demonstrated that BAT improved adipose graft retention by promoting angiogenesis, inhibiting tissue inflammation levels and the Wnt/ß-catenin pathway. LEVEL OF EVIDENCE III: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266.

Evidence of Browning of White Adipocytes in Poorly Survived Fat Grafts in Patients.

BACKGROUND: Browning adipocytes induced by burn and cancer were assumed less viable and more prone to necrosis for their hypermetabolic properties. Recent studies have shown browning of white adipose after fat engraftment in mice. OBJECTIVES: The authors sought to evaluate whether fat transfer could induce browning biogenesis in fat grafts in humans and if it is associated with graft necrosis. METHODS: Necrotic adipose grafts were excised from 11 patients diagnosed with fat necrosis after fat grafting or flap transfer. Non-necrotic fat grafts were from 5 patients who underwent revisionary surgeries after flap transfer. Histology and electronic microscopy as well as protein and gene expression of browning-related marker analyses were performed. RESULTS: Fat grafts with necrosis demonstrated a higher gene expression level of uncoupling protein-1 (greater than fivefold increase, **P < 0.01), a master beige adipocyte marker, than non-necrotic fat grafts. Electronic microscopy and histology showed that browning adipocytes were presented in necrotic adipose in patients. CONCLUSIONS: Fat transfer induced browning adipocytes in patients and was evident in patients with postgrafting necrosis.

Supplementation with Extracellular Vesicles Derived from Adipose-Derived Stem Cells Increases Fat Graft Survival and Browning in Mice: A Cell-Free Approach to Construct Beige Fat from White Fat Grafting.

BACKGROUND: Growing evidence has demonstrated that adipose-derived stem cell-derived extracellular vesicles enhance the survival of fat grafts and the browning of white adipose tissue. We evaluated whether supplementation with adipose-derived stem cell-derived extracellular vesicles promotes the survival and browning of fat grafts. METHODS: Extracellular vesicles derived from adipose-derived stem cells were injected into fat grafts of C57BL/6 mice once per week until postgraft week 12. The grafts were collected and weighed after postgraft weeks 2, 4, and 12. The histological morphology, neovascularization, and the proportion of M2 macrophages of grafts were evaluated. The ability of extracellular vesicles to promote macrophage polarization and catecholamine secretion was detected. Whether the inducement of browning adipose differentiation is extracellular vesicles or the paracrine effect of M2 macrophages polarized by extracellular vesicles was also verified. RESULTS: Grafts treated by extracellular vesicles derived from adipose-derived stem cells showed enhanced beige adipose regeneration with increased neovascularization, M2 macrophage proportion, and norepinephrine secretion at postgraft week 4. Increased retention and decreased fibrosis and necrosis were noted at postgraft week 12. The extracellular vesicles uptake by macrophages promoted M2 type polarization and catecholamine secretion while suppressing M1 type polarization. Of note, browning adipose differentiation with enhanced energy expenditure could be promoted only by the conditioned medium from extracellular vesicle-polarized M2 macrophages but not by extracellular vesicles themselves. CONCLUSIONS: Supplementation with extracellular vesicles derived from adipose-derived stem cells increases fat graft survival and browning by which extracellular vesicles-polarized M2 macrophages secrete catecholamines to promote beige adipose regeneration.

Kinin B1 Receptor Acts in Adipose Tissue to Control Fat Distribution in a Cell-Nonautonomous Manner.

The kinin B1 receptor (B1R) plays a role in inflammatory and metabolic processes. B1R deletion (B1 -/-) protects mice from diet-induced obesity and improves insulin and leptin sensitivity. In contrast, genetic reconstitution of B1R exclusively in adipose tissue reverses the lean phenotype of B1 -/- mice. To study the cell-nonautonomous nature of these effects, we transplanted epididymal white adipose tissue (eWAT) from wild-type donors (B1 +/+) into B1 -/- mice (B1 +/+→B1 -/-) and compared them with autologous controls (B1 +/+→B1 +/+ or B1 -/-→B1 -/-). We then fed these mice a high-fat diet for 16 weeks and investigated their metabolic phenotypes. B1 +/+→B1 -/- mice became obese but not glucose intolerant or insulin resistant, unlike B1 -/-→B1 -/- mice. Moreover, the endogenous adipose tissue of B1 +/+→B1 -/- mice exhibited higher expression of adipocyte markers (e.g., Fabp4 and Adipoq) and changes in the immune cell pool. These mice also developed fatty liver. Wild-type eWAT transplanted into B1 -/- mice normalized circulating insulin, leptin, and epidermal growth factor levels. In conclusion, we demonstrated that B1R in adipose tissue controls the response to diet-induced obesity by promoting adipose tissue expansion and hepatic lipid accumulation in cell-nonautonomous manners.

Remote Effects of Transplanted Perivascular Adipose Tissue on Endothelial Function and Atherosclerosis.

PURPOSE: Perivascular adipose tissue (PVAT) surrounds the arterial adventitia and plays an important role in vascular homeostasis. PVAT expands in obesity, and inflamed PVAT can locally promote endothelial dysfunction and atherosclerosis. Here, using adipose tissue transplantation, we tested the hypothesis that expansion of PVAT can also remotely exacerbate vascular disease. METHODS: Fifty milligrams of abdominal aortic PVAT was isolated from high-fat diet (HFD)-fed wild-type mice and transplanted onto the abdominal aorta of lean LDL receptor knockout mice. Subcutaneous and visceral adipose tissues were used as controls. After HFD feeding for 10 weeks, body weight, glucose/insulin sensitivity, and lipid levels were measured. Adipocytokine gene expression was assessed in the transplanted adipose tissues, and the thoracic aorta was harvested to quantify atherosclerotic lesions by Oil-Red O staining and to assess vasorelaxation by wire myography. RESULTS: PVAT transplantation did not influence body weight, fat composition, lipid levels, or glucose/insulin sensitivity. However, as compared with controls, transplantation of PVAT onto the abdominal aorta increased thoracic aortic atherosclerosis. Furthermore, PVAT transplantation onto the abdominal aorta inhibited endothelium-dependent relaxation in the thoracic aorta. MCP-1 and TNF-α expression was elevated, while adiponectin expression was reduced, in the transplanted PVAT tissue, suggesting augmented inflammation as a potential mechanism for the remote vascular effects of transplanted PVAT. CONCLUSIONS: These data suggest that PVAT expansion and inflammation in obesity can remotely induce endothelial dysfunction and augment atherosclerosis. Identifying the underlying mechanisms may lead to novel approaches for risk assessment and treatment of obesity-related vascular disease.

Sandwiched White Adipose Tissue: A Microphysiological System of Primary Human Adipose Tissue.

White adipose tissue (WAT) is a critical organ in both health and disease. However, physiologically faithful tissue culture models of primary human WAT remain limited, at best. In this study we describe a novel WAT culture system in which primary human WAT is sandwiched between tissue-engineered sheets of adipose-derived stromal cells. This construct, called "sandwiched white adipose tissue" (SWAT), can be defined as a microphysiological system (MPS) since it is a tissue-engineered, multicellular, three-dimensional organ construct produced using human cells. We validated SWAT against the National Institutes of Health MPS standards and found that SWAT is viable in culture for 8 weeks, retains physiologic responses to exogenous signaling, secretes adipokines, and engrafts into animal models. These attributes position SWAT as a powerful tool for the study of WAT physiology, pathophysiology, personalized medicine, and pharmaceutical development.

White Adipose Tissue Is a Reservoir for Memory T Cells and Promotes Protective Memory Responses to Infection.

White adipose tissue bridges body organs and plays a fundamental role in host metabolism. To what extent adipose tissue also contributes to immune surveillance and long-term protective defense remains largely unknown. Here, we have shown that at steady state, white adipose tissue contained abundant memory lymphocyte populations. After infection, white adipose tissue accumulated large numbers of pathogen-specific memory T cells, including tissue-resident cells. Memory T cells in white adipose tissue expressed a distinct metabolic profile, and white adipose tissue from previously infected mice was sufficient to protect uninfected mice from lethal pathogen challenge. Induction of recall responses within white adipose tissue was associated with the collapse of lipid metabolism in favor of antimicrobial responses. Our results suggest that white adipose tissue represents a memory T cell reservoir that provides potent and rapid effector memory responses, positioning this compartment as a potential major contributor to immunological memory.

Facial skin rejuvenation by autologous dermal microfat transfer in photoaged patients: Clinical evaluation and skin surface digital profilometry analysis.

Cumulative long-term exposure to solar ultraviolet radiation promotes premature skin aging characterized by wrinkle formation and reduced skin elasticity. In this study, we assessed whether microfat transfer could improve dermal and subcutaneous tissue thickness loss associated with photoaging. Twenty-one patients affected by facial photoaging (photodamage grade II-IV; age range 35-62 years; 19 females, 2 males; all of Caucasian origin) were treated using minimally invasive autologous dermal white fat transfer harvested with a recently designed microcannula. The results were determined by clinical assessment and patient self-evaluation and quantified by the Antera 3D® dermal digital device for noninvasive, objective, reliable, and accurate assessment of facial skin texture, color, and wrinkle characteristics. Compared with the pretreatment condition, the increment in soft tissue volume and improvement in skin quality and texture were assessed by a dermatologist after treatment. In addition, instrumental evaluation by digital skin profilometry of the treated areas revealed a 41% reduction in average wrinkle depth (7.29 ± 1.04 × 10-2 mm pretreatment vs. 4.31 ± 1.16 × 10-2 mm at 90 days posttreatment; p < 0.001), improved skin texture, more homogeneous and uniform skin color, and declined facial hemoglobin and melanin concentrations. The majority of patients (above 90%) reported improvements in self-perception. No significant complications were reported throughout the study. In conclusion, by using digital profilometry analysis as an objective and innovative tool to determine the outcome of treatment, we demonstrated that autologous microfat transfer is a safe and well-tolerated procedure with measurable beneficial effects on facial skin aging.

Adipose-derived circulating miRNAs regulate gene expression in other tissues.

Adipose tissue is a major site of energy storage and has a role in the regulation of metabolism through the release of adipokines. Here we show that mice with an adipose-tissue-specific knockout of the microRNA (miRNA)-processing enzyme Dicer (ADicerKO), as well as humans with lipodystrophy, exhibit a substantial decrease in levels of circulating exosomal miRNAs. Transplantation of both white and brown adipose tissue-brown especially-into ADicerKO mice restores the level of numerous circulating miRNAs that are associated with an improvement in glucose tolerance and a reduction in hepatic Fgf21 mRNA and circulating FGF21. This gene regulation can be mimicked by the administration of normal, but not ADicerKO, serum exosomes. Expression of a human-specific miRNA in the brown adipose tissue of one mouse in vivo can also regulate its 3' UTR reporter in the liver of another mouse through serum exosomal transfer. Thus, adipose tissue constitutes an important source of circulating exosomal miRNAs, which can regulate gene expression in distant tissues and thereby serve as a previously undescribed form of adipokine.

Unsuppressed lipolysis in adipocytes is linked with enhanced gluconeogenesis and altered bile acid physiology in Insr(P1195L/+) mice fed high-fat-diet.

High-fat diet (HFD) triggers insulin resistance and diabetes mellitus, but their link remains unclear. Characterization of overt hyperglycemia in insulin receptor mutant (Insr(P1195L/+)) mice exposed to HFD (Insr(P1195L/+)/HFD mice) revealed increased glucose-6-phosphatase (G6pc) expression in liver and increased gluconeogenesis from glycerol. Lipolysis in white adipose tissues (WAT) and lipolysis-induced blood glucose rise were increased in Insr(P1195L/+)/HFD mice, while wild-type WAT transplantation ameliorated the hyperglycemia and the increased G6pc expression. We found that the expressions of genes involved in bile acid (BA) metabolism were altered in Insr(P1195L/+)/HFD liver. Among these, the expression of Cyp7a1, a BA synthesis enzyme, was insulin-dependent and was markedly decreased in Insr(P1195L/+)/HFD liver. Reduced Cyp7a1 expression in Insr(P1195L/+)/HFD liver was rescued by WAT transplantation, and the expression of Cyp7a1 was suppressed by glycerol administration in wild-type liver. These findings suggest that unsuppressed lipolysis in adipocytes elicited by HFD feeding is linked with enhanced gluconeogenesis from glycerol and with alterations in BA physiology in Insr(P1195L/+)/HFD liver.

In vivo evidence for unidentified leptin-induced circulating factors that control white fat mass.

Fat transplants increase body fat mass without changing the energy status of an animal and provide a tool for investigating control of total body fat. Early transplant studies found that small pieces of transplanted fat took on the morphology of the transplant recipient. Experiments described here tested whether this response was dependent upon expression of leptin receptors in either transplanted fat or the recipient mouse. Fat from leptin receptor deficient db/db mice or wild-type mice was placed subcutaneously in db/db mice. After 12 wk, cell size distribution in the transplant was the same as in endogenous fat of the recipient. Thus, wild-type fat cells, which express leptin receptors, were enlarged in a hyperleptinemic environment, indicating that leptin does not directly control adipocyte size. By contrast, db/db or wild-type fat transplanted into wild-type mice decreased in size, suggesting that a functional leptin system in the recipient is required for body fat mass to be controlled. In the final experiment, wild-type fat was transplanted into a db/db mouse parabiosed to either another db/db mouse to an ob/ob mouse or in control pairs in which both parabionts were ob/ob mice. Transplants increased in size in db/db-db/db pairs, decreased in db/db-ob/ob pairs and did not change in ob/ob-ob/ob pairs. We propose that leptin from db/db parabionts activated leptin receptors in their ob/ob partners. This, in turn, stimulated release of unidentified circulating factors, which travelled back to the db/db partner and acted on the transplant to reduce fat cell size.

Exercise Effects on White Adipose Tissue: Beiging and Metabolic Adaptations.

Regular physical activity and exercise training have long been known to cause adaptations to white adipose tissue (WAT), including decreases in cell size and lipid content and increases in mitochondrial proteins. In this article, we discuss recent studies that have investigated the effects of exercise training on mitochondrial function, the "beiging" of WAT, regulation of adipokines, metabolic effects of trained adipose tissue on systemic metabolism, and depot-specific responses to exercise training. The major WAT depots in the body are found in the visceral cavity (vWAT) and subcutaneously (scWAT). In rodent models, exercise training increases mitochondrial biogenesis and activity in both these adipose tissue depots. Exercise training also increases expression of the brown adipocyte marker uncoupling protein 1 (UCP1) in both adipose tissue depots, although these effects are much more pronounced in scWAT. Consistent with the increase in UCP1, exercise training increases the presence of brown-like adipocytes in scWAT, also known as browning or beiging. Training results in changes in the gene expression of thousands of scWAT genes and an altered adipokine profile in both scWAT and vWAT. Transplantation of trained scWAT in sedentary recipient mice results in striking improvements in skeletal muscle glucose uptake and whole-body metabolic homeostasis. Human and rodent exercise studies have indicated that exercise training can alter circulating adipokine concentration as well as adipokine expression in adipose tissue. Thus, the profound changes to WAT in response to exercise training may be part of the mechanism by which exercise improves whole-body metabolic health.

Adipose tissue and breast cancer progression: a link between metabolism and cancer.

INTRODUCTION/AIMS: Obesity, an excess accumulation of adipose tissue occurring in mammalians when caloric intake exceeds energy expenditure, is associated with an increased incidence, morbidity and mortality from several types of neoplastic diseases including postmenopausal breast cancer. METHODS AND RESULTS: Several investigators have recently studied the role of human white adipose tissue (WAT) progenitors in preclinical models of breast cancer. WAT progenitors were found to promote breast cancer local growth, angiogenesis, EMT, migration and metastatic spreading. Breast cancer patients with intraepithelial neoplasia who received autologous WAT cells for breast reconstruction after surgical removal of breast cancer showed an increased risk of recurrence of local events when compared to controls. DISCUSSION/CONCLUSION: There is an urgent need for a better understanding of the role of WAT progenitors in breast cancer local and metastatic growth. A rigorous cancer screening and follow-up of patients enrolled for WAT progenitor-based therapies should be implemented.

Molecular pathways involved in the improvement of non-alcoholic fatty liver disease.

UNLABELLED: Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis are components of the metabolic syndrome. Serum leptin levels are elevated in obesity, but the role of leptin in the pathophysiology of the liver involvement is still unclear. To identify the effects and mechanisms by which leptin influences the pathogenesis of NAFLD, we performed epididymal white adipose tissue (eWAT) transplantation from congenic wild-type mice into the subcutaneous dorsal area of Lep(ob/ob) recipient mice and compared the results with those of the Lep(ob/ob) sham-operated mice. The mice were followed for 102-216 days. During killing, the transplanted mice had significantly lost body weight and exhibited significantly higher leptin levels, improved glucose tolerance, and lower liver injury scores than the sham-operated mice. Liver microarray analysis showed that novel pathways related to GA-binding protein (GABP) transcription factor targets, pheromone binding, and olfactory signaling were differentially expressed in the transplanted mice. Our data also replicate pathways known to be involved in NAFLD, such as those involved in the regulation of microRNAs, lipid, glucose, and glutathione metabolism, peroxisome proliferator-activated receptor signaling, cellular regulation, carboxylic acid processes, iron, heme, and tetrapyrrole binding, immunity and inflammation, insulin signaling, cytochrome P450 function, and cancer. CONCLUSION: wild-type eWAT transplantation into Lep(ob/ob) mice led to improvements in metabolism, body weight, and liver injury, possibly attributed to the production of leptin by the transplanted eWAT. These improvements were accompanied by the differential expression of novel pathways. The causal relationship between GABP downregulation and NAFLD improvement remains to be determined.

Short commentaries on data published by Petit et al. on locoregional risk after lipofilling in breast cancer patients.

Lipofilling is becoming part of the breast reshaping after quadrantectomy or mastectomy in breast cancer patients, but there are open questions of its safety.

"SIEF"--simultaneous implant exchange with fat: a new option in revision breast implant surgery.

UNLABELLED: A technique of implant exchange is reported using recipient-site preexpansion followed by autologous fat transplantation to the breast in 12 consecutive patients with breast implants who desired implant removal. Recipient-site preexpansion, used 2 weeks before fat grafting, may have both practical and theoretical benefits in increasing the breast subcutaneous space and stimulating the recipient-site microcellular environment overlying the prosthetic implant, allowing the subcutaneous insertion of a sufficient core volume of donor graft at the time of prosthetic explantation. In the cases described, the postexplantation breast volume at 9 months to 1 year postoperatively by quantitative three-dimensional imaging was equal to or greater than the preexplantation composite volume of breast and implant. Preexpansion before implant exchange with fat affords a more abundant space, completely independent from the subglandular or submuscular planes. In this new space, the "third space" of the breast, it is possible to technically place graft into the breast subcutaneous tissue and alleviate breast asymmetry resulting from pocket distortions caused by capsular contracture or by implant pocket drift. Observing breast augmentation with implants and with fat grafting in the same patient affords a unique opportunity to analyze some of the key differences between the two techniques. Recipient-site preexpansion and simultaneous implant exchange with fat (SIEF) should be added to the list of applications where fat grafting to the breasts may have early clinical utility and portends the use of fat used in conjunction with breast implants to achieve better patient outcomes. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, V.

Achieving aesthetic results in facial reconstructive microsurgery: planning and executing secondary refinements.

BACKGROUND: Free tissue transfer to improve bulk and contour in facial deformities has been proven useful, yet refinements that turn an acceptable result into an excellent result are essential to reconstruction. The authors reviewed their experience and described these refinements. METHODS: The charts of 371 free tissue transfer cases (1989 to 2010) performed by the senior author (J.W.S.) were reviewed. Free tissue transfer of a circumflex scapular variant flap or superficial inferior epigastric was performed to treat deformities arising from hemifacial atrophy (n = 126), hemifacial microsomia (n = 89), radiation therapy (n = 40), bilateral malformations including lupus and polymyositis (n = 50), other congenital anomalies (n = 25), facial palsy (n = 17), and burns and trauma (n = 24). RESULTS: Revision surgery planning began at initial flap operation where the flap was stretched maximally and interdigitated with recipient tissue. More tissue was required in the malar region. Revision refinement was indicated in all cases (after 6 months). Flap revision involved liposuction, debulking, reelevation, and release of tethering, followed by tissue rearrangement by means of advancement, rotation, transposition, and/or turnover flaps of subcutaneous tissues from the previous free flap. The jawline frequently required more debulking. Periorbital reconstruction was combined with lower lid support with or without canthal repositioning. Conventional face-lift techniques with the flap as superficial musculoaponeurotic system augmented the result. Autologous fat injection to the alar rim, medial canthus, upper eyelid, and lip was a useful adjunct. Severe lip deficiencies were addressed with local flaps. CONCLUSION: The keys to improving results were continual critical reassessment, open-mindedness to new approaches, and maintaining high expectations. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, V.

In situ production of innate immune cells in murine white adipose tissue.

White adipose tissue (WAT) is the focus of new interest because of the presence of an abundant and complex immune cell population that is involved in key pathologies such as metabolic syndrome. Based on in vivo reconstitution assays, it is thought that these immune cells are derived from the bone marrow (BM). However, previous studies have shown that WAT exhibits specific hematopoietic activity exerted by an unknown subpopulation of cells. In the present study, we prospectively isolated a peculiar hematopoietic stem/progenitor cell population from murine WAT. The cells are phenotypically similar to BM hematopoietic stem cells and are able to differentiate into both myeloid and lymphoid lineages in vitro. In competitive repopulation assays in vivo, they reconstituted the innate immune compartment in WAT preferentially and more efficiently than BM cells, but did not reconstitute hematopoietic organs. They were also able to give rise to multilineage engraftment in both secondary recipients and in utero transplantation. Therefore, we propose that WAT hematopoietic cells constitute a population of immature cells that are able to renew innate immune cell populations. Considering the amount of WAT in adults, our results suggest that WAT hematopoietic activity controls WAT inflammatory processes and also supports innate immune responses in other organs.

VEGF microsphere technology to enhance vascularization in fat grafting.

Vascularization is crucial for implantation of engineered tissues in reconstructive surgery. Polypeptides encapsulated in microspheres can be efficiently transported to their site of action and released in a sustained dosage. We evaluated the effect of delivering vascular endothelial growth factor (VEGF)-encapsulated microspheres in a lipoaspirate scaffold on vascularization and tissue survival. The VEGF-loaded (n=6) and empty (n=6) poly(lactic-co-glycolic acid) microspheres in human lipoaspirate and the human lipoaspirate alone (n=6) were injected subcutaneously into the flanks of athymic nude mice. Three mice from each group were killed, and grafts were explanted at weeks 3 and 6. Increases in mass and volume of VEGF samples, as well as decreases in empty and lipoaspirate-only samples, were observed at 3 and 6 weeks, reaching statistical significance at 6 weeks. Hematoxylin and eosin and CD31+ imaging demonstrated significantly greater vascularization in VEGF samples than in both the empty and lipoaspirate-only groups at both 3 and 6 weeks.