The chick retinal pigment epithelium grown on permeable support demonstrates functional polarity.
The chick retinal pigment epithelium grown on permeable support demonstrates functional polarity.
复制标题
在可渗透性支持物上生长的小鸡视网膜色素上皮表现出功能极性。
DOI：
10.1016/0014-4827(89)90092-x
复制
发表时间：
1989
影响因子：
3.7
通讯作者：
Koh,SW
中科院分区：
文献类型：
--
作者：
Koh,SW
来自鸡胚的视网膜色素上皮（RPE）在可渗透的支持物上培养。通过汇合培养物和对培养介质的分析，本研究表明在渗透膜上培养的 RPE 细胞保留了功能极性，这是 RPE 体内的一个特征。通过跟踪[3H]菊粉从培养物的顶端侧到基底侧的运动来估计汇合的RPE培养物中的细胞间通透性程度。将培养物的顶端侧暴露于[ 3 H]菊糖后24小时，顶端培养基中的 3 H浓度保持在基础培养基中的3.4至4.4倍。当 EDTA 存在时，RPE 的屏障功能消失。在汇合的 RPE 培养物中，经常观察到从培养物的顶端侧到培养物的基底侧的净单向流体运动。不同的培养物制备速率不同，最高为 1.60-1.84 μl/cm2/h。当培养物添加[35S]蛋氨酸26小时时，通过SDS-聚丙烯酰胺凝胶放射自显影评估，可以在培养基中检测到超过20条分子量范围为20,000至>250,000 Da的条带。虽然六种大分子似乎在基础培养基和顶端培养基中浓度相同，但大多数在基础培养基中浓度较高。对培养基中10％TCA可沉淀部分的分析表明，顶端培养基和基础培养基中的比活性分别为24.0±0.4×106和46.4±0.2×106(平均值±SEM，N=8)cpm/ml/mg RPE蛋白。当培养物与 VIP（血管活性肠肽）反应时，升高的细胞内环 AMP 被挤出到沐浴细胞的培养基中。然而，挤出到基础培养基中的速率是挤出到顶端培养基中的速率的两倍。汇合的 RPE 培养物的电子显微镜显示细胞的形态极化。细胞间空间似乎在细胞的顶端被由紧密连接、小带粘附连接和间隙连接组成的连接复合体封闭。
The retinal pigment epithelium (RPE) from the chick embryo was cultured on permeable support. Using confluent cultures and analysis of the incubation medium, the present study demonstrates that RPE cells cultured on permeable membrane retain functional polarity, a characteristic of the RPEin vivo. The degree of intercellular permeability in the confluent RPE cultures was estimated by following [3H]inulin movement from the apical side to the basal side of the cultures. Twenty-four hours after exposure of the apical side of the culture to [3H]inulin, the3H concentration in the apical medium remained at 3.4 to 4.4 times of that in the basal medium. The barrier function of RPE disappears in the presence of EDTA. Net unidirectional fluid movement from the apical side of the cultures to the basal side of the cultures is regularly observed in confluent RPE cultures. The rate varies among different preparations of cultures and the highest is 1.60–1.84 μl/cm2/h. When cultures are given 26 h of [35S]methionine, more than 20 bands with molecular weights ranging from 20,000 to >250,000 Da can be detected in the medium as assessed by autoradiography of SDS-polyacrylamide gels. While six macromolecules appear to be equally concentrated in the basal medium and the apical medium, the majority are in higher concentration in the basal medium. Analysis of the 10% TCA-precipitable fraction of the medium showed that the specific activities in the apical medium and basal medium were 24.0±0.4×106and 46.4±0.2×106(mean ± SEM,N= 8) cpm/ml/mg RPE protein, respectively. When cultures react with VIP (vasoactive intestinal peptide), the elevated intracellular cyclic AMP is extruded into the medium bathing the cells. However, the rate of extrusion into the basal medium is twice as fast as that into the apical medium. Electron microscopy of the confluent RPE cultures shows morphological polarization of the cells. The intercellular spaces appear to be closed at the apical side of the cells by junctional complexes consisting of tight junctions, zonular adherens junctions, and gap junctions.