Interventional‐ and amputation‐stage muscle proteomes in the chronically threatened ischemic limb

Abstract

Background Despite improved surgical approaches for chronic limb‐threatening ischemia (CLTI), amputation rates remain high and contributing tissue‐level factors remain unknown. The purpose of this study was twofold: (1) to identify differences between the healthy adult and CLTI limb muscle proteome, and (2) to identify differences in the limb muscle proteome of CLTI patients prior to surgical intervention or at the time of amputation. Methods and results Gastrocnemius muscle was collected from non‐ischemic controls (n = 19) and either pre‐interventional surgery (n = 10) or at amputation outcome (n = 29) CLTI patients. All samples were subjected to isobaric tandem‐mass‐tag‐assisted proteomics. The mitochondrion was the primary classification of downregulated proteins (> 70%) in CLTI limb muscles and paralleled robust functional mitochondrial impairment. Upregulated proteins (> 38%) were largely from the extracellular matrix. Across the two independent sites, 39 proteins were downregulated and 12 upregulated uniformly. Pre‐interventional CLTI muscles revealed a robust upregulation of mitochondrial proteins but modest functional impairments in fatty acid oxidation as compared with controls. Comparison of pre‐intervention and amputation CLTI limb muscles revealed mitochondrial proteome and functional deficits similar to that between amputation and non‐ischemic controls. Interestingly, these observed changes occurred despite 62% of the amputation CLTI patients having undergone a prior surgical intervention. Conclusions The CLTI proteome supports failing mitochondria as a phenotype that is unique to amputation outcomes. The signature of pre‐intervention CLTI muscle reveals stable mitochondrial protein abundance that is insufficient to uniformly prevent functional impairments. Taken together, these findings support the need for future longitudinal investigations aimed to determine whether mitochondrial failure is causally involved in amputation outcomes from CLTI.

S100A8 and S100A9 are elevated in chronically threatened ischemic limb muscle and induce ischemic mitochondrial pathology in mice

“…Consistent with our previous reports, 24 , 25 we found that CLTI patients displayed significant impairment in mitochondrial oxidative phosphorylation. In the present study, we used a novel CK clamp system to test the mitochondrial oxidative phosphorylation function across a range of physiologic relevant energy demands to mimic a stress test that ranges from rest to exercise.…”

“…Careful examination of a recently curated proteomics dataset from CLTI limb muscle indicated that S100A8/A9 proteins were significantly elevated in CLTI muscle compared with non-PAD control participants ( Fig 1 , A ). 25 This finding was confirmed using mRNA analysis ( Fig 1 , B ). S100A8/A9 are calcium binding proteins that belong to the S100 family.…”

“…JPST000852 [Japan Proteome Standard Repository]). 25 Although not the focus of our previous study, the protein abundance of S100A8 and S100A9 was significantly higher in the CLTI muscle than in the non-PAD control muscle ( Fig 1 , A ). These results were further confirmed at the mRNA level using real-time polymerase chain reaction.…”

“…We described the tissue processing, mass spectrometry analysis, and bioinformatics for these samples in our previous study. 25 Protein abundance for S100A8 and S100A9 from these proteomics analyses was converted to the Log2 space for group comparisons. For each comparison, protein abundance was analyzed for the group average, standard deviation, two-tailed Student’s t test (equal variance), and a Benjamini-Hochberg false discovery rate-corrected adjusted P value.…”

IGF-1 therapy improves muscle size and function in experimental peripheral arterial disease

“…A component of walking performance is related to muscular strength, power, and endurance. Various groups have documented pathological abnormalities within the skeletal muscle of PAD patients (66)(67)(68)(69)(70)(71)(72)(73)(74)(75)(76)(77)(78)(79)(80). Few clinical studies have employed objective measures of muscle strength or power in PAD patients, particularly in the context of randomized clinical trials.…”