Mechanism of stilbene glycosides on apoptosis of SH-SY5Y cells via regulating PI3K/AKT signaling pathway

Objective:To investigate the effects of stilbene glycoside(TSG)on okadaic acid-induced apoptosis in human neuroblastoma cells(SH-SY5Y)via the PI3K/AKT pathway.Methods:The optimal concentration of OA was screened by CCK-8 assay,and SH-SY5Y cells were divided into control group,model group,TSG group,LY294002 group and LY294002+TSG group.The proliferation and apoptosis in each group were detected by CCK-8 and TUNEL assays;Western blotting method and real-time fluorescence quantitative polymerase chain reaction was used to detect the expression of PI3K,P-PI3K(Y607),AKT,P-AKT(Ser473),Bcl-2 and Bax proteins.The relative protein expression was represented by P-PI3K(Y607)/PI3K,P-AKT(Ser473)/AKT and Bcl-2/Bax gray ratio.Results:CCK-8 screened the optimal concentration of OA as 40 nmol/L.Compared with the control group,the model group increased relative cell viability,decreased apoptosis rate,the pathway and apoptotic proteins expression levels of P-PI3K(Y607)/PI3K,P-AKT(Ser473)/AKT and Bcl-2/Bax were decreased,and the mRNA expression levels of PI3K,AKT and Bcl-2 were decreased.Bax mRNA expression level increased(P<0.05);Compared with model group,TSG group increased relative cell viability,decreased apoptosis rate,increased protein expression levels of P-PI3K(Y607)/PI3K,P-AKT(Ser473)/AKT,Bcl-2/Bax,and increased mRNA expression levels of PI3K,AKT,and Bcl-2.Bax mRNA expression decreased(P<0.05),LY294002 group decreased relative cell viability,increased apoptosis rate,P-PI3K(Y607)/PI3K protein expression levels were significantly decreased(P<0.05),P-AKT(Ser473)/AKT and Bcl-2/Bax protein expression levels were significantly decreased,but there was no statistical significance,PI3K,AKT and Bcl-2 mRNA expression levels were decreased,and Bax mRNA expression levels were increased(all P<0.05);Compared with LY294002 group,LY294002+TSG group increased relative cell viability,decreased apoptosis rate,and the protein expression levels of P-PI3K(Y607)/PI3K,P-AKT(Ser473)/AKT and Bcl-2/Bax were increased.The mRNA expression levels of PI3K,AKT,Bcl-2 were increased,Bax was decreased(all P<0.05).Conclusion:Stilbene glycoside may alleviate okadaic acid-induced apoptosis in SH-SY5Y cells by interfering with the PI3K/AKT signaling pathway,which in turn regulates the expression of apoptotic factors such as Bcl-2 and Bax.

A gene encoding AtPIP5K2 may be involved in regulating the sensitivity to osmotic stress

One mutant line eto with salt tolerance was screened from a T-DNA insertion mutant collection of Arabidopsis thaliana. In addition to a reduced rate of seed germination, NaCl and ABA also inhibited the growth and the greening of cotyledons of wild-type seedlings, but not the eto mutant. TAIL-PCR analysis showed that T-DNA tag insertion in the eto was located at nucleotide 27,502 in BAC F3M18, upstream （at position -487 relative to the translation initiation codon） of gene At lg77740 （encoding a putative phosphatidylinositol-4-phosphate 5-kinase, AtPIP5K2）. This inserted mutation cosegregated closely with the eto phenotype, Another analysis not only indicated that AtPIP5K2 transcript is expressed predominantly in roots and rosette leaves, but also showed the T-DNA insertion resulted higher accumulation of the AtPIP5K2 in eto mutant plants and did not influenced the expression of the upstream At lg77730 gene. This change may play an essential role in the tolerance of eto mutant plant to the osmotic stress.

Functional Cooperativity of Enzymes of Phosphoinositide Conversion According to Synergistic Effects on Pectin Secretion in Tobacco Pollen Tubes

The Arabidopsis phosphoinositide kinases PI4Kβ1 and PIP5K5 have been implicated in the control of directional vesicle trafficking underlying polar tip growth in pollen tubes. PI4Kβ1 and PIP5K5 catalyze key consecutive steps of phosphoinositide conversion, and it appears obvious that phosphatidylinositol-4-phosphate formed by PI4Kβ1 might act as a substrate for phosphatidylinositol-4,5-bisphosphate formation by PIP5K5. However, this hypothesis has not been experimentally addressed and distinct localization patterns of PI4Kβ1, PIP5K5, and also PI-synthases （PIS） generating phosphatidylinositol suggest additional complexity. Here, the synergistic functionality of enzymes of phosphoinositide conversion was assessed. In tobacco and Arabidopsis pollen tubes, phosphoinositides influence the apical secretion of pectin, and increased pectin deposition results in characteristic morphological alterations. Catalytically active and dominant negative variants of PI4Kβ1 and PIP5K5 were systematically co-expressed in tobacco pollen tubes and the incidence of morphologies related to enhanced pectin secretion was evaluated. The data support a proposed functional interplay of PI4Kβ1 and PIP5K5 at the trans-Golgi network, mediating directional vesicle trafficking. Co-expression experiments additionally including PIS isoforms, PIS1 or PIS2, indicate that pectin secretion is synergistically mediated by PI4Kβ1 and PIPSK5 acting on Ptdlns formed by PIS2 rather than PIS1. Possible ramifications for the preferential channeling of phosphoinositide intermediates between particular isoforms of PI pathway enzymes are discussed.