Investigation of the Mechanism of Post-Fracture Immune Dysfunction and the Role of Statin in Its Prevention and Treatment Strategy: from Bench to Bedside

Trauma is the leading cause of death, especially among people aged 15 through 44 years, and contributes to the major burden of disease in the world, maybe ahead of ischemic heart disease in 2030. Traumatic injury is a battle between pro-inflammatory and anti-inflammatory responses. Over-inflammation induces organ failure. As compensatory antiinflammatory response syndrome predominates, trauma results in immunosuppression and increases the susceptibility to sepsis. Systemic responses to trauma are a complex interplay involving multiple levels and, however, there is currently no trauma cell model established for investigation of post-traumatic immunosuppression. About 90% of multiply injured patients have long bone fractures and patients dying from sepsis after trauma frequently have orthopedic injuries. Therefore, orthopedic trauma is one of the maj or parts in the field of trauma research. Evidence has showed that localized interaction with the immune cell at the fracture site might be responsible for the overall systemic reaction and outcome. Monocytes are chosen to set up the model not only because of the central role in innate immunity but also of the ability to orchestrate adaptive immunity. Besides, more than any one of the other Toll-like receptor (TLR) family member, as the recognizer of damage-associated molecular patterns, TLR4 sits at the interface of microbial and sterile (such as trauma) inflammation. TLR4 is the receptor of endotoxin and TLR4 downregulation is central to endotoxin tolerance, indicating immunosuppression. Mechanism of TLR expression after bone component exposure will be set as one of our research target. Compared with other mammalian genomes, human beings have more conserved 3’ untranslated region (3’UTR) than the promoter. About half of post-transcriptional control on 3’UTR relates to microRNAs and the others are associated with RNA-binding proteins (RBPs). Recent reports demonstrate that miR- 146a impairs TLR4 downstream signaling, and three microRNA databases predict direct interaction between miR-146a and TLR4 3’UTR; hence, miR-146a is suggested to be one candidate precipitating TLR4 mRNA instability. Our preliminary results confirmed that double immunosuppressive indexes, including TLR4 downregulation and miR-146a elevation, exist in bone component-treated monocytes. The first-year goals include as follows: determination of immune status of the trauma cell model by LPS priming, elucidation of the exact sequence of inflammation and immunosuppression, differentiation trend toward macrophage induced by bone component, confirmation of TLR4 mRNA instability and its relationship between microRNAs and RBPs, as well as validating the correlation between bone component exposure, gp96 manipulation and TLR4 expression. In the second year, possible interaction between gp96 and microRNAs will be examined by transfection of their mimics, inhibitors, as well as constructs incorporating luciferase reporter and 3’UTR of gp96. The effect of specific anti-miRs will also be examined and the relationship between TLR4 and these microRNAs tested by TLR4-mutant mice. Lung is the first organ to fail because after trauma because immune dysfunction will alter pulmonary vascular permeability and increases extravasation of albumin. Animal model shows infiltration of polymorphonuclear cell and increase of myeloperoxidase. Statin can regulate immunity and protect against sepsis and lung injury by helping lung repair. Therefore, in the third year, we aim to investigate the effect of statin on postoperative pneumonia in fracture patients and validate its protective effect by cell and animal models. Furthermore, monocytes and mononuclear cells from hip fracture patients will be treated by their bony fragments and serum from hematoma at the fracture site to examine the related immune status, differentiation and apoptotic index of monocyte and T-cells, changes of serum miRNAs, and the protective effect of statin. This study will build up a through model investigating post-fracture immune dysfunction from bench to bedside and employ techniques of cell biology and molecular biology to elucidate the effect of statin on post-traumatic immune dysfunction. We hope our results will provide information for trauma care and innovate the strategy for preventing postfracture sepsis and organ failure.