Reversibility of muscular ischemia: a histochemical quantification by the nitroblue tetrazolium (NBT) test

The degree of muscular ischemia and its reversibility can be quantified in the early stages. This histochemical enzymatic study utilized Nitroblue tetrazolium (NBT) which when reduced by tissue dehydrogenase produces a blue pigment: "formazan." Seventy Wistar rats were subjected to transient hindlimb ischemia by means of a tourniquet for 3, 6, 9, 12, 15 and 18 hours, followed by reperfusion. Microsurgical muscle biopsies were obtained in each rat at 1 and 12 hours, and 3, 7, 14 days after reperfusion. Time increased in muscle staining demonstrated a succino-dehydrogenase deficit confirmed by clinical and histopathological follow-up. NBT staining time was 2 minutes (+/- 8 sec.) in the control group, between 2 and 6 minutes in the reversible ischemia group (rats with 3 and 6 hours of tourniquet), and more than 9 minutes (+/- 14 sec.) in the irreversible ischemia group (animals with more than 9 hours of tourniquet). In vascular surgery and in limb reimplantation this protocol is a practical method of evaluating cytoplasmic enzymatic activity and the status of myofibrillar oxidation in the early phases of ischemic injury, before histologic changes are clearly delineated.