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穹隆海马伞切割后大鼠海马内RUNX1T1 mRNA和蛋白的表达变化
邹琳清 金国华* 王海琴 李浩明 朱培培 秦建兵
解剖学报 ›› 2013, Vol. 44 ›› Issue (6) : 767-771.
穹隆海马伞切割后大鼠海马内RUNX1T1 mRNA和蛋白的表达变化
Changes of RUNX1T1 mRNA and protein expression in rat hippocampus after fimbria/fornix transection
目的 观察穹隆海马伞切割后不同时间点海马内RUNX1T1 mRNA和蛋白的表达变化及定位。方法 行穹隆海马伞切割术，制备模型大鼠。实验分为正常组、切割3d组、切割7d组、切割14d组、切割21d组、切割28d组。1.提取海马组织总mRNA，用Real-time PCR 检测大鼠海马组织中RUNX1T1mRNA 的表达；2.提取海马组织总蛋白，用Western blotting 检测海马内RUNX1T1蛋白的表达；3.行脑冷冻切片，行RUNX1T1免疫荧光检测和Hoechst标记，观察海马齿状回中RUNX1T1/Hoechst双标阳性细胞。 结果 Real-time PCR检测显示，切割3d组海马组织中RUNX1T1 mRNA的表达明显上调，其余各组差异不明显；Western blotting检测结果显示，正常组海马中有微量RUNX1T1蛋白表达，而切割3d组海马组织中RUNX1T1蛋白表达量明显上升，并达到高峰，7d时仍有较多的表达，此后，14d、21d、28d组中RUNX1T1蛋白表达很快又恢复到正常水平；免疫荧光检测结果表明，正常组海马齿状回中有少量的RUNX1T1/Hoechst阳性细胞，切割3d组海马齿状回中RUNX1T1/Hoechst阳性细胞数量最多，切割7d组逐渐减少，切割14d组、21d组和28d组与正常组差别不明显。结论 穹隆海马伞切割后海马内RUNX1T1 mRNA和蛋白表达在早期呈短暂性上调，并定位于海马齿状回颗粒下层，推测可能与海马神经再生过程中神经干细胞向神经元分化有关。
Objective To observe the temporal and spatial changes of RUNX1T1 mRNA and protein expression in rat hippocampus after fimbria/fornix transection at different time points. Methods Rat models were prepared by cutting fimbria fornix, and divided into normal group, cutting 3days, 7days, 14days, 21days and 28days groups. 1. The total hippocampal tissue mRNA was extracted, and the expression of RUNX1T1 mRNA was detected by real-time PCR; 2. The total protein of hippocampus was extracted, and the expression of RUNX1T1 protein was detected by western blotting; 3. Frozen brain sections were detected by RUNX1T1 immunofluorescence and Hoechst labeled, to observe the RUNX1T1/Hoechst double labeled positive cells in hippocampal dentate gyrus. Results Real-time PCR analysis showed that the RUNX1T1 mRNA expression in the hippocampus was significantly upregulated on the 3rd day after transection. Western blotting analysis indicated that the expression of RUNX1T1 protein was detected in the hippocampus of rats at basal condation, and the levels of RUNX1T1 protein were markedly increased at days 3 and 7, and peaked at day 3 after transection. In contrast, no siginificant change in the RUNX1T1 protein expression at days 14, 21 and 28. Immunostaining showed that the nember of RUNX1T1-positive cells was siginificantly higher at day 3 compared with control and decreased slowly after day 7. Conclusion The transient upregulation of RUNX1T1 mRNA and protein expression in the early phase after fimbria/fornix transection, and location in the hippocampal dentate gyrus granule lower, which may be related to the hippocampal neural regeneration.
RUNX1T1 / 海马 / 穹隆海马伞切割 / 神经再生 / 免疫荧光 / 大鼠
RUNX1T1 / Hippocampus / Fimbria/fornix transection / Neural regeneration / Immunofluorescence / Rat
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国家自然科学基金资助项目;省自然科学基金资助项目;自然科学基金资助项目
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