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Wound care SCIENCE Clinical PRACTICEWound DEVELOPMENT SCIENCE The role of the fibroblast in wound contraction and healing

Although wound contraction appears to be organised by fibroblast cells within granulation , the interaction between fibroblasts and the is not fully understood. There are two dichotomous theories: the contraction theory and the fibroblast (cell traction) theory. This article discusses each theory and suggests that both may be correct but may occur at different times along the healing pathway. Further investigation into the mechanism of foetal scarless healing may uncover a way of manipulating wound contraction leading to faster healing, reduced scarring and elimination of fibrotic diseases.

Sue Porter

(Martin, 1997), e.g a first-trimester diminishes function and appearance. KEY WORDS foetus will heal cutaneous wounds In an animal model, contraction has without scarring and intra-oral wounds been shown to begin in an acute Wound contraction can also contract and form which wound about 4–5 days after wounding, are considerably smaller than the peaking at 14 days, and progressing Extracellular matrix original wound. at roughly 0.6–0.75mm per day, Myofibroblasts depending on the shape of the wound Scarless Proliferation is characterised by and tissue type (Lawrence, 1998). , formation of a matrix of In humans, sacral wounds contract fibronectin, synthesis, rapidly because the skin is loose and proteoglycans, subcutaneous fat is present. In contrast, ealing takes place in four well- and other proteins, epithelialisation pre-tibial wounds that are stretched defined but overlapping stages: and contraction (Martin, 1997). The with the closely tethered to Hhaemostasis; inflammation; way that a wound contracts has been underlying tissue heal slowly, mostly by proliferation; and remodelling. Each subject to debate. This paper focuses epithelialisation (Butterworth, 1993). stage forms a continuum which relies on the important role of the fibroblast Wound contraction stops usually by upon a delicate balance of molecular and the debate surrounding its role in 4–6 months (Rudolph et al, 1977). signs involving an intricate series of wound contraction and closure. ordered, inter-related events that Fibroblasts are cells that are include , mitosis, neo- Wound contraction: an overview responsible for most collagen and vascular synthesis of new extracellular According to a review by Tejero- elastin synthesis and organisation matrix and the formation of a . Trujeque (2001) contraction occurs of the ECM components. They are Remodelling continues for up to 18 when the wound edges move towards indispensable in determining how months (Majno and Joris, 1996). each other in a centripetal fashion well the wound will ultimately heal. thus reducing the wound’s dimensions. Fibroblasts synthesise a fibronectin-rich Most repairs are distinct from Contraction involves a dynamic ECM as soon as they enter a wound the surrounding tissues and never process where cells organise their haematoma (Mehendale and Martin, achieve more than 80% of the tensile surrounding tissue matrix to reduce 2001). Together with abundant amounts strength of unwounded skin (Waldrop normal healing time by shrinking of the hyaluronic and Doughty, 2000). However, some the amount of extracellular matrix acid, fibronectin — a glycoprotein wounds are capable of regenerating (ECM) that needs to be produced — may serve as a channel to guide tissue without contraction and scar (Jones et al, 2004). In many respects, fibroblasts into the provisional matrix formation, leading to a perfect repair wound contraction is beneficial as it of the fibrin clot (Greiling and Clark, that matches the surrounding tissue can significantly reduce healing time 1997). appears in the body because less needs in several isoforms. Soluble fibronectin Sue Porter is Sister, Plastic Surgery Dressing Clinic, to be produced to replace tissue loss in plasma enhances blood clotting, Queen Victoria NHS Foundation Trust, East Grinstead, (Calvin, 1988). The result, however, is wound healing and phagocytosis, while West Sussex scarring, which is less desirable as it all other forms are attached to the cell

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fibrils associated with them. This is the fibroblast theory.

Myofibroblast theory It has been proposed that myofibroblasts exhibit morphological traits of both fibroblasts and smooth muscle cells (Gabbiani et al, 1971; Majno et al, 1971). The differentiated cells are presumed to be responsible for the process of wound contraction via the movement of parallel smooth muscle contracting the cell in the same way as muscle would. Myofibroblasts are rich in cytoplasmic microfilaments (actin-rich stress fibres) and the expression of alpha smooth muscle actin (αSMA), also termed stress fibres, contract the myfibroblast in a muscle-type action. Additionally, myofibroblasts exhibit cell-to-cell and cell-to-matrix (fibronexus) links

Science Photo Library joined together by gap junctions and desmosomes which are thought to be Coloured transmission electron micrograph of an elastin fibre (diagonal red band) in human connective necessary for fibroblasts to contract tissue. The elastin fibre is in contact with a fibroblast cell (upper left, yellow/blue), which is probably in a synchronised fashion (Darby et synthesising the fibre. Nearby are collagen fibres (speckled green). al, 1990). These contacts provide the force that pulls collagen 1 and III fibrils surface and are secreted into the ECM despite the formation of new collagen in the ECM towards the body of the as highly insoluble fibronectin fibrils being severely compromised. Therefore cell, resulting in a reduction in the (Alberts et al, 2002). contraction was considered to be the area of granulation tissue. The force result of forces generated by cells in the that makes the entire wound contract In fibroblasts, fibronectin fibrils are central part of the granulation tissue. is caused by many synchronised linked to integrin α2β1 transmembrane However, the authors admitted that half myofibroblast cells ‘pulling’ up tissue proteins in areas called fibrillar the guinea pigs were on the deficient and ‘fixing’ it in place with collagen adhesions that keep the cell attached diet for only a short time; therefore it (Ryan et al, 1974). Myofibroblasts to the matrix. On the cell surface the was possible that their body reserves then undergo fibronectin fibrils are elongated and of vitamin C were not depleted and (apoptosis) as the wound closes are under tension and they usually this could affect the outcome. (Guinea (Desmouliere et al, 1995). coordinate with actin stress fibres pigs have a 5–7 year lifespan, are unable within the cell. ECM signals can alter the to store ascorbic acid and will develop Myofibroblasts are thought to arise intracellular actin and thus symptoms of scurvy in 3–4 weeks on a from the conversion of the wound the tension on the fibrils. The migrating deficient diet.) fibroblast phenotype rather than from fibroblasts change their phenotype the smooth muscle cell (Darby et al, to a profibrotic type, influenced by Currently, there are two leading, 1990). How quickly they change their transforming -β1 in order but contradictory paradigms which phenotype depends upon the degree of to synthesise and then rework a new suggest a possible explanation of how mechanical tension and forces resisting collagen-rich matrix (Welch et al, 1990). fibroblasts generate and spread the the contraction. This was demonstrated force required to contract a wound. by Grinnell (1994) in his studies of Collagen is the predominant Gabbiani et al (1971) postulated that fibroblasts on floating or anchored extracellular component of the dermis. a specialised fibroblast which behaves collagen matrices. He noted that Wound contraction was initially thought similarly to a smooth muscle cell is fibroblasts changed their morphology to be caused by newly deposited responsible for ECM contraction. by developing fibronexus junctions and collagen fibres. Abercrombie et al This is the myofibroblast theory. In prominent stress fibres when added (1956) devised a method to test the contrast, Harris et al (1981) proposed into a tethered matrix. Conversely, theory using guinea pigs that were that fibroblast cells exert traction fibroblasts seeded into a free-floating given a diet deficient in ascorbic acid for as they move across the ECM. Cell collagen matrix retained a similar 15 days. Wound contraction occurred locomotion forces realign the collagen phenotype to a wound fibroblast.

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Although the theory and FPCLs continue to add to our forms the basis of current teaching it The ability of the foetus understanding of cell biology. remains speculative and controversial. to heal cutaneous wounds According to Ehrlich and Rajaratnam without scarring has McGee et al (1992) advises caution (1990) the idea has many histological attracted significant as of cells including studies to support it, but there is little interest. However, evidence morphology and gene expression physiological evidence that shows that exists that not all foetal can be induced in culture. Therefore, myofibroblast cells contract or that it can be argued that myofibroblasts the cell reduces granulation tissue as wounds heal scarlessly may be a phenomena inadvertently it contracts or that it organises and the underlying created by the culture or the scientist. collagen fibres. mechanisms remain Ehrlich (1988) has demonstrated that incompletely understood collagen matrices will contract without theory (Ferguson et al, 1996). myofibroblasts and will also contract In contrast to the myofibroblast when keratinocytes are substituted for theory, Harris et al (1981) proposed fibroblasts. Hembry et al (1986) used that wound contraction is achieved conclusions. Using an acute wound mice with tight skin to demonstrate that by many individual fibroblasts exerting model, Berry et al (1998) postulated wound contraction occurred without traction forces across the centre that differences may exist between myofibroblasts and proposed that their of the matrix. Fibroblasts exert a animal and human wound contraction function is to apply tension or maintain traction force like a treadmill on the because the behaviour of fibroblasts the new balance that has been created. collagen fibrils in the ECM which they and myofibroblasts differ in their The results of animal studies are difficult rearrange and compact, thus reducing connection with collagen. to compare as a variety of animals with its size. During cell elongation and heals predominantly by formation of dissimilar body weights have been used. spread across the ECM, shearing new dermis and re-epithelialisation, The size and shape of wound in ratio forces exert traction and thus cause with wound contraction representing a to the animal’s size is important as is the wound to contract. According small fraction of the healing process in the technique of positioning the animal to this theory, fibroblasts neither humans compared with other animals and measuring the area of the wound shorten in length, nor synchronise (Moulin et al, 2000). (Cross et al, 1995). their efforts, they are randomly orientated and cell-to-cell contact Tomasek et al (1992) proposed Fibroblasts appear in the wound is rare. The theory gained support that FPCL contraction was mediated after 2–3 days, yet myofibroblasts from Ehrlich (1988) when he by myofibroblasts, however, the FPCL, predominate at day 12 when wound found no difference in contraction which was first developed by Bell contraction is almost 80% complete in a fibroblast-populated collagen et al (1981), can closely mimic the (Darby et al, 1990; Desmouliere et al, lattice (FPCL) when biochemical and cellular aspects of 1995). Darby et al used a model were added or removed from the normal skin but does not contain to show an 85.7% wound reduction lattice. He concluded that collagen blood vessels or react the same, detected by electron micrograph, fibre reorganisation is a result of for instance it will not reproduce however, it should be noted that fibroblastic locomotion (Ehrlich and the inflammatory response. FPCLs high resolution imaging can pick up Rajaratnam, 1990). also lack the full complement of sub-cellular artefacts in addition signals and relationship variables to meaningful phenomena. Hence, Both theories have many studies between the complete range of all McGrath and Hundahl (1982) noted to support them, but they are often possible reactions in vivo that may some years earlier that a myofibroblast in vitro and involve animal models, influence cell behaviour (Kuhn et al, phenotype may be due to particular implants and collagen lattices. 2000). Conversely, for some types of culture conditions and appear as a Therefore considerable caution experimental questions it is useful consequence of contraction rather needs to be exercised when drawing to isolate a single variable of interest than its cause.

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Ehrlich et al (1994) studied αSMA nears full term. Wound size and the in and hypertrophic scars and amount of tissue damaged affects the Key Points initially found that only the nodules in extent of inflammation and hypertrophic scars were positive for response and thus scarless repair. A 8 Repair and healing can be myofibroblasts expressing αSMA. When splinter or injection site usually heals broadly described in four well- placed in culture, both hypertrophic without a scar but a full thickness defined but overlapping stages and keloid scars produced similar injury will result in a clumsy repair of haemostasis, , amounts of αSMA suggesting the with disorganised collagen and a proliferation and remodelling. cultured environment influences dense mesenchymal scar (Ferguson 8 The repair is distinct from the expression of this protein. et al, 1996). However, cases are the surrounding tissues and This may demonstrate the changes documented where infants are never achieves more than when cells are removed from their born with hypodermic needle scars 80% of the tensile strength of normal environment highlighting the following prenatal diagnostic sampling unwounded skin. importance of studying ECM and cell- (Paller and Mancini, 2005). The ability to-cell interactions in vivo (McGee et of the early gestation foetus to heal 8 Contraction involves a dynamic al, 1992). cutaneous injury rapidly and scarlessly process where cells organise appears to be intrinsic to foetal their surrounding tissue matrix Scarless wound healing cells, ECM, cytokine expression and to effect a reduction on normal The ability of the foetus to heal healing time by shrinking the amount of extracellular matrix. cutaneous wounds without scarring The relationship between has attracted significant interest. 8 In many respects, wound However, evidence exists that not the ECM and fibroblasts and how they interact contraction is desirable as it all foetal wounds heal scarlessly and can significantly reduce healing the underlying mechanisms remain to effect contraction of time by reducing the amount of incompletely understood (Ferguson granulation tissue is still granulation tissue that must be et al, 1996). There are a number of little understood... The produced to replace tissue loss, factors that influence certain foetal study of adult tissues but does result in scarring. wounds to heal with little or no scarring and fibroblasts appear to have that retain the ability to 8 Two theories about the a crucial role in this. In common with heal quickly with minimal mechanism of wound intra-oral fibroblasts, foetal fibroblasts scarring, such as oral contraction exist: the are phenotypically distinct, migrate at mucosa, and the study of myofibroblast theory and the a faster rate and are more efficient fibroblast theory. There have scarless wound contraction been no definitive conclusions, at organising the ECM environment is developing in exciting (Stephens et al, 1996). Unlike adult because a suitably accurate fibroblasts, foetal fibroblasts produce directions. experimental model of human hyalonuric acid in consistent amounts wound healing does not exist. that does not decrease with greater ultimately, genetic control. Current cell density thus allowing for faster intensive research is focused upon migration, faster repair and cross- understanding the factors involved in linking of collagen. Estes et al (1994) an attempt to benefit the quality of ways and at different times along discovered that myofibroblasts in wound healing. the healing continuum. It seems foetal wounds in sheep were absent clear that the fibroblast is not the at 75 days gestation when cutaneous Conclusion only cell to produce contraction in wounds are expected to heal without There is a stumbling block to vitro, however, the movement of a scar; but present in increasing understanding wound contraction fibroblasts through all levels of the amounts at 100 and 120 days when because a suitably accurate model of ECM during the first days of wound scarring escalated. human wound healing does not exist. healing before αSMA expression Animal models, FPCLs and implants seems to be an essential requirement Gestational age appears crucial; all offer useful insights into targeted for the wound contraction process. Ferguson et al (1996) described areas of wound contraction but each Myofibroblasts that appear in the the gradual transition or continuum has their own particular weaknesses. wound as contraction progresses may from the ability of the early foetus to The meaning of experimental results be responsible for tensile strength and regenerate all of the dermal elements is difficult to transfer to complex scar formation. Moderating the rate leading to invisible repair, to later human wounds in vivo. The answer of apoptosis during wound closure gestation where a wound which is may lie somewhere between the may be important in controlling scar barely visible, to a small mark and two theories in that both influence formation and ultimately ameliorating finally an obvious scar as the foetus wound contraction in different pathological scars. It is evident that

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gene expression and proliferation of Desmouliere A, Redard M, Darby I, Gabbiani granulation tissue in vitro, similarity to cells is controlled by a mechanical G (1995) Apoptosis mediates the decrease smooth muscle. Science 173: 548–50 force exerted on the tissue, but the in cellularity dduring the transition between granulation tissue and scar. Am J Pathology Majno G, Joris I (1996) Cells, Tissues, and suggested signalling pathways remain 146(1): 56–66 Disease: Principles of General Pathology. guesswork. Blackwell Science, Oxford Ehrlich HP (1988) Wound closure: evidence of cooperation between fibroblasts and Martin P (1997) Wound healing: aiming The relationship between the ECM collagen matrix. Eye 2(Pt 2): 149–57 for perfect skin regeneration. 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