It has previously been shown that VN can form complexes with IGF-II or IGF-I in combination with its binding proteins IGFBP-3 or -5. This study aimed to determine the efficacy of using these complexes as a treatment designed to accelerate wound healing, particularly in diabetic ulcers. The primary functions of skin cells in wound healing are attachment, proliferation and migration, thus these functions were assessed in response to these complexes in skin cells derived from patients with diabetic ulcers and from non-diabetic patients. These studies examined responses to the complexes in both skin keratinocyte and fibroblast cells. Furthermore, in order to investigate the mechanisms that underlie the responses observed, I also examined the ability of skin cells to retain these functional responses when the complexes incorporated an IGF-I analogue that does not activate the IGF receptor or when the cells had been pre-incubated with an anti-αv-integrin function blocking antibody. In addition, the ability of the cells to survive and grow when treated with the complexes under conditions mimicking the diabetic wound was assessed using growth assays in which the media contained elevated concentrations of glucose and calcium. I found that cells derived from skin from normal patients showed enhanced proliferation in response to these complexes, whereas only the presence of IGF-I and IGFBP seemed to be important in stimulating the proliferation of cells derived from diabetic patients. I also found that enhanced migration was observed in fibroblasts from diabetic ulcers in response to the complexes but these responses only required the presence of VN in normal cells. Both normal and diabetic keratinocytes showed enhanced migration in response to the complexes and the responses involved the interaction of both IGF-I and VN with their respective cell surface receptors. However the enhanced migration observed in diabetic ulcer derived keratinocytes was approximately half the level seen in normal keratinocytes. Furthermore, I showed that cells derived from skin from normal patients exhibited greater proliferation when treated with complexes in the presence of high concentrations of glucose and calcium ion compared to cells that were not treated with the complexes. Likewise, cells derived from skin surrounding diabetic ulcers were able to grow in media containing high levels of glucose and calcium when treated with VN:IGFBP:IGF-I complexes. In particular diabetic skin derived fibroblasts grown in high calcium media demonstrated enhanced proliferation when treated with the complexes, whereas diabetic keratinocyte cells seemed less affected by these conditions than their normal counterparts were. The findings in this thesis show that VN:IGFBP:IGF-I complexes can elicit enhanced growth and migration in cells derived from skin from both normal and diabetic patients. Further, these responses are maintained in conditions found in the diabetic wound microenvironment, namely in the presence of high glucose and high calcium. Together these findings demonstrate the potential of the VN:IGFBP:IGF complexes as wound healing agents to treat wounds, especially diabetic ulcers. Such delayed healing wounds represent a significant burden to health care systems and are one of the primary conditions that leads to the amputation of limbs. Current treatments do not address the co-ordination of ECM and growth factor action on cells that is here demonstrated to stimulate multiple wound healing related functional effects in skin cells. The data presented here represents important new information that may guide the design of new integrated therapeutics that may enhance the healing of recalcitrant diabetic ulcers.
| Identifer | oai:union.ndltd.org:ADTP/265661 |
| Date | January 2008 |
| Creators | Noble, Anthony M. |
| Publisher | Queensland University of Technology |
| Source Sets | Australiasian Digital Theses Program |
| Detected Language | English |
| Rights | Copyright Anthony Michael Noble |
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