We developed a large animal model, with clinically relevant gravitational loading on the lymphatic vasculature, that enables longitudinal tracking of lymphatic function in vivo as well as end-point analysis of vessel function and mechanics. One of two parallel lymphatic vessels in the sheep hind limb was ligated with the contralateral limb acting as an internal control. To determine the changes in the protein expression profiles in the lymphatic muscle cells from the remodeled vessel compared to the control, we expanded isolated LMC in vitro and performed bottom-up proteomic analysis using a Q Exactive Plus mass spectrometer. Label-free quantitative (LFQ) methods directly used the raw spectral data from parallel MS runs to determine relative protein abundances. We used both “MS/MS (MS2) spectral counting” and “precursor MS1 area” methods for label free quantitation and further contrasting the differentially expressed proteomic profiles across the control and remodeled lymphatic muscle samples to determine the alterations in mitochondrial dysfunction and elevated oxidative stress within the lymphatic muscle.