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Metabolic Fluctuations in Porcine Corneas with Different Culture Media - Oxygen Consumption in a Perfusion System
Bochert A., Berger E., Saedler J., Stave J., Guthoff R.,
Universität Rostock, Augenklinik (Rostock)
Purpose: Tissues generate their ATP-pool by aerobic and anaerobic metabolic pathways. The aerobic part is a high gaining ATP process whereas anaerobic metabolism like anaerobic glycolysis result in acidification of the tissue and a lower ATP gain. For tissues which have to achieve ATP demanding work like tissue integrity or ion pumping for example, aerobic capacity is prerequisite for a healthy tissue. Our purpose was to estimate the amount of aerobic capacity of the cornea and to assess the viability especially of the endothelium during organe culture. Therefore we examined the oxygen consumption, glucose and lactate metabolism depending on different media in an open flow perfusion system.
Methods: Investigated media were Iscoves, DMEM, MEM, Waymouth/Nutrient, M199/Nutrient, Nutrient, supplemented with 10% FCS and serum replacement (Biochrom, Germany). Two porcine corneas were kept in a perfusion chamber (Minuthchamber) with a medium flow of 2 ml per hour. Glucose, Lactate, pH and oxygen consumption were determined for 1 week. Endothelium integrity was evaluated by light microscopy and laser scanning microscopy.
Results: MEM containing 10% FCS was the medium with the highest oxygen consumption rate and lowest lactate formation and therefore low pH change. Higher amounts of lactate lead to acidification of the medium and oxygen consumption decreased in the other media. Aerobic metabolism of corneas in the different media was lower and showed sometimes a characteristic depletion towards the end of experiments (Nutrient), indicated by a higher anaerobic lactate formation and lower pH.
Conclusions: A high aerobic capacity is a prerequisite for a high ATP gain or turnover which is needed for the ionic pump activity of the endothelium. Surprisingly the medium which showed to be the best for aerobic capacity is the mostly used MEM in organe culture in contrast to the often used Iscove Medium in open-flow systems. MEM seems to be well adapted for the metabolic demands of the corneal cells especially the endothelium.