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神经损伤与再生
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神经损伤与再生  

Wnt信号通路参与脑缺血再灌注损伤后的细胞凋亡一文的图片
(Volume 8 Number 1 January 2013)


Cite this article:
Bin Liu, Jing Tang, Shiying Li, Yuqin Zhang, Yan Li, Xiaoliu Dong. Involvement of the Wnt signaling pathway and cell apoptosis in the rat hippocampus following cerebral ischemia/reperfusion injury[J]. Neural Regeneration Research, 2013, 8(1): 70-75.

 


Quantitative analysis of experimental animals

A total of 66 rats were randomly assigned to three groups: sham surgery group (no middle cerebral artery occlusion, n = 6), model group (middle cerebral artery occlusion,    n = 30) and DKK1 group (middle cerebral artery occlusion + DKK1 administration, n = 30). Six rats from each of the model and DKK1 groups were selected at 3, 6, 24, 48 and 72 hours following reperfusion for experimental analysis. Sham rats were obtained at    72 hours following reperfusion. A total of 66 rats were included in the final analysis.

 

DKK1 reduced cell apoptosis in the rat hippocampal CA1 region following cerebral ischemia/reperfusion

TUNEL staining in the sham surgery group was used to  detect minimal apoptotic cells in the hippocampal CA1 region (not examined statistically). However, in the model and DKK1 groups, the number of apoptotic cells in the CA1 region increased at 3 hours and peaked at 48 hours, following ischemia/reperfusion. Compared with the model group, the number of apoptotic cells in the DKK1 group significantly decreased at various time points (P < 0.05 or P < 0.01; Table 1, Figure 1).

Table 1 Dickkopf-1 effects on cell apoptosis (number in × 200 visual field) in the rat hippocampal CA1 region following cerebral ischemia/reperfusion injury 

Figure 1 Cell apoptosis in the rat hippocampal CA1 region (TUNEL staining, × 200).

Apoptotic cells with brown or yellow nuclei are visible in the model and Dickkopf-1 groups at various time points (arrows). Fewer apoptotic cells with weak staining are observed in the Dickkopf-1 group, compared with the model group, at corresponding time points.

DKK1 inhibited β-catenin and GSK-3β protein expression in the rat hippocampal CA1 region following cerebral ischemia/reperfusion Immunohistochemistry showed minimal β-catenin and GSK-3β expression in the sham surgery group (not examined statistically). In the model and DKK1 groups, β-catenin and GSK-3β expression was visible at various time points, and peaked at 48 hours following reperfusion (Figure 2).

Figure 2 β-catenin and glycogen synthase kinase-3β (GSK-3β) protein expression in the rat hippocampal CA1 region (immunohistochemical staining, × 200). Brown or yellow β-catenin and GSK-3β-positive cells are visible at each time point in the model and Dickkopf-1 groups (arrows). β-catenin and GSK-3β-positive cells were fewer in the Dickkopf-1 group than in the model group, at corresponding time points.

Compared with the model group, the number of GSK-3β-positive cells was significantly reduced, at corresponding time points, in the DKK1 group (P < 0.05 or P < 0.01; Table 2).

Table 2 Dickkopf-1 effects on glycogen synthase kinase-3β protein expression (number in × 200 visual field) in the rat hippocampal CA1 region following cerebral ischemia/reperfusion injury

Compared with the model group, the number of GSK-3β-positive cells was significantly reduced, at corresponding time points, in the DKK1 group (P < 0.05 or P < 0.01; Table 2). The number of β-catenin-positive cells was also decreased in the DKK1 group, compared with the model group, but there was no significant difference (P > 0.05; Table 3).

Table 3 Dickkopf-1 effects on β-catenin protein expression (number in × 200 visual field) in the rat hippocampal CA1 region following cerebral ischemia/ reperfusion injury

Western blot analysis confirmed our immunohistochemistry results, with decreased β-catenin (P > 0.05; Figure 3, Table 4) and significantly decreased GSK-3β (P < 0.05 or P < 0.01; Figure 3, Table 5) protein expression, at each time point, in the DKK1 group, compared with the model group.

Figure 3 β-catenin (A; 61 kDa) and glycogen synthase kinase-3β (B; 63 kDa) protein expression in the rat hippocampal CA1 region as determined by western blot.

β-actin (42 kDa) was used as an internal reference.

Table 4 Dickkopf-1 effects on β-catenin expression (absorbance ratio to β-actin) in the rat hippocampal CA1 region following cerebral ischemia/reperfusion injury 

Table 5 Dickkopf-1 effects on glycogen synthase kinase-3β protein expression (absorbance ratio to β-actin) in the rat hippocampal CA1 region following cerebral ischemia/reperfusion injury
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