Changes in mitochondrial physiology, calcium homeostasis and uncoupling in sepsis.

Severe infections (termed ‘sepsis‘) can lead to the malfunctioning of organs such as the lungs, heart, liver and kidneys. Even with optimal medical management involving antibiotics, pharmacological and mechanical organ support in the intensive care unit, patients with sepsis continues to have a very high mortality. Why these organs fail - the so-called syndrome of ‘multi-organ failure‘ - is still unknown. Recent evidence has suggested that the energy-producing apparatus of the cell - the mitochondrion - becomes dysfunctional, causing a lack of sufficient energy to fuel the body‘s metabolic needs. I thus propose to study changes occurring within the mitochondria during sepsis. I plan to assess whether a relationship exists between excess production by the body of potentially damaging substances, abnormal functioning of the mitochondria, and the role of specialised proteins produced by the mitochondria which could play an important protective role. This work will hopefully lead to a better understanding of this devastating condition and the potential to develop exciting new therapeutic strategies.

Increasing evidence points to mitochondrial dysfunction, with a consequent reduction in ATP production, as a fundamental mechanism underlying the development of multi-organ failure in sepsis. Inhibition of the respiratory chain is caused by increased levels of reactive oxygen species (ROS), including nitric oxide, generated by excessive inflammation. Expression of the mitochondrial uncoupling proteins (UCP) 2 and 3 are increased in many tissues during sepsis, though their functional significance remains unknown. A modest mitochondrial membrane depolarisation caused by mild uncoupling will decrease ROS production and limit mitochondrial calcium accumulation. The latter may be crucial as mitochondrial calcium uptake will, in excess, damage the organelle. As ROS also activate UCPs, increased expression of UCPs may represent part of a negative feedback, protective mechanism in sepsis. Their contribution to the bioenergetic failure seen in sepsis is not known. Likewise, there are scanty data concerning changes in mitochondrial membrane potential and calcium signaling during the septic process.

I therefore wish to explore changes in mitochondrial physiology during sepsis, focusing on the functional consequences of increased UCP expression. I will use tissues derived from a well-established 3-day rodent faecal peritonitis model (utilising both wild-type and appropriate UCP knockout mice), and from patients suffering from septic shock. I will aim to establish what relationship, if any, exists between energy production and oxygen consumption, mitochondrial [Ca2+], ROS generation, mitochondrial membrane potential and expression of UCPs. I will assess (i) the temporal association between the above variables, and their variation in relation to the progression of the septic process, including resolution, (ii) the validity of using primary cultures incubated with endotoxin compared to cells and tissues extracted from septic animals and (iii) the specific consequences - protective or harmful - of increased UCP expression. I also plan to see whether mitochondrial function in septic animals can be assessed in vivo using multi-photon confocal microscopy.