Effect Of Isoflurane On Apoptosis Of SH-SY5Y Cells Transfected With APP Gene And The Role Of Inositol-1,4,5-triphosphate Receptors

Objective: The clinical commonly used inhalation anesthetic, isoflurane,can induce neurotoxicity, lead to postoperative cognitive dysfunction,especially in the older patients with neutodegeneration. Alzheimer’sdisease(AD) is a conmmon neurodegenerative disease, β-amyloid precursorprotein (APP) is a gene associated with familial Alzheimer’s disease.Previously research has demonstrated that APP mutation will render cellsvulnerable to isoflurane-induced cytotoxicity. Our previous workdemonstrated that exposure to isoflurane may decrease cell viability and theseeffects may be connected to the disruption of intracellular calciumhomeostasis. Intracellular calcium homeostasis is primarily regulated by thethree protein receptors on the endoplasmic reticulum (ER): inositol1,4,5-trisphosphate receptors (IP3R), ryanodine receptor (RyR) andCa2+-ATPases. The IP3R on the ER membrane can cause nonphysiologicalcalcium release from the ER, this leads to depletion of ER calcium, increaseof cytosolic and mitochondrial calcium, all of which can contribute to cellapoptosis. The aim of this study is to evaluate the effect of isoflurane on theapoptosis of SH-SY5Y cells transfected with APP gene and the role of inositol1,4,5-triphosphate recepters.Methods: The SH-SY5Y cells transfected with APP gene were seeded inculture flasks with the density of1.2×104个/cm2. The cells were randomlydivided into4groups (n=6): control grop (group C), IP3R antagonist group(group X), isoflurane group (group I) and isoflurane+IP3R antagonist group(group I+X). After the cells were cultured for24h and attached to the wall,the cells were cultured routinely in group C, and Xestospongin C100nM(IP3R antagonist) was added to DMEM culture medium in groups X and I+X,and30min later the cells were exposed to1.2%isoflurane for8h in groups I and I+X. The cells were collected for examination of the ultrastructure and fordetermination of cell apoptosis, intracellular free calcium ion concentration[Ca2+]i(by flow cytometry) and expression of IP3reeptor protein (byWestern blot). The apoptosis rate was calculated.Results: Compared with group C, there was no significant change in theapoptosis rate,[Ca2+]ior IP3receptor protein expression in group X (P>0.05),while the cell apoptosis rate and [Ca2+]iwere significantly increased and IP3receptor protein expression was up-regulated in groups I and I+X (P<0.01).Compared with group I, cell apoptosis rate and [Ca2+]iwere significantlydecreased and IP3receptor protein expression was down-regulated in groupI+X (P<0.01). The pathological changes of the cells happened in groups Iand I+X, and the pathological changes were severer in group I than in groupI+X.Conclusions: These results suggest that exposure to1MAC isoflurane for8h causes cell apoptosis, the mechanism is intracellular calcium regulationdisorder-calcium overload, may partly by direct activation of IP3receptor onthe ER membrane, IP3R inhibitor could inhibit this effect partly, IP3receptor isone of the important targets.