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Normal versus hypertrophic wound healing

Butzelaar, L.

2015

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citation for published version (APA)Butzelaar, L. (2015). Normal versus hypertrophic wound healing: Does early inflammation determine finaloutcome?.

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Download date: 11. Apr. 2021

Chapter 1

General introduction and outine of the thesis

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Introduction

Skin anatomy

Skin covers the entire body and provides a barrier between the body and the outside world through its mechanical as well as immunological properties. The skin protects the body form dangers such as dehydration, UV radiation and pathogens. The skin consists of two layers: the epidermis and the dermis. The epidermis consists of keratinocytes, but also contains Langerhans cells and melanocytes. The dermis is composed of extracellular ma-trix (ECM), blood vessels, ECM producing fibroblasts and several types of immune cells. The dermis also contains skin appendices and nerve endings (figure 1). ECM consists of proteoglycans (glycosylated proteins) and several fibrous proteins including collagen and elastin1. Collagen, elastin and most of the other proteins are produced by fibroblasts1,2. Af-ter assembly, collagen undergoes some changes in order to mature and stabilize. The triple helix is first stabilized by means of hydrogen bonds3. Subsequently, several types of cross-links are formed within the triple helices and between triple helices4,5. Cross-links are also formed in elastin fibres4,5. Collagen and elastin fibre content and composition (including cross-linking) and several other proteins including proteoglycans determine the mechanical properties of skin6.

Figure 1: anatomy of the skin Adapted from Nestle OF, Di Meglio P, Qin JZ, Nickoloff BJ (2009)17.

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Skin resident immune cells include dermal Tcells and macrophages (Mφ) and epi-dermal Langerhans cells7,8. These cells are important for immune surveillance, but also prevent the immune system from reacting to harmless agents7.

Cutaneous wound healing

When the skin is damaged, its barrier function is compromised. Therefore, swift sealing and successive repair of the defect are mandatory. This is achieved through wound hea-ling, where the defect is not resolved by regeneration, but rather by deposition of fibrotic tissue, resulting in scar formation9. Fibrosis occurs in many other tissues of the human body besides skin, such as lung and liver9. The complex process of wound healing can be subdivided in roughly four successive, but partially overlapping phases: the hemostasis and inflammation phase, the proliferation phase and the remodeling phase10. In order to mini-mize blood loss, hemostasis is achieved directly after wounding through the formation of a fibrin clot by activated platelets and the coagulation cascade10. Both activated endothelium and activated platelets kick start inflammation by secreting a variety of inflammatory and chemotactic cytokines10. Neutrophils are the first inflammatory cells to arrive in the wound in response to the chemotactic stimuli1. These cells scavenge the wound for pathogens and produce cytokines that attract Mφ1. Then, the neutrophils undergo apoptosis as Mφ start to enter the wound area after approximately two days1. Mφ dispose the wound of dead cells, including neutrophils, resolve the inflammatory phase and direct wound healing towards the granulation phase1. After three days, re-epithelialization and neovascularization begin to occur and fibroblasts start to deposit immature (type III) collagen to fill the defect1. Sub-sequently, the newly formed ECM is remodeled to become stronger and more organized1. The remodeling phase lasts until approximately one year after trauma (figure 2)1.

Excessive cutaneous wound healing

Derailment of the wound healing cascade results in deposition of excess scar tissue. Two forms of excessive skin scarring exist: keloid- and hypertrophic scars (HTS). This thesis focuses mainly on HTS formation. These types of scars frequently develop as a result of burn wounds, but also after surgical trauma with incidences varying from 40 to 70% after surgical trauma and 91% after burn trauma10. HTS is defined as a scar which is raised above skin level, but remains within the boundaries of the original wound, while keloids exceed the wound margins10. Other clinical characteristics of HTS include esthetically un-desirable features such as erythema and rigidness and co-morbidity such as pain, itch and diminished range of motion when positioned over a joint11. Histological properties of HTS

12

Introduction

include increased amounts of collagen compared to normal scars and myofibroblasts which persist in mature HTS as opposed to normal mature scars10.

Figure 2: normal wound healing First, hemostasis occurs and immediately starts inflammation, followed by proliferation after approximately 3 days and remodeling which lasts for months.

Regarding possible mechanisms involved in excessive scarring, several risk factors for HTS formation have been suggested, comprising both systemic and local factors. Systemic factors include ethnic background and possibly genetic predisposition; an example of local factors is mechanical force in the form of stretch12-15.

With respect to the wound healing cascade itself, the current opinion states that HTS results from excessive and/or prolonged inflammation, overabundant proliferation and ab-normal remodeling10. Alterations of the inflammatory response are thought to play a major role in this derailment of wound healing11. Indeed, altered immune cell populations have been observed in HTS before. For example, HTS contain more Langerhans cells, which is thought to contribute to excessive inflammation in HTS formation11. Mahdavian Delavary and colleagues proposed that a prolonged inflammatory phase in hypertrophic wound healing causes Mφ to accumulate and induce overabundant ECM production8. Although Niessen and colleagues did not observe differences in Mφ numbers between HTS and nor-mal human scars of three and 12 months old, this observation does not exclude increased Mφ numbers during earlier stages of hypertrophic wound healing11.

Despite extensive research on the subject, the exact cause of HTS formation remains unknown and to date no optimal treatment is available. Even though literature suggests that the immune system plays an important role, it is unknown whether systemic or local

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Chapter 1

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immunological predisposing factors (or both) induce or contribute to HTS formation. Next to that, just a few researchers have studied human hypertrophic wound healing as early as two weeks after injury, but most studies focus on later time points in the process of hyper-trophic scar formation. Thus, the exact time of onset of the derailment of wound healing leading to HTS formation has not been established yet.

Outline of the thesis

Several risk factors for HTS formation have been described of which ethnic background is probably the best known risk factor. Chapter 2 discusses currently known risk factors for HTS formation, their levels of evidence and their influence on the individual phases of the wound healing process. In addition, risk factors for HTS are studied in chapter 3 in patients who received presternal incisions for the purpose of open heart surgery. Most open heart surgeries are performed using extra corporal circulation (ECC; heart lung machi-ne). Surgery as well as ECC induces a systemic inflammatory response16. This systemic inflammatory response may affect wound outcome, where a stronger response would result in HTS more frequently. The effect of different types of ECC on scar outcome is examined in chapter 4.

Besides in skin, fibrosis occurs in many other tissues, for example in coronary arte-ries9,17. HTS and coronary sclerosis are both forms of excessive fibrosis and have several features in common17. The possible relation of coronary sclerosis with HTS formation is examined in chapter 5.

Next to several risk factors, the immune response is thought to play an important role in HTS formation, but the precise mechanisms and moment of derailment of the wound healing process are unknown. In order to elucidate whether early inflammation could play a role, the immune response of early hypertrophic wound healing versus normal wound healing on genetic-, protein- and cellular level is examined in chapter 6. No research has been performed previously on alterations of the very early processes of wound healing in association with HTS formation.

With reference to local factors, skin of different anatomic sites is subjected to diffe-rent influences, for example UV radiation, pathogens, but also mechanical forces such as stretch. Since excessive scarring occurs more frequently in certain anatomic locations, histological and molecular properties of skin of different anatomic sites and predilection sites for excessive scarring are studied in chapter 715.

A general discussion of the results is provided in chapter 8.

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Introduction

References

1. Broughton G 2nd, Janis JE, Attinger CE. The basic science of wound healing. Plast Reconstr Surg. 2006;117:12S-34S.

2. RinnJL, Wang JK, Liu H, Montgomery K, van de Rijn M, Chang HY. A systems biology approach to anatomic diversity of skin. J Invest Dermatol. 2008;128:776-82.

3. van der Slot-Verhoeven AJ (2005). Telopeptide lysyl hydroxylase: a novel player in the field of fibrosis (Unpublished doctoral dissertation). Univer-sity of Leiden, The Netherlands.

4. Eyre DR, Paz MA, Gallop PM. Cross-lin-king in collagen and elastin. Annu Rev Biochem. 1984;53:717-48.

5. Muiznieks LD, Keeley FW. Molecular assembly and mechanical properties of the extracellular ma-trix: A fibrous protein perspective. Biochem Biophys Acta. 2013;1832:866-75

6. Kazlouskaya V, Malhotra S, Lambe J, Idriss MH, Elston D, Andres C. The utility of elastic Ver-hoeff-Van Gieson staining in dermatopathology. J Cutan Pathol. 2013;40:211-25.

7. Seneschal J, Clark RA, Gehad A, Baecher-Allan CM, Kupper TS. Human epidermal Langerhans cells maintain immune homeostasis in skin by activating skin resident regulatory T cells. Immunity. 2012;36:873-84.

8. Mahdavian Delavary B, van der Veer WM, van Egmond M, Niessen FB, Beelen RH. Macrop-hages in skin injury and repair. Immunobiology. 2011;216:753-62.

9. Stramer BM, Mori R, Martin P. The inflam-mation-fibrosis link? A Jekyll and Hyde rol for blood cells during wound repair. J Invest Dermatol. 2007;127:1009-17.

10. Gauglitz GG, Korting HC, Pavicic T, Ruzicka T, Jeschke MG. Hypertrophic scarring and keloids: pathomechanisms and current and emerging treat-ment strategies. Mol Med. 2011;17:113-25.

11. Niessen FB, Schalkwijk J, Vos H, Timens W. Hypertrophic scar formation is associated with an increased number of epidermal Langerhans cells. J Pathol. 2004;202:121-9.

12. Bayat A, Bock O, Mrowietz U, Ollier WE, Fer-guson MW. Genetic susceptibility to keloid disease and hypertrophic scarring: transforming growth factor beta 1 common polymorphisms and plasma levels. Plast Reconstr Surg. 2003;111:535-43.

13. Ketchum LD, Cohen IK, Masters FW. Hyper-trophic scars and keloids: A collective review. Plast Reconstr Surg. 1974;53:140-54.

14. Quatresooz P, Hermanns JF, Paquet P, Piérard GE. Mechanobiology and force transduction in scars developed in darker skin types. Skin Res Technol. 2006;12:279-82.

15. Verhaegen PD, van Zuijlen PP, Pennings NM, van Marle J, Niessen FB, van der Horst CM et al. Differences in collagen architecture between keloid, hypertrophic scar, normotrophic scar, and normal skin: An objective histopathological analysis. Wound Repair Regen. 2009;17:649-56.

16. Paparella D, Yau TM, Young E. Cardiopulmo-nary bypass induced inflammation: pathophysiology and treatment. An update. Eur J Cardiothorac Surg. 2002;21:232-44.

17. Stary HC, Chandler AB, Dinsmore RE, Fuster V, Glagov S, Insull W jr et al. A definition of advan-ced types of atherosclerotic lesions and a histologi-cal classification of atherosclerosis. A report from the Committee on Vascular Lesions of the Council on Arteriosclerosis, Amercian Heart Association. Circulation. 1995;92(5):1355-74.

18. Nestle OF, Di Meglio P, Qin JZ, Nickoloff BJ. Skin immune sentinels in health and disease. Nat Rev Immunol. 2009;9:679-91.

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Oude Rijn, Leiden; adventurous girl on pedestian bridge