Oxidative damage and reduction of redox factor-1 expression after transient spinal cord ischemia in rabbits.
OBJECTIVE
The mechanism of spinal cord injury has been thought to be related to the vulnerability of spinal motor neuron cells against ischemia. However, the mechanisms of such vulnerability are not fully understood. We previously reported that spinal motor neurons may be lost by programmed cell death and thus now investigate a possible mechanism of neuronal death with immunohistochemical analysis for 8-hydroxy-2'-deoxyguanosine (8-OHdG) and redox factor-1 (Ref-1).


METHODS
We used a rabbit spinal cord ischemia model with a balloon catheter. The spinal cord was removed at 8 hours, 1, 2, or 7 days after 15 minutes of transient ischemia, and histologic changes were studied with hematoxylin-eosin staining. Western blot analysis for Ref-1, temporal profiles of 8-OHdG and Ref-1 immunoreactivity, and double-label fluorescence immunocytochemical studies were performed.


RESULTS
Most motor neurons were preserved until 2 days but were selectively lost at 7 days of reperfusion. Western blot analysis of a sample from sham control spinal cord showed a characteristic 37-kDa band that was reduced after ischemia. Immunohistochemistry showed the nuclear expression of Ref-1 in motor neurons of control spinal cords, and immunoreactivity was decreased 1 day after ischemia. On the other hand, no nuclear expression was seen of 8-OHdG in motor neurons of control spinal cords, and immunoreactivity was increased 1 day after ischemia. Double-label fluorescence immunocytochemical study revealed that both 8-OHdG and Ref-1 were positive at 8 hours of reperfusion in the same motor neurons, which eventually die.


CONCLUSIONS
These results suggest that Ref-1 decreased in motor neurons after transient spinal cord ischemia and that this reduction preceded oxidative DNA damage. The reduction of Ref-1 protein at the moderately late stage of reperfusion may be one of the factors responsible for the delay in neuronal death after spinal cord ischemia.
Sakurai
Masahiro
M
Departments of Cardiovascular Surgery, Tohoku University Graduate School of Medicine, Sendai, Japan.
Nagata
Tetsuya
T
Abe
Koji
K
Horinouchi
Takashi
T
Itoyama
Yasuto
Y
Tabayashi
Koichi
K
D000080242:8-Hydroxy-2'-Deoxyguanosine; D000818:Animals; D017209:Apoptosis / Q000276:immunology*; D019757:Carbon-Oxygen Lyases / Q000032:analysis*; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000032:analysis*; D004195:Disease Models, Animal; D002546:Ischemic Attack, Transient / Q000209:etiology* / Q000276:immunology* / Q000473:pathology; D009422:Nervous System Diseases / Q000209:etiology* / Q000276:immunology* / Q000473:pathology; D009474:Neurons / Q000276:immunology / Q000473:pathology; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000276:immunology*; D011817:Rabbits; D012720:Severity of Illness Index; D013119:Spinal Cord Injuries / Q000150:complications* / Q000276:immunology* / Q000473:pathology; D013997:Time Factors
D000080242:8-Hydroxy-2'-Deoxyguanosine; D019757:Carbon-Oxygen Lyases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D003849:Deoxyguanosine