Abstract

The analysis of mitochondrial function is fundamental in the diagnosis of muscle pathologies. Long term preservation of human tissues for functional analysis is a frontier challenge in diagnostics but has potential benefits for inter-laboratory and interdisciplinary analyses. High resolution respirometry of permeabilized muscle fibers allows for quantification of mitochondrial function ex-vivo from small biopsy samples. Scientific studies on respirometry of cryopreserved muscle tissues have seldom been performed. In this study we investigated mitochondrial respiration of DMSO cryopreserved muscle tissue and compared oxygen flux rates to freshly harvested muscle from the same biopsy sample. The purpose of this study is to test the functional capacity of mitochondria in cryopreserved muscle tissue and to compare respirometry to fresh muscle mitochondrial tissue. Cryopreserved muscle showed robust coupled state 3 mitochondrial respiration comparable to fresh muscle. However, the magnitudes of state 3 (ADP-driven) respiration rates for complex I substrates with malate and glutamate and complex I and II with addition of succinate were lower than that of fresh muscle tissue. The significant increase in state 3 respiration upon addition of cytochrome c indicates that cryopreservation causes results in cytochrome c loss and potential disruption of the inner mitochondrial membrane. Optimization of the cryopreservation method will permit long-term storage of samples, allow mitochondrial function and energy metabolism to be analyzed and provide the potential for exchanging samples between research centers.