Isolation and purification of a cell adhesion factor from crayfish blood cells.
Isolation and purification of a cell adhesion factor from crayfish blood cells.
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DOI：
10.1083/jcb.106.5.1795
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发表时间：
1988-05
期刊：
影响因子：
--
通讯作者：
Söderhäll K
中科院分区：
文献类型：
--
作者：
Johansson MW;Söderhäll K
从淡水螯虾中分离的颗粒血细胞（血细胞）在体外附着并铺展在涂有淡水螯虾血细胞裂解物的盖玻片上。螯虾血浆中未检测到细胞粘附活性。细胞粘附活性仅存在于血细胞裂解物中，其中酚氧化酶原（proPO）激活系统（Soderhall和Smith，1986 a，B）已被脂多糖（LPS）、β-1，3-葡聚糖海带多糖或通过在5 mM Ca 2+中制备裂解物激活。颗粒状或半颗粒状血细胞的裂解物均可介导粘附。在A23187诱导的颗粒细胞的胞吐作用后，如果将其与海带多糖孵育，则在分泌的物质中可以产生细胞粘附活性。从活性血细胞裂解物中分离出负责细胞粘附的因子，并通过硫酸铵沉淀、阳离子交换色谱和Con A-Sepharose纯化; SDS-聚丙烯酰胺凝胶上的分子量约为76 kD。该76 kD带的抗体抑制细胞粘附。Ca ~（2+）是细胞粘附于粘附因子所必需的。用氰化物或叠氮化物，细胞附着但不能扩散。有人建议，在体内的细胞粘附因子是存储在分泌颗粒的半颗粒和颗粒细胞在一个假定的非活性前的形式，它可以在胞吐过程中释放，并在β-1，3-葡聚糖或LPS的存在下，被激活的细胞外介导的细胞附着和传播，过程中至关重要的节肢动物宿主防御。
Isolated granular haemocytes (blood cells) from the crayfish Pacifastacus leniusculus attached and spread in vitro on coverslips coated with a lysate of crayfish haemocytes. No cell adhesion activity was detected in crayfish plasma. The cell adhesion activity was only present in haemocyte lysates in which the prophenoloxidase (proPO) activating system (Soderhall and Smith, 1986a, b) had been activated; either by lipopolysaccharide (LPS), the beta-1,3-glucan laminarin, or by preparing the lysate in 5 mM Ca2+. Both lysates of granular or of semigranular haemocytes could mediate adhesion. After A23187-induced exocytosis of the granular cells, cell adhesion activity could be generated in the secreted material if it was incubated with laminarin. The factor responsible for cell adhesion was isolated from an active haemocyte lysate and purified by ammonium sulfate precipitation, cation exchange chromatography and Con A-Sepharose; it had a molecular mass of approximately 76 kD on an SDS-polyacrylamide gel. An antibody to this 76-kD band inhibited cell adhesion. Ca2+ was necessary in the medium for the cells to adhere to the adhesion factor. With cyanide or azide, the cells attached but failed to spread. It is suggested that in vivo the cell adhesion factor is stored in the secretory granules of the semigranular and the granular cells in a putative inactive pro-form, which can be released during exocytosis and, in the presence of beta- 1,3-glucans or LPS, be activated outside the cells to mediate cell attachment and spreading, processes of essential importance in arthropod host defense.