Understanding The Role Of Exercise In Lymphatic Vessel Regulation In Melanoma

Exercise has been demonstrated as an effective method to remodel tumor blood vessels and to improve immune cell infiltration in multiple mouse tumor models. However, it is unclear whether aerobic exercise alters lymphatic vasculature in tumors. The lymphatic system is important in tumor growth and metastasis because it regulates interstitial fluid pressure via lymph flow and immune cell trafficking, in conjunction with blood vessels. Lymphatic vessels are particularly important during metastasis because tumor cells escape the primary tumor through lymphatic vasculature to establish primary metastasis in the lymph nodes. Here, we evaluated the effect of aerobic exercise on lymphatic vessels and endothelial cell junctions in melanoma in male C57BL/6 J mice. BP melanoma cells from transgenic mice with BrafV600E/+ and Pten−/− mutations were injected subcutaneously. Mice were assigned to no exercise and exercise groups when tumors reached ~30 mm3. The exercise group completed treadmill running at 12 meters/minute for 45 minutes for 5 consecutive days with 2 rest days per week for 2 weeks. Tumor lymphatic endothelial cells were identified by LYVE-1, endothelial tight junctions by ZO-1, and nuclei by DAPI using immunofluorescence on frozen tumors. The number of total lymphatic vessels, elongated vessels( >100 μm), open lumens, and co-localization of ZO-1 with LYVE-1 was quantified on 6 random images per tumor using NIH Image J software. ANOVA was used for statistical analysis(p < 0.05). Tumors from exercised mice had higher lymphatic vessel density (p = 0.002) and more open lumens (p = 0.001). Surprisingly, tumors from exercised mice had fewer ZO-1 positive vessels (p = 0.015) and lower co-localization of ZO-1 and LYVE-1 (p = 0.0185) than tumors from non-exercised mice, suggesting fewer stable lymphatic vessel endothelial cell junctions. The higher lymphatic vessel density and open lumens present in tumors from exercised mice may contribute to an exercise-induced improved anti-tumor immune response, which was previously reported. However, the lower ZO-1 expression on lymphatic endothelial cells in tumors from exercised mice may indicate lymphatic vasculature dysfunction and/or hyper-permeability. Further work is needed to assess the functional changes in tumor lymphatic vasculature and metastasis by exercise. CPRIT Research Training Award CPRIT Training Program (RP210028)