Objective: To explore the role of calcium/calmodulin-dependent protein kinase Ⅱ(CaMK-Ⅱ) and protein phosphatase-2 A(PP2 A) on hyperphosphorylation of tau induced by AlCl_3 in SH-SY5 Y cell.
Methods: SH-SY5 Y cells were assigned to the control group, AlCl_3 low, middle, high exposed groups(200, 400800 μmol/L Al~(3+)), KN93 intervention group(800 μmol/L Al~(3+)+0. 5 μmol/L KN93) and sphingosine intervention group(800 μmol/L Al~(3+)+5 nmol/L sphingosine). After 48 h of exposure and intervention, the viabilities of cells were measured by CCK-8 assay, the contents of CaMK-Ⅱ and PP2 A were determined by ELISA, and the expression of tau5 and phosphorylation of Thr-181, Thr-231, Ser-262 and Ser-396 were detected by Western-Blot.
Results: The viabilities of cells in AlCl_3 middle and high exposed groups were significantly lower than that of the control group(P<0. 05). Compared with the AlCl_3 high exposed group, the viabilities of cells in KN93 intervention group and sphingosine intervention group were significantly increased(P<0. 05), as well as the difference of the cell viability between sphingosine intervention group and the control group was not statistically significant. Compared with the control group, the contents of CaMK-Ⅱin AlCl_3 low, middle, high exposed groups were significantly increased(P<0. 05), while the contents of PP2 A in those groups were significantly decreased(P<0. 05). Compared with AlCl_3 high exposed group, the contents of CaMK-Ⅱ in KN93 intervention group and sphingosine intervention group were significantly decreased(P<0. 05), and PP2 A in those groups were significantly increased(P<0. 05). The expression of tau5 and phosphorylation of Thr-181, Thr-231, Ser-396 in AlCl_3 low, middle, high exposed group showed significantly higher than those of the control group(P<0. 05), while the phosphorylation of Ser-262 in AlCl_3 high exposed group showed significantly higher than that of the control group(P<0. 05). After intervention with KN93 and sphingosine, the expression of tau5, and phosphorylation of Thr-181, Thr-231, Ser-262, Ser-396 in KN93 intervention group and sphingosine intervention group were significantly lower than those of AlCl_3 high exposed group( P<0. 05), as well as compared with the control group, the phosphorylation of Ser-262 in KN93 intervention group and Thr-181, Thr-231 and Ser-396 in sphingosine intervention group were not statistical difference.
Conclusion: AlCl_3 could increase the phosphorylation of Thr-181, Thr-231, Ser-262 and Ser396 in SH-SY5 Y cells, which mechanism may relate to the changes of CaMK-Ⅱand PP2 A. CaMK-Ⅱ mainly regulates the phosphorylation of Ser-262 induced by AlCl_3, while PP2 A mainly regulates the phosphorylation of Thr-181, Thr-231 and Ser-396 induced by AlCl_3. It is suggested that hyperphosphorylated tau protein induced by AlCl_3 is closely related to PP2 A.
Keywords: aluminum; calcium/calmodulin-dependent protein kinase Ⅱ; hyperphosphorylated tau; protein phosphatase-2A.