WGA、RCA-Ⅰ和ECL凝集素受体在正常小鼠神经管和过量维生素A酸致畸神经管发育中的作用

发布时间：2018-06-26 18:09

本文选题：神经管 + 发育　；参考：《山东大学》2007年硕士论文

【摘要】： 目的：组织和器官的形态发生(morphogenesis)是通过细胞的形态变化和运动、细胞的识别和粘附、细胞的增殖和凋亡等过程而实现的。神经管的形成和发育也是一个极其复杂的生物学过程，包括细胞增殖、凋亡、分化、识别、迁移和功能的表达等等。近年来的研究表明：神经管的发育不仅受到细胞基因表达的控制，而且还与胚胎和神经上皮产生的多种调节因子有关，凝集素受体便是这些调节因子中的一员。凝集素受体是一类分布于细胞膜表面的糖蛋白、糖脂或糖复合物。凝集素受体上具有能被凝集素特异识别并结合的糖基。我们以昆明小鼠为研究对象，用亲和细胞化学的方法对正常鼠胚神经管和过量维生素A酸致畸鼠胚神经管发育中WGA、RCA-Ⅰ、ECL三种凝集素受体的分布状况以及变化规律进行了研究，以期阐明上述三种凝集素受体在正常神经管及过量维生素A酸致畸神经管发育中的作用。方法：将健康育龄雌性昆明小鼠与正常具有生育能力的雄鼠按2∶1合笼，次日晨查见阴栓者，当天中午定为孕0.5天。孕鼠受精后第7天下午6∶30一次性灌服花生油稀释的维生素A酸30mg／kg，以获取神经管畸形的小鼠胚胎。将正常组孕鼠和维生素A酸致畸组的孕鼠分别于孕第7.5、8.5、9.0、9.5、10.5、11.5，12.5天颈椎脱臼处死并立即剖腹取出子宫。用刀片切取有膨大结节部位的子宫(含胚胎)；孕11.5d、12.5d的孕鼠在滤纸上沿子宫侧轴将子宫剪开，仔细剥离柔软的蜕膜、分离鼠胚。所得标本用Bouin固定液固定，常规石蜡包埋，做厚度为5μm的连续切片。采用SABC亲和细胞化学法检测WGA、RCA-Ⅰ、ECL三种凝集素受体在小鼠胚胎神经管的分布情况。结果： 1．WGA染色结果：在孕7.5～12.5天，WGA受体在神经管的腔面、基底膜、神经上皮细胞的胞膜和胞浆均有表达。WGA受体表达的总体趋势是先下降后上升，其最低点出现于孕9.0天。在孕9.5～12.5天，WGA受体在头部和尾部的表达水平明显高于体部。在维生素A酸致畸组，WGA受体主要表达于神经管的腔面，其它部位的表达减弱或消失。但是，在孕10.5～12.5天，WGA受体在神经管的背部呈强阳性着色。无论神经管的头部、体部，还是尾部，WGA受体在致畸组表达水平明显低于正常组。 2．RCA-Ⅰ染色结果：在孕7.5～12.5天，RCA-Ⅰ受体在神经管的腔面、基底膜、神经上皮细胞的胞膜和胞浆均有表达。在神经管未关闭之前(孕7.5～9.0天)，RCA-Ⅰ受体的表达呈现逐渐上升的趋势。在神经管关闭之后(孕9.5～12.5天)，RCA-Ⅰ受体的表达在神经管头部和尾部仍呈现逐渐上升的趋势，在神经管体部，却呈现逐渐下降的趋势，而且，头部和尾部的表达强度均高于体部。在维生素A酸致畸组，RCA-Ⅰ受体主要表达于神经管的腔面，其它部位的表达减弱或消失。但在孕11.5～12.5天，神经管的背部却呈强阳性表达。无论神经管的头部、体部，还是尾部，RCA-Ⅰ受体在致畸组表达水平明显低于正常组。 3．ECL染色结果：在孕7.5～12.5天，强阳性着色部位仅见于神经管的腔面，基底膜、神经上皮细胞的胞膜呈阳性、弱阳性或阴性着色，神经上皮细胞的胞浆呈弱阳性或阴性着色。无论在神经管的头部、体部，，还是尾部，ECL受体的表达均呈现先下降后上升的趋势，其最低点在孕9.0天。在维生素A酸致畸组，阳性着色部位与正常发育神经管基本相同。但在孕11.5～12.5天，神经管的背部却呈强阳性表达。在孕9.5～12.5天神经管的头部，孕10.5～12.5天神经管的尾部，孕11.5～12.5天神经管的体部，ECL受体的表达水平低于正常发育的神经管相应部位。结论： 1．在神经管正常发育过程中，三种凝集素受体的数量和分布状况均存在时间和空间上的差异。 2．在神经管关闭前和关闭后，三种凝集素受体的表达趋势存在明显差异。这可能与神经管关闭的形态学事件有关。 3．在神经管正常发育过程中，三种凝集素受体的表达均呈现规律性的变化。它们可能参与对神经管发育的调节。 4．三种凝集素受体在神经管表达水平的下降，尤其是WGA、RCA-Ⅰ表达水平的下降与维生素A酸致神经管畸形相关。
[Abstract]:Objective: the morphogenesis of tissues and organs (morphogenesis) is realized through the process of cell morphological changes and movement, cell recognition and adhesion, cell proliferation and apoptosis. The formation and development of neural tubes are also an extremely complex biological process, including cell proliferation, apoptosis, differentiation, identification, migration, and function expression. In recent years, studies have shown that the development of the neural tube is not only controlled by the gene expression of the cells, but also related to a variety of regulatory factors produced by the embryo and the neuroepithelium. The agglutinin receptor is one of these regulatory factors.
Lectin receptor is a kind of glycoprotein, glycolipid or glycoconjugates on the surface of the cell membrane. The lectin receptor has a glycoprotein that can be specifically identified and combined by lectin. In Kunming mice, we used an affinity cytochemistry method for the development of WGA in the neural tube development of normal rat embryo neural tube and excessive vitamin A acid teratogenic mouse embryo. The distribution and variation of three kinds of lectin receptors of RCA- I and ECL were studied in order to elucidate the role of the three lectin receptors in the development of normal neural tube and excessive vitamin A acid teratogenic neural tube development.
Methods: the female Kunming mice of healthy childbearing age and the male mice with normal fertility were caged at 2: 1. The next day was found to be the 0.5 days of pregnancy. The pregnant rats were given the vitamin A acid 30mg / kg diluted by peanut oil at 6: 30 at seventh day after fertilization, in order to obtain the mouse embryos of the deformity of the God meridian. The pregnant rats in the vitamin A teratogenic group were sacrificed on the 7.5,8.5,9.0,9.5,10.5,11.5,12.5 day of the pregnancy and removed the uterus immediately. The uterus (containing the embryo) with the enlarged nodules was cut with the blade. The pregnant mice pregnant 11.5d and 12.5d cut the uterus along the lateral axis of the uterus on the filter paper and carefully stripped the soft decidua to separate the mouse embryos. The specimens were fixed with Bouin fixed solution, paraffin embedded and continuous slices with a thickness of 5 mu m. The distribution of WGA, RCA- I, ECL three agglutinin receptors in the neural tube of mouse embryos was detected by SABC affinity cytochemistry.
Result:
1.WGA staining results: at 7.5 to 12.5 days of pregnancy, the overall tendency of WGA receptor to express.WGA receptor expression in the cavity surface of the nerve canal, the basement membrane, the cytoplasm and the cytoplasm of the neuroepithelial cells was first descended and then increased, and the lowest point appeared at 9 days of pregnancy. The expression level of WGA receptor in the head and tail was significantly higher than that in the body at 9.5 to 12.5 days of pregnancy. In the vitamin A acid teratogenic group, the WGA receptor was mainly expressed in the cavity surface of the nerve canal, and the expression of other parts was weakened or disappeared. However, the WGA receptor was strongly stained on the back of the nerve tube at 10.5 to 12.5 days of pregnancy. No matter the head, body, or tail of the nerve canal, the expression level of WGA receptor in the teratogenic group was significantly lower than that of the normal group.
2.RCA- I staining results: at 7.5 to 12.5 days of pregnancy, the expression of RCA- I receptor in the cavity surface of the nerve canal, the basement membrane, the cell membrane and the cytoplasm of the neuroepithelial cells were expressed. The expression of RCA- I receptor showed a gradual rising trend before the nerve tube was closed (7.5 to 9 days of pregnancy). After the closure of the nerve canal (9.5 to 12.5 days), the table of RCA- I receptor At the head and tail of the nerve tube, the trend of the head and tail was gradually rising, but in the nerve tube body, the trend was gradually decreasing, and the expression intensity of the head and tail was higher than that in the body. In the vitamin A teratogenic group, the RCA- I receptor was mainly expressed in the cavity surface of the nerve canal, the expression of other parts was weakened or disappeared. But in the 11.5 to 12.5 days of pregnancy, The expression of RCA- I receptor in the teratogenic group was significantly lower than that in the normal group.
The results of 3.ECL staining: in the 7.5 to 12.5 day of pregnancy, the strong positive staining site was found only in the cavity surface of the nerve canal, the membrane of the basal membrane and the neuroepithelial cells was positive, weak positive or negative coloring, and the cytoplasm of the neuroepithelial cells was weak positive or negative coloring. The expression of the ECL receptor in the head, body and tail of the nerve tube appeared first. In the vitamin A acid teratogenic group, the positive staining site was basically the same as that of the normal developing nerve canal in the vitamin A teratogenic group. But the back of the nerve tube was strongly positive in the 11.5 to 12.5 day of pregnancy. The head of the nerve tube, the tail of the 10.5 to 12.5 day of the nerve canal, and the body of the nerve tube 11.5 to 12.5 days pregnant, and the body of the nerve tube in the 9.5 ~ 12.5 days of pregnancy. The expression level of ECL receptor was lower than that of normal neural tube.
Conclusion:
1. during the normal development of the neural tube, there are temporal and spatial differences in the number and distribution of the three lectin receptors.
2. there was a significant difference in the expression of three lectin receptors before and after the closure of neural tube. This may be related to the morphological events of neural tube closure.
3. during the normal development of neural tube, the expression of three lectin receptors showed regular changes. They may participate in the regulation of neural tube development.
4. the decrease of the expression of three lectin receptors in the neural tube, especially the decrease of WGA and RCA- I expression level, is related to the neural tube defects induced by vitamin A acid.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R363

【相似文献】