Noninvasive ⁶⁴Cu-ATSM and PET/CT Assessment of Hypoxia in Rat Skeletal Muscles and Tendons During Muscle Contractions

Abstract

The purpose of the present study was to investigate exerciserelated changes in oxygenation in rat skeletal muscles and tendons noninvasively with PET/CT and the hypoxia-selective tracer 64 Cu-diacetyl bis(N 4 -methylthiosemicarbazone) (ATSM) and to quantitatively study concomitant changes in gene expression of 2 hypoxia-related genes, hypoxia-inducible factor 1a (HIF1a) and carbonic anhydrase III (CAIII). Methods: Two groups of Wistar rats performed 1-leg contractions of the calf muscle by electrostimulation of the sciatic nerve. After 10 min of muscle contractions, 64 Cu-ATSM was injected and contractions were continued for 20 min. PET/CT of both hind limbs was performed immediately and 1 h after the contractions. The exercise group (n 5 8) performed only muscle contractions as described, whereas the other group, exercise plus cuff (n 5 8), in addition underwent cuff-induced hypoxia during the first PET/CT scan. Standardized uptake values (SUVs) were calculated for the Achilles tendons and triceps surae muscles and were correlated to gene expression of HIF1a and CAIII using real-time polymerase chain reaction. Results: Immediately after the contractions, uptake of 64 Cu-ATSM was significantly increased, by approximately 1.5-fold in muscles and 1.3-fold in tendons, compared with resting conditions. The significant increase was maintained in late PET scans in stimulated muscles and tendons independently of cuff application. In muscles, SUV correlated significantly with gene expression of HIF1a and CAIII, whereas this coherence was not found in tendons. Conclusion: We found enhanced uptake of 64 Cu-ATSM in both early and late PET scans, thereby supporting the possibility that 64 Cu-ATSM registers exercise-induced transient hypoxia in both skeletal muscles and force-transmitting tendons. The fact that skeletal muscles but not tendons showed upregulation of HIF1a and CAIII could indicate that healthy tendons are less responsive than skeletal muscles to low levels of oxygen. At the onset of exercise, intracellular oxygen decreases substantially in the contracting skeletal muscles. This decrease is believed to induce immediate changes at the systemic and local levels and to constitute a main signal for more long-term muscular adjustments to exercise, such as increased skeletal muscle angiogenesis (1,2). It is unclear whether the force-transmitting tendons exhibit a similar decrease in intracellular oxygen during loading and to what extent any decrease would affect the adaptive mechanisms of the connective tissue.It has been suggested that hypoxia plays a role in the development of tendon overuse injuries, in which a decreased blood flow simultaneous with an increased activity in tendons results in local tissue hypoxia, impaired nutrition, and energy metabolism (3). In support of this suggestion have been histologic examinations of specimens removed during surgery for tendinopathy and showing hypoxic degeneration (4). Furthermore, painful Achilles tendinopathy frequently develops in the relatively hypovascular anatomi...

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“…It may seem paradoxical that creating a transient hypoxic milieu would be effective for the treatment of tendinopathy given that tendon tissue hypoxia previously has been suggested as a factor in the pathogenesis of tendinopathies 66,67 . Furthermore, a rodent study indicated that tendons may exert a less pronounced cellular response (impaired upregulation of hypoxiarelated gene expression) in response to acutely reduced oxygen levels as compared to skeletal muscle 68 , which speaks a bit against a hypoxia-mediated effect/mechanism. On the other hand, hypoxia has also been reported to improve cross-linking and mechanical properties of collagenrich tissues 69,70 .…”

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