The effect of keratinocytes differentiated from mouse embryonic stem cells on wound healing in a wound model created in mice under long-term steroid pressure

Long-term steroid use due to chronic diseases causes chronic wounds by delaying wound healing. Dexamethasone is one of the commonly used glucocorticoids. The aim of this specialization thesis is to examine the effects of keratinocyte-like cells differentiated from mouse embryonic stem cells on wound healing by histochemical and immunohistochemical methods in long-term steroid use in mice. 64 balb-c type male mice were divided into 4 groups: only the wound model was created (wound group); after the wound model was created, keratinocyte was applied (cell group); after applying 2 mg/kg dexamethasone per day for 1 week, the wound model was created and dexamethasone application was continued (steroid group); 2 mg/kg dexamethasone was applied daily for 1 week before the wound model was created, keratinocyte was applied after the wound model was created and dexamethasone application was continued (steroid group+cell group). Keratinocyte-like cells to be applied to the wound site were amplified by culturing mouse embryonic stem cells with mouse fibroblasts, differentiated by BMP-4 application, and their characterization was completed by confirming cytokeratin-8 and cytokeratin-14 levels both by immunocytochemical and flow cytometry. Keratinocyte-like cells were applied only once to the cell and cell+steroid group, and dexamethasone application was continued for 21 days in the steroid and cell+steroid group after the wound was created. Skin tissue sections taken on the 10th and 21st days after wound formation were first evaluated by hematoxylin-eosin staining. Afterwards, the distributions of cytokeratin-8, cytokeratin-14, MCP-1, TNF-α, VEGF, IL-1β and COL1A1 were evaluated immunohistochemically, the wound healing processes were monitored and the groups were compared with each other. The results showed that histochemically wound healing was positively affected by cell application in the presence of long-term steroids (21 days), immunohistochemically, keratinocyte-like cells continued to express cytokeratin at the wound site, TNF-α remained at basal level for 21 days, but negative IL-1β' was observed. In addition, the increase in MCP-1, VEGF and COL1A1 levels showed that cell application to the wound site contributed to wound healing by supporting keratinization. This specialization thesis provided a preliminary information for clinical practice, there is still a need for studies with different materials and methods.