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Objective: To investigate the cutting fornix fornix hippocampal expression differences in normal hippocampal phosphatidylethanolamine binding protein (Phosphatidylethanolamine-binding protein, PEBP). Methods: (1) Real-time PCR: 42 SD rats were randomly divided into 7 groups (n = 6). A group of normal control group, and the remaining 6 were cutting bilateral after fornix hippocampus umbrella 1,3,7,14,21 and 28 d group. The take hippocampus of rats in each group, total RNA was extracted and synthesized the first strand cDNA, the method of application of Real-time PCR analysis of rat hippocampus PEBP mRNA expression changes after cutting the fornix FIMBRIA. CT values ??of detection cutting fornix FIMBRIA is in each group and the normal control group the PEBP and GAPDH gene expression, using the 2-ΔΔCT method to analyze the results obtained as the relative expression level of PEBP genes at different time points; (2) in situ hybridization : Take six SD rats, after cutting the right side of the dome FIMBRIA 7d, perfusion-fixed preparation hippocampus frozen sections, using digoxigenin-labeled the PEBP RNA probes in the hippocampus before, after each of 1 slices, in situ hybridization, PEBP mRNA expression changes observed both sides of the hippocampus. Each slice cut side and normal side of the pyramidal cell layer of the dentate gyrus granular layer and the gate area, the detection of particles lower 0.03mm2 area of ??PEBP mRNA-positive cells counts and average gray value; (3) Western Blot: 42 SD rats were randomly divided into seven groups of six each. A group of normal control group, and the remaining six groups divided into cutting bilateral fornix FIMBRIA 1,3,7,14,21 and 28 d group. Take the hippocampus of rats in each group, total protein was extracted and measured concentration, followed by SDS electrophoresis and transferred to a membrane for immunoblotting with rabbit anti-PEBP antibody detection. Respectively, at each time point of PEBP and a β-actin positive article with the average gray value detection, the reciprocal of the ratio between the two as the PEBP proteins at different time points relative expression levels; (4) immunohistochemistry: 36 SD rats were randomly divided into 6 groups of 6. Divided into cutting the right fornix hippocampus umbrella 1,3,7,14,21 and 28d group. Rats attentive after perfusion fixation, the brain was removed, frozen sections, in the hippocampus before, and after each of immunohistochemical staining of a slicing line PEBP cutting side and normal side were counted hippocampal pyramidal cell layer dentate gyrus granule cell layer and the gate area, the number of the PEBP positive cells 0.06mm2 area of ??particles lower, and to detect the gray value; (5) at each time point data were analyzed by ANOVA using SPSS11.5 statistical software and The groups were compared by paired t test slice data in situ hybridization and immunohistochemistry. Results: (1) Real-time PCR: hippocampus PEBP mRNA relative expression in the normal group, 1.16 ± 0.10, began to increase after cutting 3d (1.51 ± 0.08), 7d (1.64 ± 0.07) reached the highest level, about the subsequent decline, 21d (1.14 ± 0.13) returned to normal levels. PEBP mRNA relative expression level of analysis of variance and pairwise comparison results show that, 3d, 7d significant difference (P lt; 0.05) with the rest between each cutting group, 3d, 7d groups and other groups between no significant difference (P gt; 0.05); (2) in situ hybridization: cutting side and normal side of the hippocampal CA1 ~~ CA3 pyramidal cell layer and dentate gyrus granule layer cells were PEBP mRNA expression, PEBP mRNA positive cells no significant difference, but the average gray value of the cutting side PEBP mRNA positive cells 78.47 ± 4.45, 102.58 ± 15.99,, the normal side-cut side of the hybridization signal was stronger than the normal side (P lt; 0.01). Observed more in the cut side of the dentate gyrus door area and SUBGRANULAR of of PEBP mRNA-positive cells (60.67 ± 6.71) and the average gray value of 67.16 ± 9.28; fewer positive cells and the normal side (50.33 ± 8.52), average gray value of 99.62 ± 13.44, on both sides of the number of positive cells and the average gray value is a significant difference (P lt; 0.05); (3) Western Blot: PEBP protein relative expression levels of the normal control group is 0.0551 ± 0.0093 began to increase after cutting 3d (0.077 ± 0.0083), 7d (0.0965 ± 0.0089) reached the highest level, and then declined, 28d (0.0505 ± ± 0.0066) about returned to normal levels. Groups PEBP relative expression amount of variance analysis and twenty-two comparison results show that, each cutting group and the normal group comparison, 1d, 28d group and the normal group, no significant difference (P gt; 0.05), the rest of each cutting group and the normal group between exist in a significant difference (P lt; 0.05); between each cut group comparison, after cutting 7d group and other groups are exist significant difference (P lt; 0.05), 3d, 14d, 21d and 1d, 28d between the existence of significant forward sex differences (P lt; 0.05), while the 3d, 14d, 21d between the non-significant with sex differences (P gt; 0.05), 1d, 28d rooms are also non-significant with sex differences (P gt; 0.05); (4) immune group of: the PEBP immunohistochemistry results showed that, PEBP expression the hippocampal CA1 ~ CA3 pyramidal cell layer of the dentate gyrus granule layer door area, SUBGRANULAR, cytoplasmic hippocampal pyramidal cell layer, cut side of each time point and teeth like gyrus granule layer PEBP number of positive cells with the normal side was no significant sex difference (P gt; 0.05), but the cutting side the PEBP-positive cells stained deepen the most obvious 7d average gray value of 68.41 ± 10.53, while the normal side of the average gray value of 91.52 ± 7.77, both sides have more significant difference (P lt; 0.01). Cutting side of the hippocampal dentate gyrus door and SUBGRANULAR visible deepening more stain the PEBP-positive cells, 7d, most significantly, the number of positive cells was 53.33 ± 4.27, and the average gray value of 65.34 ± 15.84, the number of positive cells of the normal side 37.50 ± 4.04, compared to the average gray value of 97.16 ± 11.62, cutting side of the cell number and the average gray value with normal side were significant differences (P lt; 0.05). Pyramidal cell layer of the dentate gyrus granule layer and door area after 14d SUBGRANULAR PEBP expression began to decline the 28d near normal side. Conclusion: (1) cutting fornix fornix hippocampus after organization of PEBP mRNA and protein expression levels were significantly up-regulated the expression of the two were a low to high to low expression, and the lifting of the two trends basic consistent, and 7 days after cutting peaked; (2) PEBP mRNA and protein expression in hippocampal CA1 ~ CA3 pyramidal cell layer of the dentate gyrus granule cell layer of the cells of the lower door area and particle; (3) cutting hippocampal CA1 ~ CA3 pyramidal cell layer of the dentate gyrus granule cell layer with the normal side just a stain deepened; door zone of the dentate gyrus and particles lower compared with the normal side, not only staining deepen, and an increase in the number of positive cells. Tip cutting fornix fornix hippocampus after increased in of PEBP mRNA and protein expression may be related to the hippocampus in the neural stem cells from the body or implanted neurons or cholinergic differentiation of neurons.
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