Non-invasive monitoring of mitochondrial oxygenation and respiration in critical illness using a novel technique

Abstract

Introduction: Although mitochondrial dysfunction is proposed to be involved in the pathophysiology of sepsis, conflicting results are reported. Variation in methods used to assess mitochondrial function might contribute to this controversy. A non-invasive method for monitoring mitochondrial function might help overcome this limitation. Therefore, this study explores the possibility of in vivo monitoring of mitochondrial oxygen tension (mitoPO2) and local mitochondrial oxygen consumptionin in an endotoxin-induced septic animal model.

Methods: Animals (rats n = 28) were assigned to a control group (no treatment), or to receive lipopolysaccharide without fluid resuscitation (LPS-NR) or lipopolysaccharide plus fluid resuscitation (LPS-FR). Sepsis was induced by intravenous LPS injection (1.6 mg/kg during 10 min), fluid resuscitation was performed by continuous infusion of a colloid solution, 7 ml kg(-1) h(-1) and a 2-ml bolus of the same colloid solution. MitoPO2 and ODR were measured by means of the protoporphyrin IX-triplet state lifetime technique (PpIX-TSLT). Kinetic aspects of the drop in mitoPO2 were recorded during 60s of skin compression. ODR was derived from the slope of the mitoPO2 oxygen disappearance curve. Measurements were made before and 3 h after induction of sepsis.

Results: At baseline (t0) all rats were hemodynamically stable. After LPS induction (t1), significant (p < 0.05) hemodynamic changes were observed in both LPS groups. At t0, mitoPO2 and ODR were 59 ± 1 mmHg, 64 ± 3 mmHg, 68 ± 4 mmHg and 5.0 ± 0.3 mmHg s(-1), 5.3 ± 0.5 mmHg s(-1), 5.7 ± 0.5 mmHg s(-1) in the control, LPS-FR and LPS-NR groups, respectively; at t1 these values were 58 ± 5 mmHg, 50 ± 2.3 mmHg, 30 ± 3.3 mmHg and 4.5 ± 0.5 mmHg s(-1), 3.3 ± 0.3 mmHg s(-1), 1.8 ± 0.3 mmHg s(-1), respectively. At t1, only mitoPO2 showed a significant difference between the controls and LPS-NR. In contrast, at t1 both LPS groups showed a significantly lower ODR compared to controls.

Conclusion: These data show the feasibility to monitor alterations in mitochondrial oxygen consumption in vivo by PpIX-TSLT in a septic rat model. These results may contribute to the development of a clinical device to monitor mitochondrial function in the critically ill.

Fig. 1

a Principle of the PpIX-TSLT. The pathway by which topical ALA administration enhances mitochondrial PpIX levels and the principle of delayed fluorescence detection after an excitation pulse with green (510 nm) light. Emission light is the delayed fluorescence (red light, 630–700 nm) and its lifetime is oxygen dependent. b PpIX emits delayed fluorescence after excitation by a pulse of green (510 nm) light. The delayed fluorescence lifetime is oxygen dependent according to the Stern–Volmer equation (inset), in which k _q is the quenching constant and τ₀ is the lifetime at zero oxygen. ALA 5-aminolevulinic acid, CPIII coporporphyrinogen III, PBG porphobilinogen, PO ₂ oxygen tension, PpIX protoporphyrin IX, UPIII urporphyrinogen III

Fig. 2

Principles of respirometry in the skin by oxygen-dependent quenching of delayed fluorescence of PpIX. a Baseline measurement. b Principle of the pressure-induced cessation of microvascular blood flow. V _m measurement volume

Fig. 3

Schematic flow chart of the experimental setup. ALA 5-aminolevulinic acid, LPS lipopolysaccharide, PpIX protoporphyrin IX

Fig. 4

Representative example of mitochondrial oxygen consumption measurement in a rat from the nonresuscitation group. a Before administration of LPS. b Three hours after administration of LPS. LPS lipopolysaccharide, mitoPO ₂ mitochondrial oxygen tension, ODR oxygen disappearance rate

Fig. 5

Average mitoPO₂ and ODR in the nonresuscitation group at three time points: baseline (t0) before the induction of sepsis, 3 hours after LPS infusion (t1), and after late fluid resuscitation (t2). Data presented as mean ± standard error. *Significant difference compared with baseline measurements (p <0.05). LPS lipopolysaccharide, mitoPO ₂ mitochondrial oxygen tension, ODR mitochondrial oxygen consumption