Skeletal muscle mitochondrial uncoupling in a murine cancer cachexia model

Abstract

Approximately half of all cancer patients present with cachexia, a condition in which disease-associated metabolic changes lead to a severe loss of skeletal muscle mass. Working toward an integrated and mechanistic view of cancer cachexia, we investigated the hypothesis that cancer promotes mitochondrial uncoupling in skeletal muscle. We subjected mice to in vivo phosphorous-31 nuclear magnetic resonance (31P NMR) spectroscopy and subjected murine skeletal muscle samples to gas chromatography/mass spectrometry (GC/MS). The mice used in both experiments were Lewis lung carcinoma models of cancer cachexia. A novel 'fragmented mass isotopomer' approach was used in our dynamic analysis of 13C mass isotopomer data. Our 31P NMR and GC/MS results indicated that the adenosine triphosphate (ATP) synthesis rate and tricarboxylic acid (TCA) cycle flux were reduced by 49% and 22%, respectively, in the cancer-bearing mice (p<0.008; t-test vs. controls). The ratio of ATP synthesis rate to the TCA cycle flux (an index of mitochondrial coupling) was reduced by 32% in the cancer-bearing mice (p=0.036; t-test vs. controls). Genomic analysis revealed aberrant expression levels for key regulatory genes and transmission electron microscopy (TEM) revealed ultrastructural abnormalities in the muscle fiber, consistent with the presence of abnormal, giant mitochondria. Taken together, these data suggest that mitochondrial uncoupling occurs in cancer cachexia and thus point to the mitochondria as a potential pharmaceutical target for the treatment of cachexia. These findings may prove relevant to elucidating the mechanisms underlying skeletal muscle wasting observed in other chronic diseases, as well as in aging.

Figure 1

NMR spectra from in vivo³¹P NMR spectroscopy saturation-transfer experiments performed on the hind limb skeletal muscle of control (C) mice for determination of unidirectional inorganic phosphate (Pi) to ATP flux. Representative summed ³¹P-NMR spectra acquired from C before (upper curve) and after (lower curve) saturation of the γ-ATP resonance. The arrow on γ-ATP indicates the position of saturation by radiofrequency irradiation (−13.2 ppm). ppm, chemical shift in parts per million.

Figure 2

Dynamic profile for M + 2 to M + 1 ratio (M2/M1) of glutamate mass isotopomers during [2-¹³C]acetate infusion in mouse exhibiting cancer cachexia. Squares indicate experimental data points, and the line represents the best fit for our dynamic mass isotopomer model.

Figure 3

Rates of unidirectional ATP synthesis (left plot), TCA cycle flux (TCA, middle plot) and the coupling index (right plot), calculated as the ratio of the former two variables in control (C) and tumor-bearing (TB) mice.

Figure 4

(a) TEM micrographs from healthy control muscle (A–C) and cancer-induced cachectic (D–F) gastrocnemius muscle. White arrows, normal mitochondria; black arrow, triad structure. (b) Barograph showing the difference in the mitochondrial area between control and cancer-induced cachectic gastrocnemius muscle.

Figure 4

(a) TEM micrographs from healthy control muscle (A–C) and cancer-induced cachectic (D–F) gastrocnemius muscle. White arrows, normal mitochondria; black arrow, triad structure. (b) Barograph showing the difference in the mitochondrial area between control and cancer-induced cachectic gastrocnemius muscle.