Assessment of mitochondrial activity in 3D human liver microtissues as a tool for mechanistic investigations of adverse drug effects

Abstract

Specific cellular and inflammatory factors that contribute to the severity of pulmonary dysfunction after blunt chest trauma and osteosynthesis of femoral fractures are yet not fully understood. Therefore, we investigated alterations of the cytokine productive capacity of alveolar macrophages (AM) and Kupffer cells (KC) after femoral fracture stabilized with intramedullary pin with or without blunt chest trauma.In male C57BL/6N mice an intramedullary pin was implanted in an intact femur as the sham procedure. In trauma groups mice either received an isolated femoral fracture with subsequent fracture stabilization with an intramedullary pin (group Fx) or a combined trauma of blunt chest trauma and femur fracture also stabilized by an intramedullary pin (group TTFx). Animals were sacrificed 0 h, 6 h, 12 h, 24 h and 3 d after trauma induction. Cytokine concentrations were measured in plasma and supernatant of cultivated AM and KC by FACS analysis. Pulmonary and hepatic infiltration of polymorphonuclear leukocytes (PMN) was determined by Ly6G-staining.At 6 h, isolated femoral fracture with intramedullary stabilization resulted in a significantly increased productive capacity of KC (IL-6, TNF-α, CCL2, CCL3, CCL5 and CCL7) compared to sham animals. Combined trauma additionally resulted in an increased productive capacity of AM (IL-6, TNF-α, CCL2, CCL3, CCL4, CCL5 and CCL7) at 6 h and the effect was prolonged up to 3 d compared to controls. Combined trauma also led to a significant higher amount of plasma CCL2 at 3 d and plasma CCL7 at 6 h after the insult compared to group Fx. Compared to shams, pulmonary and hepatic infiltrations of PMNs were increased in group Fx and TTFx after 6 h, but in the combined trauma model the effect was prolonged up to 3 d.An intramedullary stabilized femur fracture alone results in a significant activation of the immune response. The combination of femoral fracture and blunt chest trauma however, results in an increased and prolonged activation of the inflammatory response. Transferred to the clinical setting, these results emphasize the critical role of severe chest trauma for treatment strategies of femoral fractures in multiple trauma patients.