IN-VITRO ESTABLISHMENT OF CONTINUOUS CELL-CULTURES AND CELL-LINES FROM 10 COLONIAL CNIDARIANS
IN-VITRO ESTABLISHMENT OF CONTINUOUS CELL-CULTURES AND CELL-LINES FROM 10 COLONIAL CNIDARIANS
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DOI：
10.1007/bf00680224
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发表时间：
1994-10-01
期刊：
影响因子：
2.4
通讯作者：
RINKEVICH, B
中科院分区：
文献类型：
--
作者：
FRANK, U;RABINOWITZ, C;RINKEVICH, B
建立了静居型海洋刺胞动物Stylophora pistillata、Porites lutea、Favia favus （anthzoa、Hexacorallia、Madeporaria）、Parerythropodium fulvum fulvum、dendronephthia hemprichi、Nephthya sp. Heteroxenia fuscescence （anthzoa、Octocorallia、Alcyonacea）、Clatharia rubrinoides、Plexaura A （anthzoa、Octocorallia、Gorgonacea）和Millepora dichotoma (Hydrozoa) 10个分类群的连续细胞培养方法。从菌落碎片和/或浮藻幼虫（1993年从红海埃拉特和巴拿马圣比亚斯群岛的珊瑚礁收集）中获得各种细胞类型（5至20 mu m）的原代培养物，使用三种分离方法：机械方法，化学方法和一种新颖的方法，自发分离。细胞在改良的Leibowitz L15培养基中培养，其中含有5%至10%的热灭活胎牛血清，在海水中稀释。分离后7 ~ 20 d，原代培养细胞增殖。当发展到二次培养时，纺锤形细胞（5 - 10 μ m）逐渐取代了雌蕊花、红萼花、黄萼花和二歧花的原代培养中的所有其他细胞类型。当稀释的培养基被浓缩的培养基取代时，或当微生物污染发生时，纺锤形细胞分化成几种细胞类型。克隆了3个种（S. pistillata， C. rubrinoides和M. dichotoma）的梭形细胞的二次培养。它们产生了持续增殖的细胞系（分别为NIO-SPP-I、NIO-CR-I和NIO-MD-1）。planula的无性系起源于一个圆形细胞，开始分化并产生异质培养。以上四种培养物的样本被冷冻以备将来的工作。fuscescence、d.p hemprichi、Nephthya sp.、p.f. fulvum和F favus的次生细胞培养中，圆形细胞大小不等（5 ~ 30 μ m）。从黄叶假单胞菌培养的几个细胞发育成扁平的细胞，这些细胞通过迷路菌门真核单细胞生物特有的外质网络相互连接。我们建议这种培养方法可以作为一种普遍的方案，从其他刺胞动物生产组织培养。所建立的细胞系可用于刺胞生物和生态学的各种学科。
Continuous cell cultures from the ten taxa of sedentary colonial marine cnidarians Stylophora pistillata, Porites lutea, Favia favus (Anthozoa, Hexacorallia, Madeporaria), Parerythropodium fulvum fulvum, Dendronephthya hemprichi, Nephthya sp. Heteroxenia fuscescence (Anthozoa, Octocorallia, Alcyonacea), Clatharia rubrinoides, Plexaura A (Anthozoa, Octocorallia, Gorgonacea) and Millepora dichotoma (Hydrozoa) were established in vitro. Primary cultures of various cell types (5 to 20 mu m) were obtained from colony fragments and/or planula larvae (collected in 1993 from coral reefs at Eilat, Red Sea and from the San Bias islands, Panama) using three dissociation approaches: a mechanical approach, chemical approach and a novel approach, spontaneous dissociation. Cells were cultured in a modified Leibowitz L15 medium, with 5 to 10% heat-inactivated fetal bovine serum, diluted in seawater. Cell proliferation was observed in primary cultures within 7 to 20 d following dissociation. When developed to secondary cultures, spindle-shaped cells (5 to 10 mu m) gradually replaced all other cell types seen in primary cultures of S. pistillata, C. rubrinoides, P. f. fulvum and M. dichotoma. The spindle-shaped cells differentiated to several cell types when the diluted medium was replaced with a concentrated one, or when microbial contamination occurred. Secondary cultures of spindle-shaped cells from three species (S. pistillata, C. rubrinoides and M. dichotoma) were cloned. They gave rise to continuously proliferating cell lines (NIO-SPP-I, NIO-CR-I and NIO-MD-1, respectively). A clone from Plexaura planula, originated from one rounded cell, started to differentiate and gave rise to a heterogeneous culture. Samples of the above four cultures were frozen for future work. Rounded cells of various sizes (5 to 30 mu m) dominated secondary cell cultures of H. fuscescence, D. hemprichi, Nephthya sp., P. f. fulvum and F favus. Several cell cultures from P. lutea developed flattened cell interconnected by ectoplasmic networks characteristic of the eukaryotic unicellular organisms of the phylum Labyrinthulomycota. We propose that this culture methodology may be used as a ubiquitous protocol for producing tissue cultures from other cnidarians. The established cell lines may be used in a variety of disciplines in cnidarian biology and ecology.