Novel Gastrocnemius Muscle Characteristics in Peripheral Artery Disease Patients Associated with Impaired Functional Performance

Lower extremity peripheral artery disease (PAD) significantly affects aging populations and results in functional impairment and mobility loss. Although the clinical importance of finding efficacious interventions for PAD is well-recognized, few medical therapies are currently available. PAD is diagnosed using the ankle brachial index (ABI), a measure of blood flow to the lower extremities, and lower ABI is associated with worse function, however, low ABI alone cannot fully explain functional impairments in PAD. Small studies have reported oxidative stress, mitochondrial dysfunction and fiber damage in gastrocnemius muscle biopsies from PAD patients, suggesting skeletal muscle perturbations may contribute to functional decline. We reported highly variable fiber type composition and fiber type grouping in a small cohort of PAD patients, and lack of intermyofibrillar mitochondria (IMFM-) in the center of oxidative, myosin heavy chain (MyHC) type I fibers. We have provocative new preliminary data suggesting variability in response to ongoing denervation, and in fiber type and mitochondrial adaptations in PAD. The purpose of this study is to define specific characteristics of muscle in PAD associated with impaired walking performance through detailed immunohistochemical analyses of approximately 300 baseline gastrocnemius muscle biopsies stored in the Northwestern biorepository collected from 12 different clinical trials. This biorepository of muscle from PAD patients is one-of-a-kind and is associated with clinical and functional characteristics of the donors. We hypothesize that variability in fiber size, fiber type and mitochondrial adaptations in response to oxidative stress-induced damage and denervation across individuals with PAD will be associated with walking performance. In Aim 1, we will quantify the proportion of type I fibers with IMFM- areas that show normal type I MyHC abundance, accumulate type IIX MyHC and/or LC3, a marker of autophagy, and determine associations of these attributes with overall fiber type composition and fiber size in PAD. We hypothesize that LC3 will colocalize with IIX MyHC in IMFM- areas, suggesting incomplete autophagic clearance of IIX MyHC and mitochondrial biogenesis during fiber type transition from type IIX to type I as a result of denervation and reinnervation. In Aim 2 we will quantify denervated, NCAM positive fibers and fibers with elevated oxidative damage markers by fiber type. We hypothesize that denervation in PAD will preferentially affect fibers expressing IIX MyHC and that only IMFM-areas that accumulate IIX MyHC will be NCAM+. In Aim 3 we will correlate disease severity and walking performance with morphological characteristics of muscle in patients with PAD. Statistical modelling will be used to associate muscle characteristics, quantified in Aims 1 and 2, with function, and identify those most likely to contribute to disease severity and functional impairment. Identification of muscle adaptations associated with more favorable functional performance may facilitate development of targeted strategies to treat PAD and lay the foundation for future longitudinal studies examining the relationship between muscle properties and disease progression and variable response to pharmacologic and/or exercise interventions in patients with PAD.