Abstract 19756: Association of Increased PTEN-Induced Putative Kinase 1 (PINK1) Expression and Oxidative Damage in Gastrocnemius Myofibers of Patients with Peripheral Arterial Disease

Abstract

Introduction: Oxidative damage (OxD) determined as protein adducts of 4-hydroxy-2-nonenal (HNE) is increased in gastrocnemius myofibers of patients with peripheral arterial disease (PAD) and correlates with disease progression. Mitochondria in PAD gastrocnemius exhibit structural damage and respiratory dysfunction; however, the extent of mitochondrial damage and its relationship to OxD have not been determined. PINK1 is a glycoprotein normally expressed in low abundance on the mitochondrial external membrane. Mitochondrial damage signaled as loss of ion potential across the external membrane produces a marked increase in PINK1, which initiates mitophagy. In this study, we tested the hypothesis that mitochondrial damage is 1) prevalent among gastrocnemius myofibers of PAD patients and 2) linked to OxD. Methods: Gastrocnemius biopsies were collected from five PAD patients with claudication and five controls matched on the basis of age, gender, smoking history and type II diabetes. Biopsy specimens were fixed in methacarn, embedded in paraffin, sectioned and mounted to glass slides. Fluorescence labeled PINK1 protein and HNE adducts in individual myofibers, were quantified by quantitative fluorescence microscopy and expressed as mean pixel intensity in gray scale units (gsu) corrected for background. Results: PINK1 protein expression was increased (p=0.008) in PAD (mean ± S.E.M.; 494.78 ± 0.02 gsu) compared to control myofibers (102.45 ± 0.02 gsu) of the gastrocnemius. PINK1 was increased in 93% of the PAD myofibers. The association of increased (greater than two standard deviations above the control means) PINK1 expression and HNE adducts was determined for PAD myofibers. HNE adducts were increased in 67% of a random sample of PAD fibers expressing increased PINK1. Conclusions: Increased expression of PINK1 protein in the gastrocnemius myofibers of PAD patients with claudication, has established the presence of irreversible mitochondrial damage in a large proportion of myofibers. Elevation of OxD in 67% of PAD myofibers expressing increased PINK1 links OxD and mitochondrial damage. The data are consistent with a contribution of OxD to myofibers in PAD gastrocnemius, by damaged mitochondria.