Critical roles for EGFR and EGFR-HER2 clusters in EGF binding of SW620 human carcinoma cells.

Epidermal growth factor (EGF) signalling regulates normal epithelial and other cell growth, with EGF receptor (EGFR) overexpression reported in many cancers. However, the role of EGFR clusters in cancer and their dependence on EGF binding is unclear. We present novel single-molecule total internal reflection fluorescence microscopy of (i) EGF and EGFR in living cancer cells, (ii) the action of anti-cancer drugs that separately target EGFR and human EGFR2 (HER2) on these cells and (iii) EGFR-HER2 interactions. We selected human epithelial SW620 carcinoma cells for their low level of native EGFR expression, for stable transfection with fluorescent protein labelled EGFR, and imaged these using single-molecule localization microscopy to quantify receptor architectures and dynamics upon EGF binding. Prior to EGF binding, we observe pre-formed EGFR clusters. Unexpectedly, clusters likely contain both EGFR and HER2, consistent with co-diffusion of EGFR and HER2 observed in a different model CHO-K1 cell line, whose stoichiometry increases following EGF binding. We observe a mean EGFR : EGF stoichiometry of approximately 4 : 1 for plasma membrane-colocalized EGFR-EGF that we can explain using novel time-dependent kinetics modelling, indicating preferential ligand binding to monomers. Our results may inform future cancer drug developments.

Open grade III "floating ankle" injuries: a report of eight cases with review of literature.

INTRODUCTION: "Floating ankle" injuries result from high-energy trauma and are usually associated with extensive soft tissue damage. Such rare and complex injuries in the acute phase pose therapeutic dilemma to the treating surgeon. When salvage instead of amputation is preferred, a variety of treatment options are available ranging from open reduction along with minimal internal fixation to external fixation. In this retrospective case series study we report eight patients with open "floating ankle" injuries who had been treated with external fixation with or without internal fixation. MATERIALS AND METHODS: Eight consecutive patients (7M: 1F) with mean age of 28 years (range 18-35 years) were treated. All had Grade III open fractures of the distal tibia with foot fractures and various degrees of soft tissue injuries. ISS and MESS were used to assess injury severity. Immediate radical wound debridement; skeletal stabilization and early soft tissue coverage were done by combined trauma and plastic surgical services followed by standard post-operative rehabilitation. All the patients were assessed with SF-36 questionnaire at mean follow-up of 2.5 years (range 2-4 years). RESULTS: Three patients were treated primarily with Ilizarov ring fixators, one had hybrid fixation and the other four had Hoffman frames. Four patients needed further surgeries. One patient developed metatarsal osteomyelitis, which was debrided and treated with antibiotics. The SF-36 results were compared with age matched UK norms for men and women in all categories. Only two patients returned to their previous employment. Six suffered varying degrees of disability out of which one underwent amputation. CONCLUSIONS: External fixation with or without internal fixation is an option when salvaging rare injuries like open grade III "floating ankle" injuries. Salvaged patients do suffer a cocktail of crippling disease characterized by psycho-socio-economic and physical disability and result in increased hospital costs. We recommend more prospective studies with long-term follow-up of such complex injuries to identify the indications for salvage and also the criteria for a successful salvage.