Bone viability of amputated limbs treated with hypothermia: assessment by evaluation of mRNA levels.
We evaluated rat bone viability using a bone viability index (BVI). To evaluate hypothermic ischaemic bone injury, 21 amputated hind limbs of Fischer rats were preserved at hypothermia (4 degrees C) for 1, 3 and 6 hours. To evaluate hypothermic ischaemia/reperfusion injury, another 28 amputated limbs were transplanted to recipient rats after hypothermic ischaemia for 3 and 6 hours, respectively. Total RNA isolated from each tibia was fractionated by electrophoresis and hybridised with 32P-labelled cDNA of GAPDH, and the radioactivity of intact and degraded GAPDH mRNA measured. BVI was calculated as follows, BVI = [A / (A + B)] x 100, where A and B represent the radioactivities corresponding to intact and degraded GAPDH mRNA bands, respectively. In the hypothermic ischaemic insult group, BVIs were comparable to those of controls. However, in the 3-hour hypothermic ischaemia/reperfusion group, BVI was lower than that of the controls. Likewise, there was a significant difference between the 6-hour ischaemia/reperfusion group and controls. These results showed that bone viability decreased even after just a 3-hour hypothermic ischaemia/reperfusion.
Department of Orthopedic Surgery, Nara Medical University, Shijo 840, Kashihara, Nara 634-8522, Japan. makahane@naramed-u.ac.jp
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M
Akahane
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H
Ono
H
H
Ohgushi
Y
Y
Takakura
D000818:Animals; D015152:Blotting, Northern; D001842:Bone and Bones; D007036:Hypothermia, Induced; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011916:Rats, Inbred F344; D015427:Reperfusion Injury / Q000473:pathology*; D012092:Replantation; D014022:Tissue Survival
D012333:RNA, Messenger