Seven weeks after vehicle or streptozotocin injection, rats (referred to as non-diabetic “n,” and diabetic “d,” respectively) were randomly assigned to the control (CON) or IR group. The control groups (nCON, dCON, 8 rats per group) underwent 5 h of isoflurane anesthesia and similar surgical manipulation to the IR groups, except for hindlimb ischemia (sham-operated). The ischemia–reperfusion groups (nIR and dIR, 10 rats per group) underwent 3 h of ischemia induced by infra-renal aortic occlusion and collateral vessel ligation, followed by 2 h of reperfusion. Ischemia was clinically characterized by cyanosis and lack of an arterial pulse distal to the clamp, and biochemically by an increase in capillary blood lactate measured in the right hindlimb (Lactate Pro device, LT1710; Arkray, KGK, Japan).
After reperfusion, gastrocnemius muscles, that are considered more sensitive to IR (Charles et al., 2017 (link)), were harvested and either analyzed immediately (mitochondrial respiration) or kept in ice or in liquid nitrogen-cooled isopentane. Animals were sacrificed by heart retrieval under deep anesthesia (5% isoflurane).
After reperfusion, gastrocnemius muscles, that are considered more sensitive to IR (Charles et al., 2017 (link)), were harvested and either analyzed immediately (mitochondrial respiration) or kept in ice or in liquid nitrogen-cooled isopentane. Animals were sacrificed by heart retrieval under deep anesthesia (5% isoflurane).
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