Sepsis remains a leading cause of mortality in critically ill patients. Previous data show that patients with sepsis and critical illness (e.g., burn injury) experience a disruption of the sustained directionality of polymorphonuclear neutrophil (PMN) migration, while gaining random spontaneous PMN motility. Aberrations in directional PMN migration impair effective bacterial clearance and cause unnecessary PMN organ infiltration, leading to multi- organ damage. The molecular basis of disturbances in PMN motility is poorly understood. Preliminary data for this project indicate that inhibiting mitochondrial microtubule (MT) assembly/disassembly or inhibiting MT- dependent directional transport of mitochondria between the leading and trailing edges of PMNs causes impaired directional chemotaxis and/or spontaneous motility. It is hypothesized that: (1) MT dynamics and MT- dependent directional mitochondrial trafficking are required for sustained directionality in PMN chemotaxis; and (2) impaired directionality and spontaneous motility are causally associated with stabilized MTs, which induce decreased or aberrant mitochondrial translocation and increase basal ATP secretion in sepsis and critical illness. Preliminary studies related to the underlying molecular mechanisms showed that farnesyltransferase (FTase) inhibitor (FTI) inhibited PMN organ infiltration, promoted PMN migration to the infection site, and improved survival and bacterial clearance in septic mice. LKB1 is a master kinase that regulates a wide range of cellular processes, including MT remodeling and MT-dependent mitochondrial trafficking. In preliminary studies, (a) a chemoattractant, f-Met-Leu-Phe (fMLP) increased phosphorylation of LKB1 at Ser431 in PMNs, while farnesylation at Cys433 inhibited phosphorylation of LKB1; and (b) farnesylation-resistant LKB1 mutant (Cys433Ser) knock-in mice were protected from sepsis-induced impaired PMN directional migration.
Three Specific Aims will test the following hypotheses.
Specific Aim 1 : Determine the role of MT dynamics, MT- dependent mitochondrial trafficking, and regulated ATP secretion in the sustained directionality and spontaneous motility of PMNs.
Specific Aim 2 : Test the hypothesis that (1) FTI improves PMN directional migration by restoring MT-dependent mitochondrial trafficking; and (2) farnesylation of LKB1 is integral to aberrant PMN migration in sepsis and burn injury in mice.
Specific Aim 3 : Establish impaired directionality and spontaneous motility as new major facets of PMN dysfunction and test the following hypotheses: (1) in vitro FTI treatment ameliorates the aberrant mobility of PMNs in critically ill patients; and (2) sepsis is associated with marked increases in basal ATP release and spontaneous motility of PMNs. This proposal will establish the physiological role of dynamic MT-dependent mitochondrial translocation in directional PMN migration and the pathophysiological role of dynamic MT remodeling in impaired directionality in chemotaxis and spontaneous motility of PMNs in sepsis and critical illness. This proposal is expected to spawn new clinical trials of FTI in septic patients and/or in at-risk critically ill patients.
Neutrophils, a type of white blood cells, migrate to a site of infection to kill bacteria that have invaded into our body. In critically ill conditions, however, neutrohils often fail to eradicate bacteria because they lose their way during the migration. The proposed studies will clarify how and why directional migration of neutrophils is impaired in critically ill patients, and test the effectiveness of using a drug called as farnesylation inhibitor (used in cancer treatment) to improve neutrophil directional migration in the prevention and treatment of sepsis (severe systemic infection).
|Nakazawa, Harumasa; Ikeda, Kazuhiro; Shinozaki, Shohei et al. (2017) Burn-induced muscle metabolic derangements and mitochondrial dysfunction are associated with activation of HIF-1? and mTORC1: Role of protein farnesylation. Sci Rep 7:6618|