Pathogenesis of keloidal scars

Abstract

Introduction: Keloid disease can be defined as a dermal fibro-proliferative disorder characterised by excessive deposition of extracellular matrix components such as collagen and glycoproteins. This excessive deposition of fibrous tissue only occurs after cutaneous trauma. The purpose of this study was to understand the pathogenesis of the disease including its possible link with viruses such as human herpesviruses (HHV) and the human papillomavirus (HPV) family, since both of these viral families are known to cause skin cancer.Methods: Tissue samples were collected from keloid and non-keloid patients. The keloid samples were divided into sections; top, margin, middle, bottom and internal normal (internal control). These cells were further divided into two groups for snap freezing and placement in phosphate buffer saline (PBS). The snap frozen tissues were used to extract DNA and protein while the PBS treated samples were used to isolate and culture fibroblasts. The DNA was used to assess presence of HHV and HPV while protein was analysed by two-dimensional gel electrophoresis (2D-GE). Key transcripts from fibroblasts were extracted and profiled using reverse transcription polymerase chain reaction (RT-PCR).Results: Perturbation in cell cycle regulatory genes, especially cdc2 was observed. Mostly suppression of gene expression was notable in the lowest region of the scar. Protein profiling revealed the presence of mitochondrial-associated proteins at the margins. The presence of heat shock proteins both in the margin and in the internal control suggest that keloid patients express elevated levels of this protein both in the normal state as well as when the scar is progressing. However despite extensive testing and care to avoid presence of inhibitors the results for both HHV and HPV proved to be negative.Conclusions: Based on the observation in the cell cycle regulatory genes, cdc2 mRNA expression was shown to be down regulated across all keloidal sites of the scar including the internal controls when compared to external control. This down regulation was also evident at the protein level. Perturbations of other genes were noted as well. Fibroblasts from the bottom of the keloid scar showed down-regulation in cyclin E2, cyclin A, cyclin B and up-regulation of p27. All these data suggest the bottom to be the most quiescent part of the scar, with other parts being more active, hence the growth characteristics of the scar. Protein profiling of the scar revealed the presence of mitochondrial-associated proteins at the margins, suggesting these areas to be the most active regions of the scar.Despite the development of highly sensitive PCR-based micro-array assays, HPV could not be detected. It was proven that the micro-array developed in this section was more diverse and more accurate than the technique used currently in clinics although it did suffer from sensitivity when compared against a next generation sequencing approach.Again despite more sensitive methods to detect HHV in compare to technique seen in clinics, presence of HHV was ruled out as there were no sign of HHV in keloidal tissue.

Date of Award	1 Jan 1824
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Henry Kitchener (Supervisor), Ardeshir Bayat (Supervisor) & Philip Day (Supervisor)