Experimental Study On Cerebral Protective Effects Of Retrograde Cerebral Perfusion Via Superior Vena Cava Combined Edaravone During Deep Hypothermic Circulatory Arrest

Objective: To investigate the cerebral protective effects of Retrograde Cerebral Perfusion (RCP) combined a free radical scavenger edaravone(EDA) during deep hypothermic circulatory arrest, and explore its possible mechanism, thereby providing rational clinical application.Methods: Using rabbit deep hypothermic circulatory arrest (DHCA) model, twenty four healthy rabbits were randomly assigned toⅠ,Ⅰ,Ⅰthree groups. GroupⅠ(DHCA): After establishment of cardiopulmonary bypass, when the rectal temperature was reduced to 20℃, the circulation was arrested for 90 min. GroupⅠ( RCP):DHCA with temperature reduced to 20℃, then with retrograde perfusion through superior vena cava(SVC) for 90 min. GroupⅠ(RCP combined EDA): The edaravone was added in the prime solution at 3mg/kg, then DHCA with RCP through superior vena cava for 90 min. Each group was rewarmed slowly to 37℃. Monitering HR,MAP,pH,HCT and artery blood gas analysis. To observe the ultrastructure of brain cells using Transmission Electron Microscopy(TEM). To determine the Lactic Acid content using Lactic Acid Kit and spectrophotometric method. To detect the content of cerebral ATP applying high performance liquid chromatograpy(HPLC). Determination of the Superoxide Dismutase(SOD) activity and Malondialdehyde(MDA) content using SOD Kit and MDA Kit. To detect apoptosis by Terminal Deoxynucleotide Transferase-mediated Dexyuridine-biofin Nick End Labeling(TUNEL) method. method. Applying Immunohistochemistry analysis to detect the expression of Caspase-3 and Bcl-2.Results:(1) There was no statistically significant difference in HR,MAP,pH,HCT and artery blood gas analysis(P>0.05).(2) In groupⅠ, mass cells vacuolation and pycnosis degeneration and pile of spongiform change of white matter were found. A small amount of cells vaculolation and slight edema around interstitial cells and vessels in white matter was observed in groupⅠ, and in groupⅠonly early period near hypoxia change and brain constitution little swelling and edema were found. With transmission electron microscope, in DHCA group, hypoxia-ischemia changed manifestly as significant swelling of neuron and mitochondria, heavy hypoxia changes, widely doffing beads of rough endoplasmic reticulum, dispersion and fragmentation of organelles, and abnormal changes of nucleus. In RCP group, it showed a little swelling of neuron, mitochondria and rough endoplasmic reticulum, most of the normal chromatin and nucleoli. In RCP combined EDA group, small hypoxia changes were characterized by gentle edema of neuron, gentle swelling of mitochondria and rough endoplasmic reticulum, normal chromatin and nucleoli, and nearly intact cell.(3) Jugular vein Lactic Acid content of each group showed no statistically significant difference in CPB initiation and reducing temperature period(P>0.05). Compared with the beginning of rewarming and 37℃, Jugular vein Lactic Acid content of GroupⅠ(DHCA) was quite higher than GroupⅠ(RCP) and GroupⅠ(RCP combined EDA)(P<0.05), but no difference between GroupⅠand GroupⅠ(P>0.05). Brain constitution ATP content of each group was similar in CPB initiation(P>0.05). Rewarming to 37℃, the ATP content of each group was significantly different(P<0.05), which GroupⅠh ighest, GroupⅠf ollowed, and GroupⅠlowest. Compared with CPB initiation and 37℃in each group, the ATP content of GroupⅠand GroupⅠdecreased markedly(P<0.05), but GroupⅠvaried little(P>0.05).(4) Compared with CPB initiation and 37℃in each group, the Superoxide Dismutase(SOD) activity was significantly different(P<0.05). The SOD activity of each group showed no statistically significant difference in CPB initiation(P>0.05). However, when rewarming to 37℃, the SOD activity of each group was manifestly different(P<0.05), which RCP combined EDA group highest, RCP group second, and DHCA group lowest. The MDA content of each group showed no statistically significant difference in CPB initiation(P>0.05), but when rewarming to 37℃, the MDA content of each group significantly varied(P<0.05), which DHCA group highest, RCP group followed, and RCP combined EDA group lowest. Compared with CPB initiation and 37℃in each group, the MDA content of DHCA group and RCP group varied manifestly(P<0.05), but which showed in RCP combined EDA no statistically significant difference(P>0.05).(5) TUNEL showed there were the most positive cells in DHCA group than others(p<0.05). The amount of positive cells in RCP group was much more than in RCP combined EDA (p<0.05).(6) When rewarming to 37℃, the expression of Caspase-3 protein of each group significantly varied(P<0.05), which DHCA group highest, RCP group followed, and RCP combined EDA lowest. While rewarming to 37℃, the expression of bcl-2 protein was lowest in DHCA group than the other(p<0.05), and that was much higher in RCP combined EDA than in RCP group (p<0.05). Compared with CPB initiation and 37℃in each group, The expression of bcl-2 protein of RCP group and RCP combined EDA edhanced manifestly(P<0.05), but which reduced significantly in DHCA group (P<0.05).Conclusion: (1) RCP combined a free radical scavenger EDA during deep hypothermic circulatory arrest can supply oxygenated blood to the brain, but also reduce the toxic effects of large amounts of free radicals from hypoxic cerebral cells, which can reduce irreversible damage of the ultrastructure of neuron and reduce neuronal apoptosis in cerebral cortex. (2) RCP combined EDA during deep hypothermic circulatory arrest can significantly inhibit the neur apoptosis by downgrading the expression of Caspas-3 protein and upgrading the expression of Bcl-2 protein. (3) RCP combined EDA has better effect of cerebral protection compared to single RCP via the superior vena cava, which is a effective method of cerebral protection in the aortic arch surgery involved atherosclerosis.