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PFKFB3-mediated glycolysis rescues myopathic outcomes in the ischemic limb
Terence E. Ryan, … , Espen E. Spangenburg, Joseph M. McClung
Terence E. Ryan, … , Espen E. Spangenburg, Joseph M. McClung
Published August 25, 2020
Citation Information: JCI Insight. 2020;5(18):e139628. https://doi.org/10.1172/jci.insight.139628.
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Research Article Muscle biology Vascular biology

PFKFB3-mediated glycolysis rescues myopathic outcomes in the ischemic limb

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Abstract

Compromised muscle mitochondrial metabolism is a hallmark of peripheral arterial disease, especially in patients with the most severe clinical manifestation — critical limb ischemia (CLI). We asked whether inflexibility in metabolism is critical for the development of myopathy in ischemic limb muscles. Using Polg mtDNA mutator (D257A) mice, we reveal remarkable protection from hind limb ischemia (HLI) due to a unique and beneficial adaptive enhancement of glycolytic metabolism and elevated ischemic muscle PFKFB3. Similar to the relationship between mitochondria from CLI and claudicating patient muscles, BALB/c muscle mitochondria are uniquely dysfunctional after HLI onset as compared with the C57BL/6 (BL6) parental strain. AAV-mediated overexpression of PFKFB3 in BALB/c limb muscles improved muscle contractile function and limb blood flow following HLI. Enrichment analysis of RNA sequencing data on muscle from CLI patients revealed a unique deficit in the glucose metabolism Reactome. Muscles from these patients express lower PFKFB3 protein, and their muscle progenitor cells possess decreased glycolytic flux capacity in vitro. Here, we show supplementary glycolytic flux as sufficient to protect against ischemic myopathy in instances where reduced blood flow–related mitochondrial function is compromised preclinically. Additionally, our data reveal reduced glycolytic flux as a common characteristic of the failing CLI patient limb skeletal muscle.

Authors

Terence E. Ryan, Cameron A. Schmidt, Michael D. Tarpey, Adam J. Amorese, Dean J. Yamaguchi, Emma J. Goldberg, Melissa M.R. Iñigo, Reema Karnekar, Allison O’Rourke, James M. Ervasti, Patricia Brophy, Thomas D. Green, P. Darrell Neufer, Kelsey Fisher-Wellman, Espen E. Spangenburg, Joseph M. McClung

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Figure 6

PFKFB3 expression increases glycolytic flux in skeletal muscle and endothelial cells in vitro, resulting in enhanced hypoxia tolerance and angiogenesis.

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PFKFB3 expression increases glycolytic flux in skeletal muscle and endot...
Skeletal muscle cells (myotubes) and HUVECs (endothelial cells) were treated with control (AAV-GFP) and overexpression (AAV-PFKFB3) constructs. (A and B) Treatment was validated at both the mRNA (A, n = 3/group) and protein (B) level. (C and D) PFKFB3 overexpression increased both basal and maximal glycolytic flux in skeletal muscle cells (C, n = 7–8/group) resulting in improved cell survival/viability in hypoxia (D, n = 4/group). (E–G) PFKFB3 overexpression also increased both basal and maximal glycolytic flux in endothelial cells (E and F, n = 3–4/group), which resulted in enhanced endothelial cell tube formation (G, n = 3-4/group), an in vitro model of angiogenesis. Increased angiogenesis could be blocked by treatment with PFK15 (inhibitor of PFKFB3). **P < 0.01, ***P < 0.001, and ****P < 0.0001 versus control (AAV-GFP or DMSO) using ANOVA with Tukey’s post hoc for comparisons. φP < 0.05 for virus effect (E) using ANOVA (1-way in A and G, 2-way in C–E) with Tukey’s post hoc for comparisons. Values are presented as mean ± SEM.

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