title: Evaluation and comparison of the effect of hypothermia and ozone on ischemia-reperfusion injury of skeletal muscle in rats.
abstract: BACKGROUND
  Tourniquet-induced ischemia-reperfusion, which affects local and distant organs, is very common in orthopedic surgery. Hypothermia is used in traumatic tissue during ischemic period commonly. Ozone (O3) has been recommended as a novel therapeutic agent in various medical conditions. The objective of the study was to evaluate and compare the effect of hypothermia (H) and O3 on ischemia-reperfusion injury of skeletal muscle in rats by measuring oxidative parameters and inducible nitric oxide synthase (iNOS) levels.
  
  
  METHODS
  Eighteen rats (Wistar albino) were separated into five groups randomly (sham, IR, IR + H, IR + O3, IR + H + O3; n = 6). The lower right extremity of all rats was subjected to 2 h of ischemia and 22 h of reperfusion clamping the common iliac artery and using the rubber-band technique at the level of the lesser trochanter under general anesthesia. Two hours of hypothermia were applied during the first 2 h of reperfusion in two groups. O3 was applied in two groups. All rats were sacrificed after the IR period with high dose of anesthesia. The tibialis anterior muscle and blood were saved. Levels of superoxide dismutase, glutathione peroxidase, MDA, NOx, and interleukin-1β were measured in the muscle. Creatinine kinase, lactate dehydrogenase, aspartate aminotransferase, urea, creatinine, and electrolytes were measured in serum. Immunohistochemical iNOS staining was performed on muscle samples.
  
  
  RESULTS
  The levels of MDA, NOx, and interleukin-1β in muscle were raised in the IR group compared with those in the sham group. The same parameters were lower in the groups of IR + H, IR + O3, and IR + H + O3 compared with those in the IR group. Superoxide dismutase and glutathione peroxidase activities in muscle were lower in the IR group compared with those in the sham group; however, same parameters were higher in the groups of IR + H, IR + O3, and IR + H + O3 compared with those in the IR group. Score and intensity of iNOS staining in skeletal muscle in the IR group was increased compared with that in the sham group and decreased in the groups of IR + H, IR + O3, and IR + H + O3 compared with that in the IR group. Levels of creatinine kinase, aspartate aminotransferase, and K in the three treatment groups decreased compared with those in the IR group.
  
  
  CONCLUSIONS
  These findings showed that hypothermia, which has more affect, and O3 decreased the tourniquet-induced IR injury in the rat's muscle-skeletal system by reducing the levels of oxidative and nitrosative stress parameters and enhancing antioxidant enzymes. Hypothermia and O3 had no synergistic effect. Hypothermic reperfusion and O3 preconditioning might be beneficial in skeletal muscle IR injury-associated tourniquet.
authors.affiliation: Department of Orthopedic Surgery, Gulhane Military Medical Faculty, Ankara, Turkey.
authors.forename: Huseyin
authors.initials: H
authors.lastname: Ozkan
authors.affiliation: Department of Orthopedic Surgery, Agrı Military Hospital, Agrı, Turkey. Electronic address: safakekinci@yahoo.com.
authors.forename: Safak
authors.initials: S
authors.lastname: Ekinci
authors.affiliation: Department of Physiology, Gulhane Military Medical Faculty, Ankara, Turkey.
authors.forename: Bulent
authors.initials: B
authors.lastname: Uysal
authors.affiliation: Department of Orthopedic Surgery, Malatya Military Hospital, Malatya, Turkey.
authors.forename: Faruk
authors.initials: F
authors.lastname: Akyildiz
authors.affiliation: Department of Orthopedic Surgery, Gulhane Military Medical Faculty, Ankara, Turkey.
authors.forename: Selim
authors.initials: S
authors.lastname: Turkkan
authors.affiliation: Department of Orthopedic Surgery, Erzurum Military Hospital, Erzurum, Turkey.
authors.forename: Omer
authors.initials: O
authors.lastname: Ersen
authors.affiliation: Department of Orthopedic Surgery, Gulhane Military Medical Faculty, Ankara, Turkey.
authors.forename: Kenan
authors.initials: K
authors.lastname: Koca
authors.affiliation: Department of Sport Medicine, Gulhane Military Medical Faculty, Ankara, Turkey.
authors.forename: Mehmet Murat
authors.initials: MM
authors.lastname: Seven
mesh_terms: D000818:Animals; D007036:Hypothermia, Induced; D008297:Male; D018482:Muscle, Skeletal / Q000098:blood supply*; D018384:Oxidative Stress; D010126:Ozone / Q000627:therapeutic use*; D011897:Random Allocation; D017208:Rats, Wistar; D015427:Reperfusion Injury / Q000628:therapy*
chemical_list: D010126:Ozone
keywords: GSH-Px MDA; Hypothermia; IL-1B; IR injury; NOx; Ozone; SOD; iNOS