| Objective:to observe the effects of combining the injection of H102with transplanting HUCMSC on learning and memory function, and then detect the expression of P-tau, GSK-3β, PP-2A and PP-1in the brain of APP transgenic mice.Methods:1.Collect the umbilical cord of the newborn by cesarean section under sterile conditions, separate and culture the cells to observe the shape and the surface marker by FCM and then mark them with Dil.2. The APP transgenic mice were randomly divided into model group, the H102treatment group, the HUCMSC2-weeks group, the HUCMSC4-weeks group, the H102+HUCMSC group, and a group of C57BL/6J mice with the same age and background was set as normal. Model group and the normal control group were treated with nonnal saline solution, while the H102treatment group and H102+HUCMSC group were treated with compounds (H102in normal saline solution), the HUCMSC2-weeks and the HUCMSC4-weeks group were transplanted with HUCMSC. The ability of spatial reference memory was determined by Morris Water Maze.3. Immunohistochemical stain and western blot were used to detect the content of P-tau, GSK-3p, PP-2A and PP-1in the brain of the APP transgenic mice.Results:1. UC is a rich source of HUCMSC, and HUCMSC expressed strongly CD13, CD29, CD106and CD105, so they have the features of the MSC. After marked, it can be saw red fluorescence on the most HUCMSC. And after transplanted, the HUCMSC with red fluorescence can also be seen in the slice.2. The Morris water maze test:(1) Place Navigation Training:The escape latency of mice were much longer in the model group than in the nomal group (P<0.01). There was no difference between the HUCMSC2-weeks group and the model group (P>0.05), but the escape latency of mice was shorter in HUCMSC4-weeks group, H102treatment group and the H102+HUCMSC group(P<0.01). The escape latency of the mice in the H102treatment group was similar with the H102+HUCMSC group, but which in the HUCMSC group was much longer(P<0.05, P<0.01). (2) Spatial Probe Test:Compare with normal group, the times of the mice of the model group past the position of the platform were significantly less (P<0.01), and the initial angles were significantly larger (P<0.01). There was no difference between the HUCMSC2-weeks group and the model group (P>0.05). In addition, compared with the model group, the times of the mice of the HUCMSC4-weeks group, H102treatment group and H102+HUCMSC group past the position of the platform were significantly more (P<0.05), and the initial angles were much smaller (P<0.01).3. The test of immunohistochemistry and western blot(1) The test of immunohistochemistry:There were much more cells which expressed P-tau and GSK-3β in the brain of the mice of the model group than in the nomal group (P<0.01), but compared with the model group, there were little in the HUCMSC4-weeks group, H102treatment group and the H102+HUCMSC group (P<0.01). At the same time, we can see lots of cells which expressed PP-2A and PP-1in the brain of the mice in the HUCMSC4-weeks group, H102treatment group and the H102+HUCMSC group, and there were less in the model group(P<0.01).(2) The test of western blot:The content of the P-tau in the brain of the model group mice is more than that in the nomal group (P<0.01), but the PP-2A is muce less (P<0.01). The level of the protein expression of P-tau in the brain of the mice of the HUCMSC4-weeks group, H102treatment group and the H102+HUCMSC group is lower than that of the model group (P<0.01), but the level of the PP-2A is higher (P<0.01).Conclusions:Through reversing the production of A(3to reduce the content of the P-tau and GSK-3β and improve the expression the PP-2A and PP-1, injecting H102together with transplanting HUCMSC not only can reduce the damage of the neurons but also can promote the reconstruction of the nervous system, and then advance the ability of learning and memory of the APP transgenic mice. |
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