Mark Wefers Bettink

Chapter 4 76 inadequate calibration. On the contrary, in this study the COMET showed the tendency to underestimate mitoPO 2 in the case intracellular PO 2 was artificially increased to arterial oxygen levels. The mitoPO 2 measured with the COMET monitor is higher than expected. MitoPO 2 appears to be, depending on the measurement site and respiratory rate of the tissue, much closer to microvascular oxygen tension [20, 21], and thus closer to tissue and/ or interstitial oxygen levels [22, 23], than anticipated [7]. There are several reasons why mean mitoPO 2 in a tissue sample cannot be an order of magnitude lower than microvascular and interstitial oxygen tension; First, oxygen does not disappear stepwise but gradual, so several mitochondria will see aPO 2 close to intravascular values. Second, oxygen diffuses also from large vessels so contribute to cellular oxygen delivery [24], so several mitochondria have a higher PO 2 than capillary oxygen tension. Third, the mitoPO 2 will not be substantially lower than interstitial PO 2 because the oxygen gradient over the cell membrane is small [1]. Typically reported baseline mitoPO 2 values are 40-70 mmHg. Other oxygen measurements in the skin are matching these values [23]. Importantly, it has been demonstrated in both a preclinical [25] and clinical setting [4] that mitoPO 2 provides different information than hemoglobin saturation-based techniques like near- infrared spectroscopy. Application of cyanide on the skin led to a temporary block of mitochondrial respiration and abolishment of the oxygen gradient. In the first series of 9 investigated subjects, the timing between the laser pulse and the end of the off-gating of the photomultiplier (PMT) in the COMET proofed too long to adequately detect the short delayed fluorescence lifetimes caused by the artificially high intracellular PO 2 . The gating itself is necessary to prevent damage to the sensitive detector due to laser light and prompt fluorescence [4], and its timing is a trade-off between several factors, foremost the ability to accurately measure high mitoPO 2 (supraphysiological) and protecting the detector. After adjustment of this timing in the firmware, PMT gating interference was sufficiently reduced to allow collection of the delayed fluorescence signal after topical application of cyanide. Due to this adaptation, we were able to demonstrate that mitoPO 2 , as measured with COMET, corresponds well to PaO 2 in the absence of mitochondrial oxygen consumption. Under more physiological circumstances the timing of the PMT gating is much less critical as delayed fluorescence lifetimes are longer and easier to detect. COMET measured very low mitoPO 2 after oxygen deprivation and overall the calibration of the device seems adequate for its purpose.