Fibroblasts and Wound Healing: an Update

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Editorial
Fibroblasts and wound healing: an update
Heather E desJardins-Park1, Deshka S Foster1& Michael T Longaker*,1
1Hagey Laboratory for Pediatric Regenerative Medicine, Department of Surgery, Division of Plastic Surgery, Stanford University
School of Medicine, Stanford, CA 94305, USA
*Author for correspondence: longaker@stanford.edu
“Increased comprehension of dermal fibroblast heterogeneity may yield both mechanistic
insights into existing therapies and inspiration for novel therapeutics targeting specific cell
populations in wound healing and scarring”
First draft submitted: 30 May 2018; Accepted for publication: 13 June 2018; Published online:
31 July 2018
Keywords: broblast •injury •regeneration •scar •stem cell •therapy •wound healing
Wound healing and regenerative medicine are intimately linked. While any dermal wound in an adult human, even
if treated, will result in scarring [1], the ‘holy grail’ of wound healing is ‘scarless wound healing’: wound repair via
the regeneration of functional, native tissue. Scarring and pathological wound healing states, such as hypertrophic
scarring and keloids, represent an enormous clinical and financial burden on our healthcare system. Unfortunately,
there are few truly effective therapies that hasten healing while reducing scar burden.
In the setting of skin injury, wound healing following hemostasis occurs in three overlapping stages: inflammation,
proliferation and remodeling [2]. Fibroblasts are critical in all three phases, playing a key role in the deposition of
extracellular matrix (ECM) components, wound contraction and remodeling of new ECM. Since our previous
review [3], recent work continues to show the striking heterogeneity of skin fibroblasts. The concept that dermal
fibroblasts represent multiple distinct subpopulations is an important advancement in our understanding of skin
pathophysiology and serves as a new perspective from which the innovation of novel wound therapies may be
possible. Herein, we will discuss recent advancements in the understanding of fibroblast heterogeneity as it pertains
to cutaneous wound healing and relevant developments in clinical wound therapies.
Dening broblast subpopulations
Recent basic science research in wound healing has increased its focus on understanding the lineages, identities
and roles of fibroblasts in various tissues. Rigorous characterization of dermal fibroblast heterogeneity based on cell
surface markers has proved challenging, as cell surface marker expression is highly variable and no single marker
identifies this cell type [4]. However, in recent years, distinct populations of dermal fibroblasts have been elucidated.
In 2013, Driskell et al. showed that dermal fibroblasts arise from two different lineages. The upper dermal lineage
is involved with hair follicles, while the lower synthesizes ECM and engages with adipocytes [5]. Notably, the lower
lineage was found to be largely responsible for dermal repair following wounding, explaining why scar tissue in
humans is particularly ECM-rich and lacks hair follicles [5].
In 2015, Rinkevich et al. described the discovery of a ‘scarring fibroblast’ lineage responsible for depositing
the vast majority of dorsal scar tissue in mice [6]. These cells are defined by lineage positivity for the homeobox
transcription factor EN1. The authors found that these same cells could be reliably identified via expression of the
marker CD26 and that ablation of these cells reduced scarring, although this also delayed wound healing. More
recently, Hu et al. found that PRRX1 demarcates the ventral lineage of murine scar-producing fibroblasts [7].
Novel insights into stem cell contributions to wound healing
In order to fully understand fibroblast heterogeneity, these cells’ lineages must be characterized. Plikus et al.showed
that during wound healing, adipoyctes can be generated from myofibroblasts (activated fibroblasts involved in
wound contraction), suggesting significant lineage plasticity in what were previously understood to be terminally
Regen. Med. (2018) 13(5), 491–495 ISSN 1746-0751 49110.2217/rme-2018-0073 C
2018 Future Medicine Ltd

Editorial desJardins-Park, Foster & Longaker
Topical application
of growth factors
(e.g., PDGF, EGF)
can increase
broblast
proliferation and
accelerate wound
closure
Topical application
of cell-based skin
substitutes
(e.g., Grax®) protect
the wound site
and provide
healing factors
Targeting broblast
mechnotransduction
decreases brosis in
scarring
Red blood cell Platelet Fibrin Neutrophil Cytokines
Macrophage Fibroblast ECM Adipocyte
Figure 1. Wound healing pathophysiology and novel therapeutics for wound healing. An overview of cutaneous
wound healing pathophysiology with a summary of recent wound healing therapeutics of note (discussed in depth in
the text). (A–C) A review of cutaneous wound healing pathophysiology.(A)During the rst stages of wound healing,
platelets are recruited to the open wound and deposit brin (which serves as a preliminary extracellular matrix) to
arrest bleeding. (B) During the next stages of wound healing, immune cells including neutrophils followed by
macrophages are recruited to the wound and clear dead tissue and debris in preparation for healing. New blood
vessels sprout around the site. Fibroblasts are recruited to the site in anticipation of scar formation. Keratinocytes
begin to migrate to cover the cutaneous wound surface. (C) Finally, during the remodeling phases of wound healing,
the keratinocytes have covered the site. Below the broblasts deposit new extracellular matrix replacing the brin
plug, which is then remodeled to form the nal scar. New blood vessels are pruned and nerves begin to regenerate to
the site. (D–F) Novel therapeutics for wound healing. (D) Growth factors such as PDGFs can be provided directly to the
wound to stimulate broblast proliferation and accelerate wound closure. (E) Cell-based skin substitutes
(e.g., Grax R
, Osiris Therapeutics, MD, USA) can be applied directly to the wound site to protect it and directly
provide factors including cells involved in wound healing. (F) Fibroblast mechanotransduction plays a role in
stimulating scar production. Treatments that target broblast mechanotransduction, such as modulation of the FAK
pathway, are being explored in the wound setting to decrease scar brosis and potentially improve cosmesis. Key for
cell types used in illustrations provided in box at bottom.
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Fibroblasts & wound healing: an update Editorial
differentiated fibroblasts [8].Geet al. also demonstrated significant lineage infidelity among cells involved in wound
healing (including epidermal and hair follicle cells). They showed that this lineage infidelity is induced by stress-
response-related transcription factors and is transient in healing but persistent in the setting of cancer [9].Inthis
regard, cancer cells can co-opt regenerative mechanisms seen in healing.
Such examples of lineage plasticity are dismantling the concept of distinct populations of stem cells that supply
each cell type in a wound. For example, it is possible that under conditions of homeostasis, epidermal and hair
follicle fibroblast lineages are distinct but under stress conditions this distinction is blurred [9]. These findings might
explain why response to tissue injury can be highly variable across different individuals and pathological states.
Fibroblast heterogeneity across pathological wound healing states
Human wound healing may be viewed as a spectrum, with typical scar formation representing the ‘normal’
phenotype; chronic wounds at one extreme and hyperproliferative scarring and even keloids at the other. Previous
studies have begun to investigate mechanisms by which fibroblast dysfunction could contribute to pathological
wound healing states.
Multiple studies have shown that fibroblasts from chronic nonhealing wounds display abnormal phenotypes,
including decreased proliferation, early senescence and altered patterns of cytokine release [10], as well as abnormal
MMP and TIMP activity [11]. Conversely, keloid fibroblasts are known to exhibit increased proliferation and
decreased apoptosis [12], and it has been suggested that keloid fibroblasts induce an abnormal phenotype in
surrounding quiescent fibroblasts via paracrine signaling [13], explaining the observation that keloids outgrow the
initial wound boundaries. Early observations, such as that made by Wang et al. that hypertrophic scar fibroblasts
most closely resemble fibroblasts from deeper dermal layers [14], have alluded to the potential significance of different
fibroblast subgroups in pathological healing states. However, this complex biology and its clinical implications have
yet to be fully elucidated. We hope that research efforts will continue to explore these pathologies through the lens
of fibroblast heterogeneity in the skin, to shed new light on the mechanisms behind scarring disease.
Fibroblast-focused therapeutics
There have been many wound therapeutic innovations in scar modulation since our previous review. We will limit
our discussion to those most relevant to fibroblasts, including therapies involving the delivery of viable fibroblasts
to the wound site and manipulation of fibroblast behavior.
Growth factor therapies
Several growth factors, including PDGF, FGF and TGF-β, are known to stimulate fibroblast division, activity and/or
differentiation. But to date, only one growth factor, a recombinant human PDGF formulation (REGRANEX Gel,
Smith & Nephew, London, UK), has been approved for chronic wounds. Other growth factors including FGF and
TGF-βhave failed to robustly demonstrate improved wound healing in human patients [15,16]. The general failure
of growth factor therapies is likely due to multiple limitations including short half-life following delivery. While
attempts have been made to integrate biomaterials for controlled growth factor release [17], major breakthroughs
have yet to make their way into clinical practice.
Cell-based therapies
Effective wound treatments demand an agent that reflects the complex in vivo milieu of cell types and growth
factors. One approach has been to deliver viable allogeneic cells, including fibroblasts, to the wound site. These
cells do not persist indefinitely [18], but instead serve as a source of growth factors and cytokines to support the
function of the patient’s own cells.
Cell-based therapies are most often used for chronic wounds, perhaps because as previously mentioned these
patients’ own cells may be incompetent for wound healing. Cell-based therapies approved for use in wounds
incorporate a varying range of cells, from fibroblasts only to fibroblasts plus keratinocytes, or even fully cryopreserved
skin [19,20,21]. A more recent development in cell-based wound therapies, Grafix (Osiris Therapeutics, MD, USA),
consists of cryopreserved placental tissue including placental ECM and fibroblasts [22].
While all of these products are used clinically for a large variety of wound types, their full mechanism of
action remains unknown and in many cases their efficacy has not been robustly established in vivo [23].Increased
characterization of the different skin cell populations may enable the development of increasingly effective cell-based
treatments.
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Editorial desJardins-Park, Foster & Longaker
Novel directions: targeting regeneration & broblast mechanotransduction
In recent years, growing mechanistic understanding of fibroblast function has led to therapeutically promising
discoveries. Wnt signaling is known to be critical for skin differentiation and Wnt-responsive dermal fibroblasts
play a key role [24]. As such, Wnt-3a and FGF-9 (which triggers and amplifies Wnt expression and activation in
dermal fibroblasts) are being developed as therapies with the potential to achieve scarless healing with features of
regeneration [25,26,27].
It has also been established that mechanical forces play a role in the development of pathological scars [28].
Mechanotransduction in wound fibroblasts occurs via focal adhesion complexes, which link the ECM to the
intracellular cytoskeleton [29]. In 2011, Wong et al. demonstrated that FAK activation occurs following cutaneous
injury and that fibroblast-specific FAK inhibition decreases scarring in mice [30]. These results suggest that fibroblast
mechanotransduction may be a rich target for novel antiscarring wound therapies, and elucidating the molecular
pathways linking mechanotransduction and fibrosis is an active area of current research. Figure 1 illustrates the
stages of wound healing and fibroblast-related wound therapies.
Conclusion & future perspective
Understanding dermal fibroblast heterogeneity is a lofty but important goal. Major advancements have been made
in the last several years to expand our knowledge of the different populations, signaling pathways and cellular niches
of the diverse fibroblasts in mouse and human skin. Further work is needed to fully elucidate the contributions of
different dermal fibroblast lineages to wound healing, characterize the most specific markers for each cell population
and translate these populations from mice to humans. Increased comprehension of dermal fibroblast heterogeneity
may yield both mechanistic insights into existing therapies and inspiration for novel therapeutics targeting specific
cell populations in wound healing and scarring. We anticipate that in the near future, the field of wound healing
therapeutics will expand to mirror our growing understanding of the diversity of cells involved in this biology.
Financial & competing interests disclosure
MT Longaker is a co-founder of, has an equity position in, and serves on the board of Neodyne Biosciences, Inc., a startup company
which developed a device to reduce mechanical tension on wounds to minimize post-operative scarring. The authors have no other
relevant afliations or nancial involvement with any organization or entity with a nancial interest in or nancial conict with
the subject matter or materials discussed in the manuscript apart from those disclosed. This includes employment, consultancies,
honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.
No writing assistance was utilized in the production of this manuscript.
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