The role of Toll-like Receptor 4 in diabetic foot ulceration

Introduction Diabetes Mellitus has reached epidemic proportions. Foot ulceration is a multifactorial complication of diabetes associated with marked morbidity and mortality. Innate-immune Toll-like receptor 4 (TLR4) mediated inflammation has been implicated in the systemic pathogenesis of diabetes and may contribute to impairment of wound healing. This thesis investigates the effect of high glucose and hypoxic conditions on TLR4 activation and signalling in vitro and in vivo. Method Human skin biopsies were stained for H&E and TLR4. Fibroblasts cultured at physiological glucose concentration (5.5mM) were exposed to pathological glucose concentrations (25mM), with duplicate samples placed in a hypoxia chamber. The effect of TLR4 and its signalling pathway was assessed through specific inhibitors. Diabetes was induced in wild-type (WT) and TLR4 knock-out C57BL/6 mice by intra-peritoneal injection of low-dose streptozocin. Hindlimb ischaemia was induced by femoral artery ligation four weeks post- streptozocin, and a full thickness skin wound inflicted below the knee. Wound healing was assessed via digital planimetry at days 3, 7 and 14 post surgery. Results Diabetic-ischaemic ulcers demonstrated greater inflammatory cell infiltration (p=0.0001) and TLR4 expression (p=0.038). Hypoxic and high glucose (25mM), conditions led to an increase in TLR4 protein expression, apoptosis and IL-6 release. Inhibition with MyD88 inhibitory-peptide, TLR4 neutralising-antibody and specific TLR4 antagonist ameliorated the effects of high glucose and ischaemia (p<0.05). In vivo, wound healing was significantly impaired in the diabetic-ischaemic group at day 14 (p<0.05). Diabetic-ischaemic wounds in TLR4 KO mice exhibited significantly improved healing rates compared to those in WT mice at all time points. Conclusion Diabetic-ischaemic ulceration is associated with increased cellular inflammation and TLR4 expression. Hypoxia stimulates up-regulation of TLR4 protein expression and this effect is exaggerated by hyperglycaemia. Inhibition of TLR4 significantly reduced TLR4 protein expression, apoptosis and IL-6 release, suggesting a protective effect. In TLR4 KO mice, there is a significant improvement in the healing of diabetic-ischaemic wounds compared to WT. We propose a synergistic effect between hypoxia and hyperglycaemia impairing wound healing exists, through TLR4-mediated inflammation

The "Woundosome" Concept and Its Impact on Procedural Outcomes in Patients With Chronic Limb-Threatening Ischemia

This editorial assembles endovascular specialists from diverse clinical backgrounds and nationalities with a global call to address key challenges to enhance revascularization in chronic limb-threatening ischemia (CLTI) patients.- Dedicated below-the-ankle (BTA) angiography and revascularization is underutilized in ischemic foot treatment. Existing guidelines do not address comprehensive BTA vessel analysis. CLTI trials also often lack data on in-line arterial flow to the ischemic lesion and BTA vessel evaluation, hindering outcome assessment.- Dedicated multi-planar angiographic evaluation of the distal microcirculation is key: Direct arterial flow or good-quality collaterals are crucial in influencing wound healing and need to be assessed diligently to the level of the distal ischemic wound territory, termed “woundosome.”- An important primary emphasis of future trials should be on validating technologies and strategies for assessing tissue perfusion before, during, and after revascularization undertaken to heal tissue loss in CLTI patients. This will allow determination of a potentially significant delta in tissue perfusion prior to and following intervention at the “woundosome” level. Once changes in arterial perfusion have been identified as positively correlated to wound healing, these could serve as a much-needed novel primary technical outcome measure for patients with tissue loss undergoing surgical, hybrid, or endovascular revascularization