In reconstructive surgery, skeletal muscle may endure protracted
ischemia before reperfusion, which can lead to significant
ischemia/reperfusion injury.
Ischemic postconditioning induced by brief cycles of reperfusion/reocclusion at the end of
ischemia has been shown to salvage skeletal muscle from
ischemia/reperfusion injury in several animal models. However,
ischemic postconditioning has not been confirmed in human skeletal muscle. Using an established in vitro human skeletal muscle hypoxic conditioning model, we tested our hypothesis that hypoxic postconditioning salvages ex vivo human skeletal muscle from
hypoxia/reoxygenation injury and the mechanism involves inhibition of opening of the
mitochondrial permeability transition pore (mPTP) and preservation of
ATP synthesis. Muscle strips (~0.5×0.5×15mm) from human rectus abdominis muscle biopsies were cultured in Krebs-Henseleit-
HEPES buffer, bubbled with 95%N(2)/5%CO(2) (
hypoxia) or 95%O(2)/5%CO(2) (reoxygenation). Samples were subjected to 3h
hypoxia/2h reoxygenation. Hypoxic postconditioning was induced by one or two cycles of 5min reoxygenation/5min
hypoxia after 3h
hypoxia. Muscle injury, viability and
ATP synthesis after 2h of reoxygenation were assessed by measuring
lactate dehydrogenase (LDH) release, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium
bromide (MTT) reduction and
ATP content, respectively. Hypoxic postconditioning or treatment with the
mPTP-opening inhibitors
Cyclosporine A (CsA, 5×10(-6)M) or N-Methyl-4-isoleucine
Cyclosporine (NIM811, 5×10(-6)M) 10min before reoxygenation decreased LDH release, increased MTT reduction and increased muscle
ATP content (n=7 patients; P<0.05). Conversely, treatment with the
mPTP opener
Atractyloside (5×10(-6)M) 10min before hypoxic postconditioning abolished its protective effect (n=7 patients; P<0.05). We conclude that hypoxic postconditioning effectively salvages human skeletal muscle from
hypoxia/reoxygenation injury by inhibition of
mPTP opening and preservation of
ATP synthesis during reoxygenation.