Mark Wefers Bettink

Chapter 6 110 data were recorded starting 1 hour before administration of LPS until discharge 8 hour post-LPS, and at corresponding timepoints in the control group. Body temperature was measured using an infrared tympanic thermometer (FirstTemp Genius 2; Sherwood Medical, Mansfield, UK). As described previously [12], headache, nausea, shivering, muscle and back pain were scored on a six-point Likert scale (0 = no symptoms, 5 = worst ever experienced), resulting in a total sickness score of 0–25. In the endotoxemia protocol, the radial artery was cannulated using a 20-gauge arterial catheter (Angiocath; Becton Dickinson, Sandy UT, USA) and connected to an arterial pressure monitoring set (Edwards Lifesciences, Irivine, CA, USA) to allow continuous monitoring of blood pressure and continuous blood sampling for blood gas analysis various timepoints (see Fig. 1). In the control group, blood pressure was measured noninvasively using a cuff. Measurement of mitoPO2 and mitoVO2 using the COMET monitor Oxygen-dependent quenching of the delayed fluorescence lifetime of mitochondrial PpIX is the first known method to measure mitoPO 2 in living cells and tissues, in a non- invasive and feasible manner in humans. PpIX is the final precursor of heme in the heme biosynthetic pathway. PpIX is synthesized in the mitochondria and administration of 5-aminolevulinc acid (ALA) substantially enhances the PpIX concentration. We used ALAcare plasters for PpIX enhancement. Photoexcitation of PpIX populates the first excited triplet state and causes the emission of red delayed fluorescence. The delayed fluorescence lifetime is inversely related to the mitoPO 2 according to the Stern–Volmer equation. The background of the delayed fluorescence lifetime technique is extensively described elsewhere[8,14–16]. The light source and the detection system are the two core components of the COMET monitor (Photonic Healthcare, Utrecht, the Netherlands). A 515 nm pulsed laser, pulse duration 60ns, with a 10Hz repetition rate illuminates the intra cellular accumulated PpIX. The fluorescent signal is projected on a gated red-sensitive photomultiplier tube. The light emitted by the sensor is divergent and safe for eyesight at any distance. A detailed description of the COMET measuring system can be found elsewhere[7]. Local oxygen consumption is measured as mitochondrial oxygen consumption (mitoVO 2 ), by pressure-induced occlusion of the microcirculation thereby stopping local oxygen supply. mitoVO 2 and mitoPO 2 were measured with the COMET at baseline, 1.45 hours, 4 hours and 7 hours post-LPS administration, and at corresponding timepoints