Srihawong Thanida, Kuwana Takashi, Siripattarapravat Kannika, Tirawattanawanich Chanin
Center for Agricultural Biotechnology, Kasetsart University, NakhonPathom, Thailand.
Int J Dev Biol. 2015;59(10-12):453-60. doi: 10.1387/ijdb.140287ct.
Avian primordial germ cells (PGCs) are destined to migrate a long distance from their extra embryonic region via the vascular system to the gonadal ridges where they form the germ cells. Although PGC migration is crucial for a genetic continuation to the next generation, the factors and mechanisms that control their migration remain largely unknown. In the present study the chemotactic effect of stem cell factor (SCF) was examined on chicken blood circulating PGCs (cPGC), employing 3D chemotaxis slides and time-lapsed imaging analyses as an in vitro study model. Upon in vitro exposure to an SCF gradient, 77.1% (54 out of 70) of cPGCs showed a clear response, of which 48.1% (26 out of 54) polarized with the consecutive formation of a persistent membrane protrusion and significant directional migration towards the gradient and the others showed transient membrane protrusions. In contrast, the controls and apparently SCF unresponsive cPGCs and c-kit-negative red blood cells (RBCs) showed only cytoplasmic cycling with random formations of membrane blebbing and no directional migration. Significant (p < 0.05) differences between the SCF-treated and control cPGCs and RBCs were found in the migration parameters of eccentricity, accumulated and Euclidean distances, and migration velocity. The SCF-treated PGCs also revealed a chemotactic response, as judged by their significant displacement of center of mass and Rayleigh test. Complete inhibition of all the SCF-induced responses in PGCs was found following pretreatment of the cPGCs with 10 µM of the c-kit inhibitor, STI57l, prior to SCF exposure. In addition, cPGCs were found to be positive for c-kit expression using a polyclonal goat anti-mouse c-kit primary antibody, suggesting that the cPGCs were capable of SCF sensing and the potential involvement of SCF/c-kit in the chemotactic migration. Therefore, SCF is suggested to function as a chemoattractant in the migration of chicken cPGC.
禽类原始生殖细胞（PGCs）注定要从其胚外区域经血管系统长途迁移至性腺嵴，在那里它们形成生殖细胞。尽管PGC迁移对于遗传延续至下一代至关重要，但控制其迁移的因素和机制仍 largely未知。在本研究中，采用 3D 趋化性载玻片和延时成像分析作为体外研究模型，检测了干细胞因子（SCF）对鸡血液循环中的 PGCs（cPGCs）的趋化作用。在体外暴露于 SCF 梯度后，77.1%（70 个中的 54 个）的 cPGCs 表现出明显反应，其中 48.1%（54 个中的 26 个）发生极化，随后持续形成持久的膜突起并朝着梯度方向进行显著的定向迁移，其他的则表现出短暂的膜突起。相比之下，对照组以及明显对 SCF 无反应的 cPGCs 和 c-kit 阴性红细胞（RBCs）仅表现出细胞质循环，随机形成膜泡，无定向迁移。在偏心率、累积距离和欧几里得距离以及迁移速度等迁移参数方面，发现 SCF 处理的 cPGCs 与对照组的 cPGCs 和 RBCs 之间存在显著（p < 0.05）差异。经 SCF 处理的 PGCs 还显示出趋化反应，这通过其质心的显著位移和瑞利检验得以判断。在用 10 µM 的 c-kit 抑制剂 STI57l 对 cPGCs 进行预处理后，再暴露于 SCF，发现 PGCs 中所有 SCF 诱导的反应均被完全抑制。此外，使用多克隆山羊抗小鼠 c-kit 一抗发现 cPGCs 对 c-kit 表达呈阳性，这表明 cPGCs 能够感知 SCF，且 SCF/c-kit 可能参与趋化迁移。因此，提示 SCF 在鸡 cPGC 的迁移中起趋化因子的作用。
