REGENERATIVE MEDICINE Scarless Wound Healing: Chasing the Holy Grail Graham G. Walmsley, B.A. Zeshaan N. Maan, M.B.B.S., M.S. Victor W. Wong, M.D. Dominik Duscher, M.D. Michael S. Hu, M.D. Elizabeth R. Zielins, M.D. Taylor Wearda, B.A. Ethan Muhonen, B.A. Adrian McArdle, M.B., B.Ch., B.A.O. Ruth Tevlin, M.B., B.Ch., B.A.O. David A. Atashroo, M.D. Kshemendra Senarath-Yapa, M.A., M.B.B.Chir. H. Peter Lorenz, M.D. Geoffrey C. Gurtner, M.D. Michael T. Longaker, M.D., M.B.A. Stanford, Calif.; and Baltimore, Md.

Summary: Over 100 million patients acquire scars in the industrialized world each year, primarily as a result of elective operations. Although undefined, the global incidence of scarring is even larger, extending to significant numbers of burn and other trauma-related wounds. Scars have the potential to exert a profound psychological and physical impact on the individual. Beyond aesthetic considerations and potential disfigurement, scarring can result in restriction of movement and reduced quality of life. The formation of a scar following skin injury is a consequence of wound healing occurring through reparative rather than regenerative mechanisms. In this article, the authors review the basic stages of wound healing; differences between adult and fetal wound healing; various mechanical, genetic, and pharmacologic strategies to reduce scarring; and the biology of skin stem/progenitor cells that may hold the key to scarless regeneration.   (Plast. Reconstr. Surg. 135: 907, 2015.)

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dult mammalian wound healing involves a complex series of molecular and cellular events that typically culminate in a fibrotic “patch-like” repair. Wound healing can be considered in three overlapping stages. The first stage, inflammation, begins with the formation of a fibrin clot in conjunction with platelets at the site of injury. This provides a temporary extracellular matrix, and a stimulus that recruits inflammatory cells to the wound environment. The first immune cells to arrive are neutrophils, whose primary function is prevention of bacterial infection. They are found in highest concentrations 1 to 2 days after injury.1,2 Days 2 and 3 after wounding see the arrival of circulating monocytes and macrophages,3 which play a role in the transition between the inflammatory stage and subsequent proliferative stage of wound healing. Early wound macrophages From the Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Plastic and Reconstructive Surgery, and the Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine; and the Department of Surgery, Division of Plastic and Reconstructive Surgery, Johns Hopkins University School of Medicine. Received for publication May 15, 2014; accepted August 14, 2014. Copyright © 2015 by the American Society of Plastic Surgeons DOI: 10.1097/PRS.0000000000000972

serve to phagocytose apoptotic neutrophils and clear debris. In days 5 to 7 after injury, wound macrophages adopt a more antiinflammatory role in preparation for tissue rebuilding. Moreover, secretion of growth factors, such as plateletderived growth factor and transforming growth factor (TGF)-β, allows macrophages to stimulate fibroblast migration and activation.1,4 The proliferative phase of wound healing is estimated to begin within 48 hours to 10 days after wounding and is characterized by replacement of the temporary, fibrin-based extracellular matrix with granulation tissue,3 which consists of a vascularized extracellular matrix formed by fibroblasts and endothelial cells. The formation of granulation tissue facilitates reepithelialization, which occurs through the migration and proliferation of keratinocytes, beginning hours after wounding.5 The migrating fibroblasts secrete newly formed extracellular matrix, composed of glycosaminoglycans, proteoglycans, and collagen, although the proportion of type III to type I collagen is higher in early scar tissue than Disclosure: The authors wish to voluntarily declare a potential conflict of interest involving Neodyne Biosciences. Dr. Gurtner and Dr. Longaker are founders and serve on the board of directors. The other authors have no financial information to disclose.

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Plastic and Reconstructive Surgery • March 2015 in unwounded tissue or mature scar.1 In parallel, myofibroblast-mediated wound contraction reduces the original wound size by bringing wound edges closer together.1,3 Furthermore, migration of endothelial cells, mediated largely by vascular endothelial growth factor production from macrophages and fibroblasts,1,4 and subsequent angiogenesis of the newly formed tissue are responsible for the red hue of immature scars. In humans, wound closure is achieved primarily through granulation tissue formation and reepithelialization, whereas in loose-skinned animals (e.g., mice), wound closure is achieved primarily through contraction. Although collagen deposition by fibroblasts and myofibroblasts is typically at its maximal level 3 weeks after injury, at this point, wound strength is at a minimum. Subsequent remodeling of collagen, along with a shift toward a more normal proportion of type I to type III collagen, results in a mature scar that has 80 percent of the strength of normal skin after 3 to 4 months.1 Remodeling, the final phase of wound healing, begins after reepithelialization and wound closure, typically from 14 to 21 days after injury, and can last for over 1 year.3 Reorganization of extracellular matrix by matrix metalloproteinases and collagenases is accompanied by decreased cellularity and vascularity of scar tissue. Epithelial

appendages (i.e., hair follicles, sweat glands, and sebaceous glands) are not reformed, and a healed scar therefore represents a “bare area.” Thus, the final product of the body’s response to injury is a relatively acellular mass of epithelialized extracellular matrix that has a distinct life cycle of its own (Fig. 1).1,6–8 Although normal wound healing typically follows the phases and time frames outlined above, the degree of scarring occurs along a continuum. The first type of pathologic scarring (hypertrophic) typically develops following wound closure under excess tension or infection.9 Hypertrophic scarring is characterized by wide, raised scars that remain within the original borders of injury and have a rapid growth phase of 6 months with gradual regression to a flat scar over a period of 1 to 3 years.10 Another type of pathologic scarring (i.e., keloids) differs markedly from both normal and hypertrophic scars. Keloids appear as firm, bosselated tumors that extend beyond the original borders of injury, do not regress spontaneously, and can develop up to several years after cutaneous injury.11

FETAL SCARLESS WOUND HEALING Early fetal skin has unique wound healing capabilities, with an intrinsic ability to

Fig. 1. Schematic representing the timing of major clinical manifestations and their underlying biology in the life cycle of scar formation. Erythema, heat, and swelling occur from days 0 to 6 and are attributable to platelet degranulation, vasodilation, and leukocytic infiltration. Joint contractures peak at approximately 3 to 6 months, and are caused by altered matrix metalloproteinase expression, changes in extracellular matrix composition, collagen cross-linking, and increased myofibroblasts at the scar site. Finally, pruritus and pain typically peak at 9 months and are caused by small-fiber neuropathy and histamine release.

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Volume 135, Number 3 • Scarless Wound Healing regenerate tissue rather than form a scar.12,13 Understanding the mechanisms underlying regenerative healing in the fetus may potentially inform scarless healing in adults. After wounding, fetal skin is capable of rapid reepithelialization,14–16 with minimal inflammation.17 Fetal skin undergoes many changes during development. A single layer of ectodermal cells, seen at days 7 and 8 in human gestation, changes into stratified, keratinized epithelium between 22 and 24 weeks. Keratin expression varies throughout development and differs greatly from that of adult epithelium.18 The composition of extracellular matrix is thought to contribute significantly to fetal healing. Fetal tissues display an elevated ratio of type III to type I collagen in comparison with adult tissues.19 The loose network of extracellular matrix helps to promote cellular migration, proliferation, and differentiation, thought to play an important role in wound healing.15,20 The extracellular matrix of fetal tissue is also rich in hyaluronic acid, a glycosaminoglycan that promotes a microenvironment conducive to cell motility and inhibits the proliferation of fetal fibroblasts.21,22 The key differences between fetal and adult wounds are summarized in Table 1.12–22 Although the environment within which the fetus resides has been thought to contribute to its regenerative healing,23,24 prior studies using transplantation methodologies have suggested that the ability of fetal skin to heal scarlessly is cell intrinsic. Longaker and colleagues showed that full-thickness adult sheep skin transplanted onto the dorsa of 60-day gestational fetal lambs healed with scar formation after incisional wounding.25 In a reciprocal experiment, Lorenz et al. showed that human fetal skin transplanted subcutaneously into adult athymic mice recapitulated the scarless phenotype after incisional wounding.13 Collectively, these experiments suggest that properties intrinsic in the transplanted tissues are more significant than extrinsic

environmental factors in determining the outcome of wound healing and suggest that therapies directly targeting the fibroblasts largely responsible for scar formation may be effective. In summary, understanding fetal wound healing offers a blueprint for manipulating postnatal healing to be more fetal-like.

SKIN STEM CELLS IN WOUND HEALING To date, exciting advances in genetic lineage-tracing technologies, cellular assays, and imaging techniques have revealed important stem and progenitor cell reservoirs in the interfollicular epidermis, the eccrine sweat glands, and the hair follicle.26 Given that scarring following a full-thickness excisional wound is defined as a bare area lacking sweat glands, sebaceous glands, and hair follicle regeneration, a thorough understanding of these stem and progenitor cells will likely contribute to achieving the ultimate goal of scarless wound healing following injury. Interfollicular Epidermis The epidermis is a dynamic structure tasked with protecting the body from the outside world. As such, it frequently sustains damage and must repair itself.3 This healing is accomplished through the contributions of numerous tissuespecific stem and progenitor cell populations located throughout the epidermis and its associated appendages.27 During wound healing, a population of “slow-cycling cells” is proposed to exhibit autocrine regulation28 until mobilized by a wound healing response. When wounding occurs, slow-cycling cell division frequency increases, providing excess daughter cells to help repair the damage. Given the gathering evidence for behavioral plasticity in injured epithelia, with progenitors capable of reverting to multipotent states and multipotent cells differentiating to fill unipotent roles,29,30 these data hold exciting ramifications

Table 1.  Key Differences between Adult and Fetal Wound Healing Inflammation Hyaluronic acid21,22 Collagen15,20 Keratin14,18 Mast cells15,16 TGF-β12–14 Stem cells12 17,18

Fetal Wounds

Adult Wounds

Few or no inflammatory cells High, prolonged levels, promotes cellular movement Elevated ratio of type III compared to type I K8 and K19 are present during fetal development Low numbers and less mature Low expression of TGF-β1 and TGF-β2 Higher levels of MSCs at injury sites with ­accompanying E-cadherin–positive cells

Many inflammatory cells Lower levels, inhibits cellular movement Elevated ratio of type I compared to type III Absence of K8 and K19 Higher numbers, more mature High expression of TGF-β1 and TGF-β2 Lower levels of MSCs at injury sites without accompanying E-cadherin–positive cells

MSCs, mesenchymal stem cells.

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Plastic and Reconstructive Surgery • March 2015 for researchers hoping to shift wound healing away from scarring and toward a more regenerative phenotype. Sweat Glands Eccrine sweat glands (henceforth “sweat glands”) are widely distributed across the adult human body, and can reach a density of up to approximately 700 glands/cm2 of adult human skin.31 Loss of sweat glands in burn patients remains an unsolved problem. Recent groundbreaking studies of the biology and development of these epidermal appendages have identified gland-specific progenitors,32 and revealed the roles of these cells in development, homeostasis, and wound repair.32,33 In elucidating the developmental lineage and healing capabilities of sweat gland structures, the common heritage of all epidermal appendages is further underscored. Developmentally, nascent sweat glands form from a downfolding of multipotent, K5- and K15-positive, epidermal basal progenitors. As morphology becomes more specified, these multipotent progenitors further differentiate into unipotent luminal and myoepithelial progenitor populations. Two sweat apparatus progenitor populations are of particular interest in epidermal regeneration and in considering potential therapeutic strategies. These are the progenitor populations housed in the myoepithelium and the multipotent progenitors found in the sweat gland duct. When engrafted into murine back skin, myoepithelial progenitors have intriguingly been observed to form new epidermis,32,34 which reflects the common multipotent K14-positive basal cell origins of both the interfollicular epidermis and the sweat glands. Regarding native wound healing responses, the luminal and myoepithelial progenitors in the secretory coil have only been observed to proliferate and contribute to the repair of deep glandular wounds.32 In contrast, progenitors in the sweat gland duct have been noted to proliferate and contribute to healing scratch wounds in the interfollicular epidermis. This observation, speculated to be orchestrated by a mechanism of keratinocyte outgrowth,33 is of immense interest to translational medicine. As physicians and researchers further decipher the myriad iterations of healing strategies used throughout the epidermis, we may concomitantly open the door to a future where therapeutically selecting from among epidermal progenitors becomes reality.

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Hair Follicles The mammalian hair follicle is a dynamic structure known to house numerous progenitor cell populations and contribute to the repair of injured skin.35–38 The rapid turnover of these cells makes lineage analysis a considerable challenge, but a multipotent progenitor population capable of regenerating basal keratinocytes has been characterized.39 Cells in the bulge are multipotent, have high proliferative potential, and are capable of giving rise to all epithelial layers in the hair follicle.40 During wound healing, cells in the bulge and upper follicle migrate and contribute to interfollicular epidermis regeneration.41 A recent study successfully generated folliculogenic human epithelial stem cells from induced pluripotent stem cells and demonstrated differentiation along all hair follicle lineages in skin reconstitution assays.42 Given that loss of dermal appendages is an unwanted feature of cutaneous scarring, the transplantation of induced pluripotent stem cell– derived folliculogenic epithelial stem cells holds promise for the regeneration of normal skin architecture following cutaneous injury.

FIBROBLAST LINEAGES Dermal fibroblasts represent a heterogeneous population of stromal cells and are responsible for scar deposition following cutaneous injury. In 1987, Cormack demonstrated that two subpopulations of fibroblasts reside within the papillary and reticular dermis, respectively.43 Later, a third subpopulation was found associated with hair follicles in the dermal papilla region of the follicle.44,45 When cultured, these fibroblast subtypes exhibit differences in growth potential, morphology, and growth factor/cytokines profiles.46–49 However, these and other studies examining fibroblast heterogeneity have largely failed to convincingly demonstrate developmental and functional diversity among fibroblasts in vivo, particularly in the case of dermal skin fibroblasts. A recent article by Driskell et al. revealed that skin connective tissue contains two distinct lineages of fibroblasts and that each lineage exhibits specific functional properties.50 The first lineage gives rise to the upper dermis and is responsible for hair follicle formation, and the second gives rise to the lower dermis and is responsible for the deposition of connective tissue following excisional wounding. The study demonstrated, for the first time, that distinct embryonic origins for functional subtypes of fibroblasts exist. Most importantly from the perspective of scarless

Volume 135, Number 3 • Scarless Wound Healing wound healing, the study showed that epidermal B-catenin activation stimulates the expansion of the upper dermal lineage and renders excisional wounds permissive to hair follicle growth. As our understanding of the heterogeneous fibroblast population continues to evolve, novel molecular and cellular targets for the modulation of both scar deposition and dermal appendage regeneration will likely emerge.

STRATEGIES TO REDUCE SCARRING Scars resulting from surgical incisions represent an enormous market totaling over 100 million cases per year in the developed world.51 Cutaneous injury initiates a complex cascade of molecular and cellular events within a dynamic extracellular matrix, features of which are common among fibrotic diseases (Fig. 2).2, 52–54 For this reason, treatment modalities capable of broadly influencing biological repair pathways hold the greatest potential for scar reduction. Strategies to minimize the appearance of these

scars and restore the integrity of normal skin will be reviewed here. Current Pharmaceutical Strategies Topical therapy and intralesional injections of corticosteroids have formed the mainstay of nonsurgical therapy for the treatment of keloids and hypertrophic scars for many years. Although this approach can be effective, with success rates of 50 to 100 percent reported in the literature,55–57 it is also associated with hypopigmentation, dermal atrophy, telangiectasia, widening of the scar, and delayed wound healing.56 Although the precise mechanisms through which steroids exert their effects in this context have not been fully elucidated, their principal role is thought to be inhibition of the inflammatory response, suppressing fibroblast growth and promoting collagen degradation.57 Triamcinolone, the most widely used agent, is able to inhibit TGF-β1 expression and induce apoptosis in fibroblasts,58 and inhibit TGFβ1, COL4A1, and COL7A1 in keratinocytes.59

Fig. 2. Schematic representing the common biology of fibrotic diseases such as idiopathic pulmonary fibrosis, renal fibrosis, scleroderma, cirrhosis, and scarring. In general, transient or chronic inflammatory states characterized by the presence of neutrophils, macrophages, and lymphocytes stimulate resident fibroblasts to produce and secrete collagen proteins, which then crosslink to form mature collagen fibers. The link between inflammation and fibrosis includes key mediators such as the mechanoresponsive focal adhesion kinase–monocyte chemoattractant protein pathway, and the TGF-β and Wnt signaling pathways.

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Plastic and Reconstructive Surgery • March 2015 Evidence supporting the use of calcium antagonists as a standalone treatment remains scarce, although some studies have used them as an adjunct to other therapies. Intralesional verapamil used as an adjunct to surgical excision and topical silicone was found to increase the cure rate of keloids from 18 percent with the latter two treatments alone, to 54 percent in combination with verapamil.60 Although the mechanism of action remains poorly delineated, there is evidence that verapamil injection decreases collagen production.61 Verapamil has also been shown to reduce the production of interleukin-6 (which is thought to be a mitogenic signal for keratinocytes) and vascular endothelial growth factor in fibroblasts harvested from the central portions of keloid scars.62 Topical agents based on plant extracts also exist. Mederma (Merz Pharmaceuticals, Frankfurt, Germany) is a topical gel used to improve scar appearance, with the extract from the common onion as the main active ingredient. Onion extract contains a bioflavonoid with both antihistamine and antiproliferative effects.63 Although studies have shown that Mederma improves collagen organization following excisional ear wounding in rabbits,63 a 2006 clinical trial found no statistically significant difference in hypertrophic scar appearance compared with standard petroleum emollient.64 These findings illustrate a common pattern seen in treatments attempting to reduce scar formation: success in animal models is often not recapitulated in human clinical trials. Gene Targets A number of genes involved in the scarring response have been identified, typically influencing fibrosis by regulating collagen production and degradation. Molecularly targeted therapies seek to either inhibit or up-regulate the pathways controlled by these genes to decrease collagen synthesis.65–68 TGF-β is a major regulator of fibroblast physiology. TGF-β expression has been strongly implicated in the fibroblast-to-myofibroblast transition and in the production and secretion of extracellular matrix. The significant role of TGF-β has garnered it attention as a potential target for fibrosis therapy. The gene is expressed as three isoforms in humans with differential activity. TGF-β1 and TGF-β2 both promote fibrosis, whereas TGF-β3 has been shown to decrease fibrosis and scarring.69,70 Therapeutic strategies targeting TGF-β signaling involve the use of either TGF-β1 and TGF-β2 inhibitors and neutralizing antibodies or up-regulating TGF-β3 signaling.

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Neutralizing antibodies bind directly to TGFβ1 and TGF-β2, preventing them from binding to and activating TGF-β receptors. In animal models, these antibodies have successfully reduced fibrosis across a number of organs, including a reduction in cutaneous scarring following injury.71–74 Unfortunately, the success of TGF-β antibodies in animal models has not translated across to human clinical trials. Metelimumab, the first anti–TGF-β antibody used in clinical trials for the treatment of systemic sclerosis, demonstrated no significant benefits.75 The TGF-β and platelet-derived growth factor inhibitor imatinib mesylate was also unsuccessful in phase II clinical trials targeting scleroderma.76 Up-regulating TGF-β3 has also demonstrated efficacy in animal models, reducing scarring in a number of studies.77–80 Unfortunately, unpublished data from an international phase III clinical trial has shown that TGF-β3 has no significant effect on scarring (Juvista EU; Renovo, Bristol, United Kingdom). A number of other genes involved in fibrosis have also been addressed in animal models, including modulation of Wnt signaling,65 targeting the hedgehog pathway,81 interleukin-10, which has been implicated in a variety of mechanisms leading to fibrosis,69,82–86 histone deacetylase,66 early growth response gene-1,82 and heat shock protein 90.67 Unfortunately, efficacy is limited to animal studies. Although murine models for both normal87 and hypertrophic scarring88 exist, the discrepancy between animal and clinical trial results highlights differences in signaling between the two species and suggests the need for alternative approaches to address the signaling overlap. Dermal Substitutes Dermal substitutes provide coverage to the wound site and establish a matrix that encourages engraftment and proliferation of endogenous cells and enhances the function of transplanted cells. Acellular matrices, derived from animal or human tissue through the removal of living cells, are biocompatible, remain morphologically similar to natural tissue structure, and have mechanical properties similar to host skin.89 Acellular matrices derived from human dermis currently in clinical use include AlloDerm (LifeCell Corp., Branchburg, N.J.), DermaMatrix (Synthes, West Chester, Pa.), and FlexHD (Ethicon, Inc., Somerville, N.J.). Dermal substitutes using human chorion and amnion have also been used. Among these, Epifix is composed of dehydrated human amnion/chorion membrane (MiMedx Group, Kennesaw, Ga.). Grafix (Osiris Therapeutics, Inc.,

Volume 135, Number 3 • Scarless Wound Healing Columbia, Md.) is cryopreserved placental membrane with native mesenchymal stem cells used as a covering to promote healing and tissue repair. Xenografts are skin substitutes for use in humans harvested from animals. The OASIS Wound Matrix (Cook Biotech, Inc., West Lafayette, Ind.), derived from the submucosal layers of porcine jejunum, is one of the more commonly used non–human-derived acellular matrices in clinical use. Similarly, Surgisis (Cook Biotech) is a biological graft extracted from porcine small intestinal submucosa. More recently, fetal bovine dermal matrices have becomes available, including PriMatrix (TEI Biosciences, Inc., Waltham, Mass.) and SurgiMend (TEI Biosciences). Synthetic fabricated scaffolds are typically composed of proteins normally present in extracellular matrix. Integra (Integra, Plainsboro, N.J.) is composed of bovine collagen and chondroitin6-sulfate cross-linked with glutaraldehyde with a silicone membrane as a temporary epidermis.90 Chitosan-based scaffolds have shown promise as a supportive template for fibroblast attachment.91 A biodegradable porous scaffold composed of collagen, hyaluronic acid, and gelatin accelerated wound closure by providing optimal pore size/ water absorption for human skin cell growth.92 Pullulan-collagen composite dermal hydrogels have been shown to accelerate early wound healing, potentially by modifying cell recruitment and augmenting granulation tissue formation.93 Matrices seeded with cells may also play a role in preventing scar formation. Although useful for enhancing the rate of wound healing, particularly in the context of burns and large excisional wounds, dermal substitutes have not been shown to significantly impact scarring. In the future, the seeding of purified stem cell populations and niche components onto dermal substitutes may allow for a more significant shift away from repair and toward regeneration. Progress on this front will first require a more thorough characterization of skin stem cell populations and their niches. Mechanical Offloading An alternative approach to limiting fibrosis in the setting of cutaneous injury is mechanical offloading. Mechanical tension plays a significant role in the development of fibrosis, activating a number of mechanoresponsive signaling pathways, with focal adhesion kinase being one of the best described.53,94 Plastic surgeons have long been aware of the presence of Langer lines in human skin, which correspond to the natural

orientation of collagen fibers within the dermis. Incisions made parallel to Langer lines generally heal with less scar (because of reduced tension) than those placed perpendicular.95 Paper tape96,97 and silicone sheets98–100 have been reported to reduce scarring, with a proposed mechanism related primarily to tensile reduction. In a randomized controlled trial of 39 patients, the delayed application of paper tapes to closed surgical wounds for up to 12 weeks postoperatively was shown to reduce scar size and decrease the likelihood of developing a hypertrophic scar.96 A study involving 195 patients with torso scars (63 taped and 73 controls) found an overall improvement in scar appearance in taped compared with control scars.97 Success in animal models has also been reported, with microporous paper tape application demonstrating reduced hypertrophic scarring in a rabbit ear model.101 In an attempt to further improve the action of tapes, hybrid materials such as Dynaclose (mediGroup Australia Pty Ltd., Melbourne, Victoria, Australia), a tape with a central region of silicone elastomer, have been developed.102 However, when used as an alternative to sutures, paper tape alone had no significant effect on scar size versus sutures alone in closing abdominal wounds.103 Similarly, the use of Steri-Strip S (3M, St. Paul, Minn.) tapes compared to sutures alone for the closure of median sternotomy incisions demonstrated no significant difference in scar size.104 Silicone gel sheets have been widely used for many years despite an overall lack of knowledge regarding the underlying mechanism and limited evidence supporting clinical efficacy. Although studies have shown that silicone gel sheets effectively reduce tension on the wound site,105 other mechanisms for the scar-reducing effects such as hydration of the stratum corneum,106,107 direct modification of extracellular matrix composition,108,109 and decreased TGF-β2 expression in scar fibroblasts110,111 have been proposed. Silicone gel sheeting placed over evolving hypertrophic and keloid scars led to reductions in scar size in 17 of 20 patients.98 Another study following 30 patients with scars of different types—including superficial scars, hypertrophic scars, and keloids—found that silicone gel application initiated a few days after wound closure led to improvements in scar grade after 6-month follow-up.99 Recently, more focused efforts to provide mechanical offloading during wound healing have demonstrated efficacy in both animal and human clinical trials in reducing fibrosis by simultaneously influencing multiple signaling pathways with

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Plastic and Reconstructive Surgery • March 2015 a mechanoreceptive element. A phase I clinical trial112 and a multicenter, randomized, controlled trial113 have demonstrated that stress-shielding of half of an abdominal incision leads to significant improvements in scar appearance compared with the contralateral side. In addition to scarring following primary surgery, mechanical offloading offers an opportunity to reduce scarring following scar revision surgery. The device (Neodyne Biosciences, Inc., Menlo Park, Calif.) significantly decreased scarring following revision surgery in a randomized controlled trial of 12 patients.114 Collectively, these findings suggest that the pathways involved in fibrosis are too complex to currently target pharmacologically, and that device approaches appear highly promising for reducing scarring following primary and revision surgery.

CONCLUSIONS Our current understanding of wound healing and cell subpopulations within the skin has permitted the development of scar-reducing therapies with only marginal effectiveness. Although differences between wound healing in fetal and adult skin have been well-characterized, the cellular mechanisms governing fetal skin regeneration remain poorly understood. This is not surprising considering that we are only now beginning to precisely characterize distinct functional lineages in the skin. Achieving scarless wound healing in the adult will require not only an understanding of signaling molecules and growth factors but also a thorough understanding of lineage-specific cellular origin and function during both fetal and adult stages. Michael T. Longaker, M.D., M.B.A. Department of Surgery Division of Plastic and Reconstructive Surgery Stanford University School of Medicine 257 Campus Drive Stanford, Calif. 94305-5148 [email protected] Geoffrey C. Gurtner, M.D. Department of Surgery Division of Plastic and Reconstructive Surgery Stanford University School of Medicine 257 Campus Drive Stanford, Calif. 94305-5148 [email protected]

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