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

Objective: To investigate the cerebral protective effects of Retrograde Cerebral Perfusion (RCP) 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 to Ⅰ , Ⅱ ,Ⅲ three groups. Group I (DHCA): ascending aorta and right atria were cannulated for establishing CPB, temperature was reduced to 30℃, and ascending aorta was obstructed. When temperature reached to 20℃, circulation is arrested for 90 min. Group Ⅱ(Antrograde Cerebral Perfusion, ACP): regular CPB with temperature reduced to 20℃, descending aorta was obstructed, then with antrograde cerebral perfusion through aorta arch for 90 min. Group Ⅲ (RCP): CPB with temperature reduced to 20℃, then with retrograde perfusion through superior vena cava(SVC) for 90 min. Each group was rewarmed slowly after 10 min, to 37 ℃ and lasted for 120 min. Continuously monitering ECG, EEG, MAP, rectal temperature and artery blood gas analysis, determination of the Lactic Acid content using Lactic Acid Kit and spectrophotometric method. Applying high performance liquid chromatograpy (HPLC) to detect the content of cerebral ATP, observing the ultrastructure of brain cells using Transmission Electron Microscopy (TEM) and detecting apoptosis by Terminal Deoxynucleotide Transferase-mediated Dexyuridine-biofm Nick End Labeling (TUNEL) method. Applying Semiquantu RT-PCR and Immunohistochemistry analysis to detect the transcription and expression of Caspase-3 mRNA and Bcl-2/Bax. Results: There was no statistically significant difference in MAP, Hb, Hct and artery bloodand gas analysis. Compared with the beginning of rewarming and 37°C, Jugular vein Lactic Acid content of Group I (DHCA) was quite higher than group II (ACP) and group III (RCP) (p<0.05), but no difference between II (ACP) and group III (RCP). Brain constitution ATP content of each group was similar (p>0.05) in CPB initiation, circulatory arrest 90 min and rewarm to 37°C, ATP content of group I was much lower than others (p<0.05). In group I , mass cells vacuolation and pycnosis degeneration and pile of spongiform change of white matter were found. Earlier period new hypoxia change was observed in group II, and in group III only earlier period neur hypoxia change and brain constitution little swelling and edema were found.. With transmission electron microscope, we can see: in DHCA group, hypoxia-ischemia changes manifested as swelling of neuron, widely doffing beads of rough endoplasmic reticulum, dispersion of organelles, and swelling of mitochondria, heavy hypoxia changes were also found widely and severely as doffing beads of rough endoplasmic reticulum, fragmentation of organelles, severe swelling of mitochondria, severe vacuolation, and abnormal changes of nuclei. In ACP and RCP perfusion groups, light hypoxia changes were characterized by gentle oedema of neuron, gentle swelling of mitochondria and rough endoplasmic reticulum, nearly intact cell structure, and mostly normal chromatin and nucleolus. TUNEL showed there were much more positive cells in group I than others (p<0.05). At the onset of rewarming, the expression of caspase-3 mRNA and protein was significantly higher in group I than that in group II and III (p<0.05), but no significant difference between group II and III (p>0.05), while rewarming 2h, much more higher than start in group I (p<0.05), but no obvious alteration in group II and III(p>0.05). While rewarming begining, the expression of bcl-2 protein was lower in group I than other two (p<0.05). After rwarming 2h, the expression was obviously upgraded in group II and III (p<0.05), but no significant change in group I (p>0.05). The expression of Bax protein was just like caspase-3 protein, significantly higher in group I than that in group II and III (p<0.05), but no significant difference between group II and III(p>0.05), while rewarming 2h, much more higher than start in group I (p<0.05), but no obvious alteration in group II and III (p>0.05).Conclusion: (1) The methods of RCP via superior vena cava not only simplified the complexity of surgical operation, also kept ample perfusion for brain, suited the study of cerebral protection in operation on aorta. (2) RCP via superior vena cava ensured enough cerebral blood supply, relieved the apoptosis of cortex cells, maintain the neuronal normal form, avoided ultrastructural irreversible lesion. (3) The methods of RCP via superior vena cava inhibited the neur apoptosis by downgrading the expression and transcription of caspas-3mRNA. (4) RCP via superior vena cava significantly upgraded the expression of bcl-2 protein, while downgraded the expression of Bax protein, inhibited the occurrence of neur apoptosis.