Males, but Not Females, Demonstrate Mitochondrial Dysfunction in the C26 Model of Cancer Cachexia

Cancer cachexia is characterized by progressive muscle wasting that can lead to symptoms such as anemia, severe weight loss, and fatigue. These symptoms can lead to limitations in activities of daily living and can cause resistance to chemotherapy treatments in cancer patients. There are no current treatments available to treat cancer cachexia and a critical need remains to identify mechanisms of cancer cachexia. Recently, our group identified mitochondrial disfunction precedes muscle atrophy in males but not females in a model of lung cancer induced atrophy. However, it is unknown whether this finding is replicated when studying a different type of cancer. PURPOSE: This study set out to determine if mitochondrial respiration is impaired in the plantaris muscle in a well-established colon cancer model of cachexia. METHODS: The time-course study consisted of male and female mice in four groups per sex: An age-matched control (PBS), and three groups implanted with C26 tumors. Tumor growth for 10-15 days, 20 days, and 25 days. Tumors were implanted bilaterally into the hind flank for a total of 1X106 cells PBS (one-half per each hindflank). The plantaris was weighed for wet mass then teased into small fiber bundles and permeabilized for the quantification of mitochondrial function. Mitochondrial dysfunction was classified by a decrease in the respiratory control ratio (RCR), which is the ratio of state 3 (maximal ADP stimulated respiration) to state 4 (oligomycin-induced leak respiration). Male and Female data were analyzed separately using a one-way ANOVA. RESULTS: The tumor burden increased as the number of days increased. Male RCR showed a mean difference in RCR at the early timepoint (10-15 day, p=0.058) and demonstrated significantly lower RCR at the 20 day timepoint compared to PBS control (20d= 1.170± 0.094, PBS= 2.41 ± 0.13, p=0.031). Interestingly, RCR was not significantly different between male PBS and 25 days (1.864± 0.21, p=0.084). RCR in the plantaris from females was not different among any of the groups (p=0.401). CONCLUSION: Along with our previously published data in a lung cancer model, these data indicate that the mechanisms of muscle atrophy are sex dependent. Specifically, mitochondrial dysfunction appears to play an important role in cancer-induced atrophy in male, but not female, mice

The time-course of cancer cachexia onset reveals biphasic transcriptional disruptions in female skeletal muscle distinct from males

Abstract Background Cancer-cachexia (CC) is a debilitating condition affecting up to 80% of cancer patients and contributing to 40% of cancer-related deaths. While evidence suggests biological sex differences in the development of CC, assessments of the female transcriptome in CC are lacking, and direct comparisons between sexes are scarce. This study aimed to define the time course of Lewis lung carcinoma (LLC)-induced CC in females using transcriptomics, while directly comparing biological sex differences. Results We found the global gene expression of the gastrocnemius muscle of female mice revealed biphasic transcriptomic alterations, with one at 1 week following tumor allograft and another during the later stages of cachexia development. The early phase was associated with the upregulation of extracellular-matrix pathways, while the later phase was characterized by the downregulation of oxidative phosphorylation, electron transport chain, and TCA cycle. When DEGs were compared to a known list of mitochondrial genes (MitoCarta), ~ 47% of these genes were differently expressed in females exhibiting global cachexia, suggesting transcriptional changes to mitochondrial gene expression happens concomitantly to functional impairments previously published. In contrast, the JAK-STAT pathway was upregulated in both the early and late stages of CC. Additionally, we observed a consistent downregulation of Type-II Interferon signaling genes in females, which was associated with protection in skeletal muscle atrophy despite systemic cachexia. Upregulation of Interferon signaling was noted in the gastrocnemius muscle of cachectic and atrophic male mice. Comparison of female tumor-bearing mice with males revealed ~ 70% of DEGs were distinct between sexes in cachectic animals, demonstrating dimorphic mechanisms of CC. Conclusion Our findings suggest biphasic disruptions in the transcriptome of female LLC tumor-bearing mice: an early phase associated with ECM remodeling and a late phase, accompanied by the onset of systemic cachexia, affecting overall muscle energy metabolism. Notably, ~ 2/3 of DEGs in CC are biologically sex-specific, providing evidence of dimorphic mechanisms of cachexia between sexes. Downregulation of Type-II Interferon signaling genes appears specific to CC development in females, suggesting a new biological sex-specific marker of CC not reliant on the loss of muscle mass, that might represent a protective mechanism against muscle loss in CC in female mice

Supplemental table 4

Common DEGs (up- and downregulated) at 24h, 48h, 72h, 168h and merging all timepoints together between males and females.</p

Supplemental table 1

“KEGG” pathway analysis with a background reference gene list comprised of all genes in our dataset (ShinyGO 0.77).</p

Supplemental table 6

Common pathways (up- and downregulated) at 24h, 48h, 72h and 168h between males and females.</p

Supplemental table 2

“GO Cellular Component” pathway analysis with a background reference gene list comprised of all genes in our dataset (ShinyGO 0.77). </p

Supplemental table 5

Pathway analysis (G:profiler) in the total mouse transcriptome.</p

Supplemental table 3

Overlapping DEGs (up- and downregulated) across all timepoints within each sex.</p

Supplemental Figure 1

Venn diagrams presenting the shared MitoCarta genes, overlapping with up- (A) and downregulated (B) DEGs at all timepoints, between males and females. Light red circles represent males and light green circles represent females.</p