Sjövall 2013 Thesis Lund University - Sweden

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Sjövall F (2013) Mitochondrial function in sepsis - temporal evolvement of respiratory capacity in human blood cells. Thesis Lund University - Sweden 83pp.

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Sjoevall F (2013) Thesis Lund University - Sweden

Abstract:


O2k-Network Lab: SE Lund Elmer E


Labels: MiParea: Respiration, mt-Medicine, Patients  Pathology: Sepsis 

Organism: Human  Tissue;cell: Blood cells, Platelet  Preparation: Intact cells, Permeabilized cells 


Coupling state: LEAK, ROUTINE, OXPHOS, ET  Pathway: N, S, ROX  HRR: Oxygraph-2k 


Sepsis is a devastating disease that is caused by the host’s response to an overwhelming infectious process. As sepsis progresses, organs distant from the site of infection become affected and sepsis-induced multiple organ failure ensues. An impaired immunologic response, including dysfunctional peripheral blood immune cells has been described as part of the septic syndrome. Mitochondrial dysfunction has been suggested to be a contributing factor in the pathogenesis of these alterations and restoration of mitochondrial function has been implicated as a prerequisite for the recovery from sepsis. Further, platelets have been proposed to serve as a surrogate tissue in evaluation of systemic mitochondrial dysfunction. The overall aim of this thesis was to evaluate the temporal evolution of mitochondrial respiratory function in platelets and peripheral immune cells during the course of sepsis. In the first study we established methodology and performed a thorough assessment of normal human platelet respiratory function ex vivo from healthy individuals in a wide age-span using high-resolution respirometry. We concluded that freshly isolated platelets, intact or permeabilised, were well suited for studying human mitochondria ex vivo. With different titration protocols, detailed information of the cellular respiratory capacities could be obtained and we deemed this approach suitable for evaluating endogenous mitochondrial capacity as well as alterations of mitochondrial function induced by exogenous factors. In the two subsequent studies we examined mitochondrial respiratory function in platelets and peripheral blood immune cells (PBICs) of patients with severe sepsis or septic shock and studied its evolvement during the first week following admission to the intensive care unit. In both cell types we found that mitochondrial respiration (per cell) gradually increased during the week analysed. In platelets, this increase was higher in patients who subsequently died. Also, in platelets, we observed reduced respiratory control ratios of intact platelets when the cells where suspended in the patient’s own plasma. As markers for mitochondrial content we measured mitochondrial DNA (mtDNA), cytochrome c (Cyt c) and citrate synthase (CS). There was a difference between the two cell types in that the markers were profoundly more increased in PBICs compared to platelets even though they displayed approximately the same levels of increase in mitochondrial respiration. In the final study of this thesis we evaluated cytokines and nitric oxide in the plasma from the septic patient cohort since these signaling molecules have been demonstrated to enhance mitochondrial respiration through stimulation of mitochondrial biogenesis. Of ten different cytokines and NO analysed, IL-8 levels correlated positively with both maximal ATP-generating as well as maximal non-ATP12 generating rates of respiration in samples from the latest time point evaluated. Further, the plasma level of IL-8 was higher in non-survivors in samples taken at day 6-7 compared to survivors. In conclusion, this thesis demonstrates that circulating blood cells exhibit increased respiratory capacities throughout the first week of sepsis. This increase seems to be accomplished by different mechanisms; in PBICs by increased mitochondrial mass as indicated by elevated levels of mitochondrial markers, and in platelets possibly by a post-translational regulation of mitochondrial respiratory capacity. In addition, a plasma factor seems to be able to induce increased uncoupling of respiration in platelets during sepsis.