Toll-Like Receptor Signalling In Diabetic Wound Healing

UoM administered thesis: Phd

  • Authors:
  • Robert Harrison


Wound healing is a complex process defined by three distinct yet overlapping stages: inflammation, proliferation and remodelling. Macrophages play a key role in all three stages, as they can switch between pro-inflammatory 'M1-like' states and anti-inflammatory 'M2-like' states. In approximately 4-10% of patients with diabetes, aberrant inflammation leads to the formation of nonhealing diabetic foot ulcers (DFUs). One of the reasons for DFU formation is a failure to switch between these M1 and M2 states. In this thesis, I aim to elucidate whether dysregulation within the toll-like receptor (TLR) signalling pathway plays a role in the formation of an excessive pro-inflammatory phenotype. An in vitro functional screen of TLRs in which macrophages were stimulated with the specific TLR agonists showed that diabetic macrophages express significantly higher levels of pro-inflammatory cytokines. However, the fold increase of cytokine production is much greater in non-diabetic macrophages. The expression levels of TLRs were not different between non-diabetic and diabetic macrophages. Next, non-diabetic and diabetic mice were given a bone marrow (BM) transplant of cells with a knockout of the TLR adaptor protein Myd88. Interestingly, although ndb mice with Myd88-/- BM healed wounds significantly slower than their counterparts with wild-type BM, there was no difference between the rate of wound healing in diabetic mice with wild-type BM and those with Myd88-/- BM, implying a significant role for Myd88- independent inflammatory signalling. Finally, ChIP-seq was used to examine the differences in NF-kB binding. It was discovered that NF-kB binds to DNA overwhelmingly in distal intergenic regions in diabetic macrophages compared with non-diabetic macrophages, suggesting dysregulation of chromatin conformation in diabetic macrophages. Together these results demonstrate that even though diabetic macrophages are inherently more pro-inflammatory, they fail to actively mount an inflammatory response in the same way as non-diabetic macrophages. This primarily seems to be because of great changes in the regulation of inflammatory transcription factor binding rather than aberrant TLR activity.


Original languageEnglish
Awarding Institution
Award date1 Aug 2019