239 General Discussion TNF-α, G-CSF, GRO-α (CXCL1) and IL-8 (CXCL8)) were highly increased in wound tissue from animals and patients [18–23]. Furthermore burns increased the level of MCP-1 (CCL2), a chemoattractant for monocytes and supporter of differentiation of monocytes towards macrophages [24,25]. Based on this cytokine milieu, the excessive response of both neutrophils and monocytes is expected to be severe. On the other hand, in wound tissue from burn patients we found that lymphocyte attractants (MIP-1α (CCL3), MIP-1β (CCL4) and RANTES (CCL5)) peaked at the second or third week after burn injury [26]. This could mean that while the innate immune system is highly activated immediately after burn, lymphocyte activation could be delayed. This is in line with a studies from Finnerty et al. and Jeschke et al. who showed elevated levels of cytokines that activate innate immune cells in burned children, adults and elderly [27–30]. Other studies have shown that the cytokine profile after burn is associated with an immediate, severe and prolonged innate immune response and the levels of IL-1, IL-6 and MCP-1 can even be linked to increased morbidity and mortality [31,32]. The accumulation of activated immune cells can cause damage to surrounding tissues. This then leads to the release of more DAMPs and cytokines, establishing a vicious circle of inflammation that delays wound healing [10,33]. In Chapter 6, we used full skin equivalents (FSEs), consisting of fibroblasts and keratinocytes, to study skin development and skin regeneration after burn injury. In Chapter 7, either monocytes or T cells. were incorporated into these FSEs. An important advantage of this models is combinations of incorporated cells lead to the release of specific cytokines. For example, we discovered that IL-6, IL-8 and MCP-1 were highly expressed in FSE models that included only fibroblasts and keratinocytes. This high expression was probably related to a stress response induced by the in vitro culture of these cells [34]. When burn injury was inflicted on these FSE models without immune cells the production of IL-4, IL-6, IL-8 and IL-12p70 increased even further. The release of IL-1β, which was highly expressed after burns in animals and humans, was only detected when monocytes were included in FSE models. Likewise, the expression of IFN-γ and IP-10 was significantly higher when T cells were present in the models, regardless of burn injury. The increased percentage of T cells expressing IP-10 receptor CXCR3 (CD183) might be related to this. These findings demonstrate that interactions between skin cells and immune cells can drive cytokine production and will impact the immune response. Other researchers also showed that co-culture of T cells with keratinocytes increased the levels of chemokines, particularly MCP-1, MIP-3α (CCL20), MIG (CXCL9) and IP-10 (CXCL10) [35–37]. Presumably, T cells produce IFN-γ which stimulates keratinocytes to produce chemokines, especially in inflamed tissue [35,38]. Crosstalk might also happen between T cells and fibroblasts during inflammation [39], which could play a role in (preventing) fibrosis. 8
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