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Abstract
Epidermolysis Bullosa (EB) is a group of inherited blistering disorders in response to mechanical trauma due to gene mutations. Despite great strides in research on the treatments for EB and correlations between genotype and phenotype, no definitive treatment is available. There are four aspects contained in this thesis. Firstly, a double-blind placebo-controlled trial to determine the clinical and molecular responses of intradermal cultured allogeneic fibroblast injections in chronic ulcers of five patients with generalised severe recessive dystrophic EB (RDEB-GS) which is due to COL7A1 mutations. My part in this study was the lab work including the analysis of the collagen VII gene and proteins expression. The clinical results showed that all five patients had rapid improvements in wound healing of both paired sites. My research showed that the healing could be independent of type VII collagen as only 3 of 5 patients had increased type VII collagen protein and COL7A1 mRNA, yet 2 of the 5 patients with healing wounds had negative collagen VII expression. This was also the first study to show that injections of suspension solution with or without allogeneic fibroblasts were able to improve the wound healing in chronic non-healing RDEB-GS wounds. This treatment could provide a feasible safe therapy for wound healing in RDEB patients. Secondly, we investigated whether quantitative immunofluorescence mapping of parental carriers could be used to determine the type of EB in situations where the EB infant has already died from a severe form of EB without testing. Our findings are the first to indicate that the carriers have a mild to moderately reduced relevant protein level in comparison with normal control skin. Thirdly, this is the first study of a family with EB simplex and ankyloblepharon. The splice site mutation KRT5 1474+1 G>A led to 129bp missing in exon 9, resulting in reduced expression of keratin 5 in the skin. Further, the last four amino acids of domain 2B and the domain H2 of keratin 5 were not important for maintaining proper function of the protein, as expression of a transcript lacking sequences coding for these amino acids was demonstrated in the clinically normal father. Fourthly, a family non-Herlitz junctional EB with LAMB3 mutations R635X and 1588del AG was studied to detect the consequence of gene mutations at the mRNA and protein levels. We found that1588delAG played a central role in protein expression and clinical manifestations probably because it led to a shortened mRNA transcript and reduced expression of β chain and γ2 chain of LM-332. Together, the data presented in this thesis demonstrates that injections of suspension solution, with or without allogeneic fibroblasts, could provide a feasible safe therapy for wound healing in RDEB patients, and studies on families with EB could help us to better understand the relationship between genotype and phenotype in EB.