PI3K/Akt/mTOR Tor Signaling Pathway Regulates Autophagy Induced By Hippocampus CA1 Area Of Rats With Vascular Dementia

Objectives On the basis of the interference of PI3K/Akt/m TOR pathway inhibitor LY294002 and rapamycin in vascular dementia model rats, to observe the pathway related protein expression of PI3 K, Akt and m TOR and the influence on expressions of autophagy related protein LC3 and Beclin1, to discuss the regulation of PI3K/Akt/m TOR signaling pathway in this process.Methods 1 192 SD rats were randomly divided into sham operation group, vascular dementia model group, LY294002 group and rapamycin group. Meanwhile, each group was further divided into 1, 2, 4 and 8 weeks, four subgroups after the success of model preparation(n=12). 2 Vascular dementia rat models were established by blocking four vessels. 3 Stereotaxis of lateral ventricle: Bregma points to the left or right next to open0.9 mm, back to 1.5 mm, drill hole with the micro dental drill and pull out the drill after significant breakthrough, reach the surface of the dura mater. Drawing 10μl, 10mmol/L LY294002 with microsyringe in the syringe, insert the needle along the borehole slowly reaching the depth of 3.8 mm, and inject uniformly in 5 minutes, retain the needle for 5minutes, and then pull the needle slowly. The step of rapamycin group is the same as the step of LY294002 group. 4 The expressions and location of PI3K、Akt and m TOR were detected by immunohistochemistry assay. 5 The ratio of autophagy-associated protein LC3II/LC3 I and Beclin1 were measured by Western blotting. 6 The expression and location of LC3 were detected in CA1 area of hippocampus by fluorescence microscope.7 We observed tissue slices with microscope camera OLYMPUS(400×), then analyzed them by Motic Med 6.0 digital medical image analysis system. The results of Western Blot were analyzed with Image J. 8 Application SPSS 17.0 statistical software for data analysis at last. Measurement data are presented as mean±SD( sx ±), using analysis of single variance, P<0.05 for the difference was statistically significant.Results 1 The immunohistochemical results of PI3K: The expressions of PI3 K were obviously increased at all-time points in the VD group compared with those in the shamoperation group(P < 0.05 or P < 0.01); compared with VD group, the expressions of PI3 K had no obvious change at all-time points in the LY294002 group and in the rapamycin group(P>0.05).2 The immunohistochemical results of Akt: The expressionsof Akt were obviously increased at all-time points in the VD group compared with these in the sham-operation group(P < 0.05 or P < 0.01); compared with VD group, the positive expression of Akt protein showed lower at every time point of the LY294002group(P < 0.05 or P < 0.01); the expression of Akt had no obvious change at alltime points in the rapamycin group(P> 0.05).3 The immunohistochemical results of m TOR: The expressions of m TOR were obviously increased at all-time points in the VD group compared with these in the sham-operation group( P < 0.05 or P < 0.01);Compared with VD group, the positive expression of m TOR protein showed lower at every time point in the LY294002 group and in the rapamycin group(P < 0.05 or P <0.01).4 Beclin1 result in western blot: The expression of Beclin1 was obviously increased at all-time points in the VD group compared with these in the sham-operation group(P < 0.05 or P < 0.01). Compared with VD group, Beclin1 was increased at all time points in the LY294002 group and in the rapamycin group(P < 0.05 or P < 0.01).5 LC3 result in western blot: The expression of LC3 was obviously increased at all-time points in the VD group compared with these in the sham-operation group(P < 0.05 or P< 0.01). Compared with VD group, LC3II/LC3 I ratio was increased at all time points in the LY294002 group and in the rapamycin group( P < 0.05 or P < 0.01). 6 LC3 labeling result in immunofluorescence: Fluorescence microscope results indicated that the sham-operation group of the forth week had the low expression of LC3; the expression of LC3 was obviously increased at 4 week in the VD group; LC3 was increased at 4week in the LY294002 group and in the rapamycin group.Conclusions 1 The over expressions of PI3K、Akt and m TOR protein in CA1 area of hippocampus of vascular dementia rats and PI3K/Akt/m TOR signaling pathway are involved in the pathogenesis of vascular dementia.2 PI3K/Akt/m TOR signaling pathway play inhibitory effect in vascular dementia.