The Effect Of Bcl-2and Caspase-3on The Cortical And Hippocampal Neuronal Damage In AD Rat Model Induced By Aβ1-42Oligomers

Objective: To investigate the role of Bcl-2/Caspase-3in AD rat model neuronaldamage which induced by Aβ1-42oligomers microinfusion into right lateralcerebral ventricle.Methods: After screening with Morris water maze,60ratswith escape latency less than60s were randomly assigned equally into fourgroups (15rats in each group).there are normal group, PBS control group,Aβ1-42fiber group, Aβ1-42oligomers group. According to the《brain stereotaxicatlas》,using the stereotaxic instrument positioned the rat right lateral cerebralventricle,and then fix the micro-dosing device in the right lateral cerebralventricle. Through the micro-dosing system, Aβ1-42fiber(concentration1μg/μl) or Aβ1-42oligomers(concentration1μg/μl) was injected into the rightlateral cerebral ventricle of rats, daily inject5μg, administration5days for preparation AD rat model; the same method was used in PBS control group togive the same volume of PBS; normal group without any treatment. Fourth weekafter modeling,morris water maze was performed to detect the changes in theability of learning and memory in rats. After the behavioral testing,4rats wererandomly chosen from each group to do the heart perfusion with4%paraformaldehyde,removed the skull carefully, and then separated the braintissue with glass dissecting needle, made brain tissue paraffin slice. Theremaining rats were killed by enuleation of eyeball, rapid peeled the rat’s cortexand hippocampus on the ice tray, stored at-80℃。RT-PCR method was used todetect the expression of Bcl-2mRNA and Caspase-3mRNA of the cortex andhippocampus in AD rats, and western blot assay was carried out to detect Bcl-2protein relative expression and the activity of Caspase-3protein in the rats’cortex and hippocampus. Used HE staining and optical microscope to observethe morphological changes of rats’ cortex and hippocampus. The data wasanalyzed with SPSS19.0statistical software,the results were recorded as x±s,P<0.05showed a statistically significant difference. Results:(1) Morris watermaze behavioral test results: before modeling, there was no significantdifference in the four groups of rats escape latency period (P>0.05). Aftermodeling,the escape latency were prolonged than before moldeling(P<0.05) inAβ1-42oligomers group rats and Aβ1-42fiber group rats; while the normalgroup and PBS control group rats’ escape latency showed no statistical significance before and after modeling(P>0.05);compared with normal group（26.200±3.985）and PBS control group（28.000±7.906）,the escape latencyin Aβ1-42oligomers group （49.200±7.517）and Aβ1-42fiber group（37.200±5.937）were prolonged(P<0.05);the escape latency in Aβ1-42oligomers group was much longer than Aβ1-42fiber group(P<0.05).(2)Bcl-2mRNA、Caspase-3mRNA test results: compared with the normal groupand PBS control group, the expression levels of Bcl-2mRNA in the cortex andhippocampus were decreased (P<0.05) in the model group rats; while theexpression level of Bcl-2mRNA in Aβ1-42oligomers group rats’ cortex andhippocampus decreased more significantly than A β1-42fibergroup(P<0.05);the expression of Caspase-3mRNA in Aβ1-42oligomers groupand Aβ1-42fiber group were increased (P<0.05) compared with normal grouprats’ cortex and hippocampus, the change was much more obvious in Aβ1-42oligomers group; the levels of Caspase-3mRNA were no statistical significance(P>0.05) between normal group and PBS control group rats’ cortex andhippocampus.(3) Western blot results: compared with normal group and PBScontrol group,the relative expression of Bcl-2protein was decreasedsignificantly（P<0.05） and the activity of Caspase-3protein was increasedobviously(P<0.05) among AD model group rats’ cortex and hippocampus, thesedifferences were statistically significant; while the Aβ1-42oligomers groupchanged more significant; there were no significant difference(P>0.05) between normal group and PBS group rats’ Bcl-2protein relative expression andCaspase-3protein activity of cortex and hippocampus.(4) HE staining results:the nuclei of the normal group and PBS group rats’ cortical neurons stainedblue,obvious nucleolus and abundant cytoplasm; the nuclei of Aβ1-42fiberrats’ cortical neurons are hyperchromatic,nuclear membrane structure is notclear; Aβ1-42oligomers rats cortical neurons injury are more serious, nucleistaining was dark purple,dense cytoplasm, nuclear chromatin margination. Thenormal group and the PBS group rats’ hippocampal neuron number was rich, thenuclei were oval or round,staining was light blue, dyeing uniformity, nucleolusis obvious; Aβ1-42fiber group rats hippocampal neuronal nuclear stainingdeeper than normal group; irregular shape; Aβ1-42oligomers group ratshippocampal neurons were in disorder, cell body shrinkage, nuclear staining wasdeep blue purple,nuclear chromatin condensation, dense, and deep stained.Conclusions:(1) The modeling method of perfuse Aβ1-42into rats’ right lateralcerebral ventricle can successfully establish more ideal AD rat model.(2) Aβ1-42oligomers and Aβ1-42fiber all can cognitive dysfunction in rat,but Aβ1-42oligomers is more seriously affect the brain’s learning and memory in rats.(3) A β1-42oligomers and A β1-42fiber can inhibit Bcl-2expression,up-regulate the activity of Caspase-3in rat’s cortex andhippocampus.(4)Aβ1-42oligomers and Aβ1-42fiber can cause damage toneurons in rat’s cortex and hippocampus,in which Aβ1-42oligomers has much greater neuronal toxicity.(5) Bcl-2and Caspase-3play an important role in rat’sneuronal damage which induced by Aβ1-42oligomer, the mechanism of actionis worth in-depth study.