Wound healing is a highly orchestrated process that is evolutionarily conserved and involves complex interactions of extracellular matrix (ECM) molecules, soluble mediators, various resident cells, and infiltrating immune cells. The healing process consists of three consecutive yet overlapping phases: inflammation with blood clotting, new tissue formation and tissue remodeling [1-4]. The onset of wound healing occurs immediately upon injury by the formation of a hemostatic blood clot with a platelet plug that seals the wound and initiates inflammation (Figure 3.1a). Inflammatory cells are attracted to the wound site by signaling of the activated platelets to prevent infection and remove debris. In addition, these inflammatory cells release growth factors, cytokines and proteinases, which initiate the phase of tissue formation [1-4]. During this phase, migrating fibroblasts proliferate and produce large amounts of ECM. This new tissue is called granulation tissue because of the granular appearance of the numerous capillaries that are sprouting at the wound edge as angiogenesis induces the development of new vasculature [3, 4]. In the final phase, granulation tissue is replaced by mature scar tissue through matrix remodeling. Eventually, the inflammatory response abates and the cellularity normalizes [3, 4]. Impaired wound healing represents a serious cause of morbidity and mortality, primarily affecting aged individuals (i.e. ≥ 65 years) and diabetic or immunosuppressed patients, as well as patients receiving chemo- or radiotherapy [3, 6]. Chronic wounds are, by definition, wounds that have failed to progress through the stepwise process of physiologic healing. Instead, they are trapped in a phase of pathologic inflammation that causes a delayed, incomplete and uncoordinated healing process and impairs the restoration of anatomic and functional integrity of the tissue [6, 7]. At the opposite end of the spectrum, excessive healing causes fibrosis with overabundant collagen deposition and reduced remodeling. This fibrosis results in loss of the anatomical structure of the tissue and thus compromises its function [8].
38 | Introduction – Wound healing and cancer
Figure 3.1 - Wound healing versus invasive tumor growth. a. Normal tissues have a highly organized structure
with epithelial cells upon a basement membrane which separates them from the vascularized stromal compartment. Upon tissue damage, activated platelets form a hemostatic plug and release vasoactive factors that regulate
vasoconstriction and formation of the fibrin clot. The activated platelets also release chemotactic factors like TGF-β
and PDGF that initiate the formation of granulation tissue as well as the activation of fibroblasts and remodeling of the ECM. In addition, granulocytes and monocytes are recruited, and the venous network is restored. After re- epithelialization and healing of the wound, signaling subsides. b. Invasive carcinomas are less organized. The interaction of neoplastic cells with other cell types (i.e. mesenchymal, hematopoietic and lymphoid) causes the production of a chaotic vascular organization of blood and lymphatic vessels by angiogenesis an lymphangiogenesis. Neoplastic cells release cytokines and chemokines that are mitogenic and/or chemoattractants for granulocytes, mast cells, monocytes, macrophages, fibroblasts and endothelial cells. In return, these activated fibroblasts and inflammatory cells produce cytokines and chemokines that are mitogenic for neoplastic and endothelial cells.
3.1.2 “Cancers are wounds that do not heal”
Already in 1863 Rudolf Virchow proposed that malignant transformation represents the most severe complication of both impaired and excessive healing. He hypothesized that chronic irritation and previous injuries are a precondition for carcinogenesis [3]. The American pathologist Harald Dvorak then postulated in 1986 that “cancers are wounds that do not heal” [9]. He had recognized remarkable similarities between the granulation tissue of healing skin wounds and the composition of the stroma of malignant tumors. Therefore, he presumed that tumors activate the wound healing response of their host for the formation of the tumor stroma. Both postulations have been supported by numerous clinical observations and experimental studies [3].
Introduction – Wound healing and cancer | 39
Hallmark Cancer Wound healing
Proliferative signaling Sustained Transient
Evasion of growth suppression Sustained Transient
Cell migration With invasion and
metastasis
Without invasion and metastasis
Enabling replicative immortality Yes No
Angiogenesis Sustained Transient
Cell death Resisting Transient increase
Inflammation Sustained Transient
Table 3.1 - Comparison between the hallmarks of cancer and wound healing. Adapted from Arwert et al. [1].
Although many components of wound healing have been found in cancer, there are also important differences as illustrated in figure 3.1 and summarized in table 3.1. The most important difference is that wound healing is a self-limiting process whereas cancer is not [1]. This difference is caused by a dissimilar expression and activation of microenvironmental factors [1, 3]. Thus, the released growth factors, cytokines and chemokines show striking similarities between healing wounds and tumors [10]. However, these factors differ in the kinetics of their expression and become constitutively activated in solid tumors (Table 3.2) [1]. Furthermore, vessels in wounds and tumors are initially immature and leaky. This hyperpermeability allows the continuous release of plasma proteins and the deposition of a fibrin and fibronectin matrix, typical for most cancers and for healing wounds. This leakiness of the vessels with the deposition of the matrix is an acute and transient event in wound healing, whereas it is a persistent event in most cancers [3].
3.2 Angiogenesis and lymphangiogenesis as part of tumor vascularization
3.2.1 Introduction
The formation of new blood vessels is crucial for the supply of oxygen and nutrients to the healing tissue as well as to growing solid tumors. Neovascularization has an essential role in the growth of tumors beyond a diameter of 2mm and in metastasis [11]. Angiogenesis is the predominant approach of neovascularization in wound healing as well as in tumors [3]. Upon the onset of angiogenesis in healing wounds, also lymphangiogenesis is initiated in order to reconstruct the lymphatic vasculature. This process is particularly important for metastasis in tumorigenesis [12].
40 | Introduction – Wound healing and cancer
Functions in wounds Functions in cancer
Growth Factors
EGF family Epidermal and mesenchymal regeneration; accelerates wound healing
Cancer cell invasion, macrophage signaling and autocrine growth of tumor cells
FGF family Early angiogenesis, fibroblast proliferation and re-epithelialization via keratinocyte migration
Angiogenesis and fibroblast proliferation
TGFβ family Attracts neutrophils and macrophages, mediates ECM deposition, angiogenesis, epithelial cell migration and wound healing
Tumor development, tumor cell invasion and metastasis PDGF Attracts neutrophils and macrophages, and
mediates ECM deposition and angiogenesis.
Recruits inflammatory cell infiltration and mediates angiogenesis and lymphangiogenesis
VEGF Angiogenesis Tumor cell invasion and angiogenesis
Cytokines and chemokines
IL-1α/β Fibroblast and keratinocyte proliferation and neutrophil recruitment
Tumor cell proliferation, angiogenesis and inflammation
IL-6 Fibroblast proliferation and neutrophil recruitment
Tumor development, tumor cell invasion and metastasis
TNF Leukocyte infiltration Tumor promotion or suppression CSF1 Recruitment of macrophages and
re-epithelialization
Tumor cell invasion and migration MCP1 Macrophage recruitment, angiogenesis, re-
epithelialization and ECM production
Monocyte recruitment, tumor cell invasion and metastasis
CXCL1 Neutrophil infiltration, epithelial migration and neovascularization
Angiogenesis, invasion and migration CXCL2 Epithelial proliferation Recruits inflammatory cell infiltration
and migration CXCL8
(also known as IL-8)
Inflammation, wound contraction and epithelial proliferation
Angiogenesis, migration and invasion
CXCL12 Angiogenesis Migration, invasion and angiogenesis
Table 3.2 - Cytokines, chemokines and growth factors that influence wound healing and tumor progression.
Cytokines, chemokines and growth factors are included on the basis that they have been shown to influence both wound healing and tumor invasion or progression in vivo. CSF: colony stimulating factor, CXCL: C-X-C motif ligand, EGF: epidermal growth factor, FGF: fibroblast growth factor, IL: interleukin, PDGF: platelet-derived growth factor, MCP1: monocyte chemoattractant protein 1, TGF: transforming growth factor, TNF: tumor necrosis factor, VEGF: vascular endothelial growth factor. Adapted from Arwert et al. [1].