gms | German Medical Science

Objectives: Surgical treatment after multiple trauma (MT) is important in stabilising the patient but is considered the so-called 'second hit'. Invasive, prolonged surgery causes excessive inflammation and may be detrimental to a patient’s condition. This lies at the base of two main trauma-treatment strategies that mainly differ in their invasiveness in the acute phase after MT: Early-Total-Care (ETC) and Damage-Control-Orthopaedics (DCO). Both treatments have pros and cons, but exact cellular mechanisms that cause their differential effects on fracture healing are not fully known. The fracture hematoma (fxH) is a key element in initiating and prolongating the fracture healing cascade. Cellular communication mechanisms greatly depend on proteins. Although proteomics is increasingly applied in trauma research, this mainly focusses on circulatory proteins. The aim of this study was therefore to investigate the proteome of the fxH from a MT model, comparing two surgical treatments.

Methods: The porcine MT model consisted of bilateral femur fracture, blunt chest trauma, liver laceration, and controlled haemorrhagic shock. Animals were operatively and medically stabilised and monitored under ICU standards for 72 hours. Two fracture treatments were applied; intramedullary nailing (ETC; n=7), and external fixation (DCO; n=8). FxH was sampled, snap-frozen and stored at -80°C. Samples were sectioned and protein isolated. Proteins were gel separated and bands were collected and digested. Digested samples were run in an HPLC system. Proteome discoverer software was used for protein identification, and abundance and fold change calculations. The swiss-prot Sus scrofa database was used (TaxID 9823).

Results and conclusion: Label-free proteomics analysis was performed to investigate the differences in protein expression between the ETC and DCO groups, identifying a total of 2,311 proteins. Protein interaction networks were generated using STRING software. Of these proteins, 30 proteins showed a statistical difference (p<0.05; FC cutoff 1.5-fold) between the groups. Among those, 19 and 11 proteins showed higher abundance in the ETC and DCO groups respectively. These proteins were mainly involved in signalling pathways related to osteogenic differentiation and immune regulation.

This study is the first to describe the fxH proteome, showing that label free proteomics is a suitable analytical tool for protein analysis in fxH. The invasiveness of the surgical intervention had a clear effect on the fxH proteome at the injury site. Treatment-specific proteome changes were identified, linked to key processes in inflammation and fracture healing, such as the activation of the complement system, osteoblastogenesis, and immune cell attraction. These proteome changes indicate that ETC seemed to elicit more advanced fracture healing responses in the acute phase after trauma, while DCO prompted more balanced systemic inflammatory responses.