Role of Heme Oxygenase in the Protection Afforded Skeletal Muscle During Ischemic Tolerance

Abstract

Ischemic tolerance (IT) is known to improve resistance to ischemia/reperfusion (I/R)-induced injury; however, the mechanisms remain unknown. The authors hypothesized that induction of heme oxygenase (HO), a heat shock protein, would provide anti-inflammatory benefits during IT, thereby preventing leukocyte-derived I/R injury. Male Wistar rats were randomly assigned to sham (n = 4), I/R (n = 9), preconditioning (PC)+I/R (n = 7), chromium mesoporphyrin, to inhibit HO (CrMP; n = 4), or PC+I/R+CrMP (n = 6) groups. PC consisted of 5 cycles of I/R, each lasting 10 min, induced by tightening a tourniquet placed above the greater trochantor of the hindlimb. Twenty-four hours later, the hindlimb underwent 2 h of no-flow ischemia followed by intravital microscopy during 90 min reperfusion to assess capillary perfusion (#/mm), tissue injury (ratio of ethidium bromide to bisbenzimide labeled cells/100 microm2), leukocyte rolling (Lr, #/1000 microm2), and adhesion (La, #/1000 microm2) in postcapillary venules of the extensor digitorum longus (EDL) muscle. In the I/R group, Lr was significantly increased (7.1 +/- 0.4) compared to sham (3.1 +/- 0.4). PC+I/R increased Lr (10.8 +/- 0.72), which was further exacerbated by the removal of HO (14.2 +/- 1.3). La (7.8 +/- 2.0) was significantly increased compared to sham (2.4 +/- 0.9), while PC returned La back to sham levels (1.9 +/- 0.7). Removal of HO activity, via CrMP, had no significant effect on La (3.9 +/- 0.7). However, CrMP removed the protection to microvascular perfusion (I/R = 9.4 +/- 1.1, PC = 16.6 +/- 1.8, sham = 20.5 +/- 2.8, PC+CrMP+I/R = 12.3 +/- 2.3) and prevented protection from ischemia-induced tissue injury. The data suggest that HO is an important protective mechanism during IT in skeletal muscle, but such protection was by mechanisms other than altered leukocyte-endothelial cell interaction.