AMPKα1 is necessary for extracellular matrix homeostasis in mouse Achilles tendon

Abstract

Tendinopathy is a disorder that affects ~3.5 million people in the US1 and is caused by poor self-repair of tendon. Tendinopathy is characterized by degenerative extracellular matrix (ECM), increased cell density, and poor biomechanical function.2 AMP-activated protein kinase (AMPK), an energy stress sensor, is a potential regulator of ECM remodeling in musculoskeletal tissues.3,4 Aging connective tissues (e.g., skeletal muscle) has exhibited reduced sensitivity to AMPK activation and impaired glucose sensitivity.5 Furthermore, cartilage-specific loss of Prkaa1 (AMPKα1) and Prkaa2 (AMPKα2), resulted in ECM degeneration and cell death.4 Yet, if and how AMPK regulates ECM remodeling and cell survival remains unknown. Because tendon is an ECM-rich tissue with low cell turnover, our objective was to identify the necessity of AMPKα1 for tendon cell maintenance and tissue function throughout the lifespan.In this study, we tested the hypothesis that loss of AMPKα1 in tendon impairs ECM remodeling, cell survival, and mechanical function of mouse Achilles tendons in an age-dependent manner. Wildtype (WT; ScxCre-negative floxed) and Prkaa1fl/fl; ScxCre ( cKO) mice were generated to evaluate Achilles tendon structure and function at 1-, 3-, and 9-months (1M, 3M, 9M) using histological and biomechanical assays. Quantitative data were compared using two-tailed unpaired t-tests or 2 way-ANOVAs with Sidaks multiple comparisons tests, where appropriate. cKO tendons developed ECM degeneration by 3M which further progressed by 9M as calcified deposits (lesions). Additionally, 3M cKO tendons had fewer cells compared to WT tendons (WT: 2244 ± 553 cells/mm2; cKO: 1402± 343 cells/mm2, p=0.0188). Additionally, by 9M, cKO tendons developed larger ECM lesions with a similar cell density compared to WT tendons (WT: 2.50 ± 3.26 % area; cKO: 17.11± 8.88 % area p=0.0450). The mechanical function of cKO tendons was impaired by 3M, as maximum load and stress was significantly reduced compared to WT (WT: 9.599 ± 1.486 mm2; cKO: 5.028± 0.981 mm2, p=0.0007). Additionally, cross-sectional area of cKO tendons was significantly increased compared to WT (WT: 0.172 ± 0.459 mm2; cKO: 0.258± 0.058 mm2, p=0.0373). Loss of AMPKα1 in tendon led to larger yet weaker tendons suggesting deficiencies in ECM quality which was supported by histological evidence of premature calcified deposits. Our conclusions support our hypothesis that AMPK is necessary for tendon homeostasis, cell survival, and mechanical function and is the first study to show maintenance of tendon ECM is dependent on AMPKα1 throughout adulthood.1. Millar, et. al Nat Rev Dis Primers 7, 1 (2021); 2. Fouda, et. al Am J Transl Res 9, 4341-60 (2017); 3. Grieve, et. Al Scand J Med Sci Sports 22, 55-63 (2012); 4. Zhou, S. Sci Rep 7, 43245 (2017); 5. Salminen, et.al Ageing Res 11 230-241(2012) ACKNOWLEDGEMENTS: NSF GRFP (to LH); NIH R01AR079367 (to MLK); P30AR069620. Thanks to Steph Steltzer for assistance with X-ray imaging. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.