(PTFS15-02-26) Long-Chain Polyunsaturated Fatty Acids Mitigate In Vitro Skeletal Muscle Wasting Induced by Colorectal Cancer via a Lipoxygenase (LOX)-dependent Pathway

Objectives: Up to 50% of adults with colorectal cancer (CRC) are at risk of developing cachexia, featured by the loss of skeletal muscle mass and function. Long-chain polyunsaturated fatty acids (LC-PUFAs) and their bioactive metabolites, termed specialized pro-resolving lipid mediators (SPMs), exhibit anti-inflammatory properties and promote muscle repair. This study aimed to examine the effects of n-3 and n-6 LC-PUFAs on CRC-induced muscle wasting and the underlying cellular and molecular mechanisms.

Methods: Differentiated murine C2C12 skeletal muscle cells were treated with conditioned media derived from CT26 colorectal carcinoma cells. Individual PUFAs including arachidonic acid (ARA, 20:4n-6), eicosapentaenoic acid (EPA, 20:5n-3), docosapentaenoic acid (DPA, 22:5n-3), and docosahexaenoic acid (DHA, 22:6n-3) were added to C2C12 cells at 25 μM with or without the presence of LOX inhibitors including NDGA or BLX-3887 for 72 hours. Immunocytochemistry was used to visualize cell morphology, and myotube diameter was quantified to assess muscle wasting. Conditioned media was obtained from a C2C12-CT26 co-culture model receiving individual PUFAs, and targeted LC-MS/MS was used to profile lipid metabolites produced by the co-culture. Differences between groups were tested by a one- or two-way ANOVA followed by pairwise Holm-Šidák post hoc tests, and statistical significance was set as P ≤ 0.05.

Results: ARA, EPA, DPA, and DHA each mitigated CT26-induced muscle wasting by increasing myotube diameter, with EPA exerting the greatest protective effect. Inhibition of LOX activity completely blocked the benefits of LC-PUFAs, indicating a critical role of LOX-derived PUFA metabolites. In response to PUFA treatments, muscle-cancer co-culture produced bioactive SPMs derived from LOX-dependent pathways, including lipoxins (e.g., LXA₄), D-series resolvins (e.g., RvD1), and E-series resolvins (e.g., RvE1).

Conclusions: LC-PUFAs including ARA, EPA, DPA, and DHA protected C2C12 muscle cells against CT26-induced cachexia. The protective effect depended on LOX-catalyzed PUFA metabolism, suggesting a potential mechanism by which PUFAs mitigate CRC-induced cachexia.

Funding Sources: The study is supported by College of Agriculture startup funding at Purdue University and the 2023 Indiana Center for Musculoskeletal Health (ICMH) Cancer Team Trainee Pilot Award.