Studies On The Correlation Between FOXO3a And Glioma

Objective1. To investigate the expression of FOXO3a in human gliomaand the correlation with proliferation marker Ki-67, prognosticindicator p27^kip1and their clinicopathologic significance. Besides,discuss the effect of abnormal expression of FOXO3a in theoccurrence and the development of the glioma.2. To evaluate the specific effect of FOXO3a during gliomacell growth.Methods1. Immunohistochemical technique was used to detect theexpression levels of FOXO3a, p27^kip1and Ki-67in70cases ofglioma tissue. The glioma cases characterized in this study included:34grade Ⅱ（26fibrillary astrocytoma,1protoplasmic astrocytoma,2gemistocytic astrocytoma,3mixed oligo-astrocytoma and2ependymoma）;24grade Ⅲ （22anaplastic astrocytoma and2anaplastic oligodendroglioma）;12grade Ⅳ （8glioblastoma,3malignant ependymoma and1medulloblastoma）.2. Serum starvation and release were used to synchronizelymphoma cells. Flow cytometer was used to analyze cell cycle. The expression and subcellular localization of FOXO3a and p27^kip1weredetected by western blot and subcellular fractionation. LY294002was used to treat U87MG cell and T98G cell, the expression andsubcellular localization of FOXO3a and p27^kip1were investigated bywestern blot, immunoflurescence and subcellular fractionation.Results1. FOXO3a expression directly correlated with malignant gradeof glioma. FOXO3a positive expression was significantly associatedwith pathologic stage of the glioma. On other hand, no significantdifference were seen regarding patients’ age, gender, KPS score,tumor location, type of surgery, tumor diameter, vessel density, ornecrosis. In all cases of glioma analyzed, FOXO3a expression waspositively associated with p27^kip1（Spearman’s r=0.845, P<0.001）and was negatively associated with Ki-67expression （Spearman’s r=-0.555, P<0.001）.There was also a marked correlation betweenp27^kip1and Ki-67（Spearman’s r=-0.647, P<0.001）. By using theKaplan-Meier analysis, patients in the low expression FOXO3a（≤23.5%） group were significantly associated with poor overallsurvival （P<0.001）. Multivariate analysis using the Cox’sproportional hazards model showed that FOXO3a protein was anindependent prognostic indicator for patients’ overall survival （P<0.001）.2. Advances in the cell cycle study have revealed: FOXO3aincreased as the cells were treated with serum addition as well as thesame kinetics was observed at p27^kip1. Subcellular fractionationshowed that FOXO3a was mainly present in the cytoplasm of cellsreleased from Gkip11, leading to the dramatically reduced p27nuclearaccumulation. Administration of PI3K pharmacologic inhibitorLY294002abrogated this effect by regulating FOXO3a expressionand subcellular localization.Conclusions 1. The expression of FOXO3a was closely correlated with theaggressiveness of glioma. It indicates that disregulation of FOXO3amay cause the cell cycle abnormal and promote cell proliferation,thus may play key roles in the occurrence and development ofglioma.2. In vitro, FOXO3a modulated the cell cycle by transcriptionalregulation of p27^kip1. It suggests that FOXO3a has the potential to asa tumor suppressor gene. Gene therapeutic approaches aimed atPI3K or the pharmacologic inhibitors of PI3K to down-regulateP-FOXO3a expression could be developed for the management ofglioma.