Department of Biological Science, National University of Singapore Kent Ridge, Singapore, Singapore.
Front Physiol. 2013 Jun 7;4:135. doi: 10.3389/fphys.2013.00135. eCollection 2013.
This study aimed to test the hypothesis that branchial osmoregulatory acclimation involved increased apoptosis and replacement of mitochdonrion-rich cells (MRCs) in the climbing perch, Anabas testudineus, during a progressive acclimation from freshwater to seawater. A significant increase in branchial caspase-3/-7 activity was observed on day 4 (salinity 20), and an extensive TUNEL-positive apoptosis was detected on day 5 (salinity 25), indicating salinity-induced apoptosis had occurred. This was further supported by an up-regulation of branchial mRNA expression of p53, a key regulator of cell cycle arrest and apoptosis, between day 2 (salinity 10) and day 6 (seawater), and an increase in branchial p53 protein abundance on day 6. Seawater acclimation apparently activated both the extrinsic and intrinsic pathways, as reflected by significant increases in branchial caspase-8 and caspase-9 activities. The involvement of the intrinsic pathway was confirmed by the significant increase in branchial mRNA expression of bax between day 4 (salinity 20) and day 6 (seawater). Western blotting results revealed the presence of a freshwater Na(+)/K(+)-ATPase (Nka) α-isoform, Nka α1a, and a seawater isoform, Nka α1b, the protein abundance of which decreased and increased, respectively, during seawater acclimation. Immunofluorescence microscopy revealed the presence of two types of MRCs distinctly different in sizes, and confirmed that the reduction in Nka α1a expression, and the prominent increases in expression of Nka α1b, Na(+):K(+):2Cl(-) cotransporter 1, and cystic fibrosis transmembrane conductance regulator Cl(-) channel coincided with the salinity-induced apoptotic event. Since modulation of existing MRCs alone could not have led to extensive salinity-induced apoptosis, it is probable that some, if not all, freshwater-type MRCs could have been removed through increased apoptosis and subsequently replaced by seawater-type MRCs in the gills of A. testudineus during seawater acclimation.
在从淡水到海水的逐渐适应过程中，攀鲈鳃中的渗透调节适应性涉及到凋亡的增加和富含线粒体的细胞（MRC）的替换。在第 4 天（盐度 20）观察到鳃 caspase-3/-7 活性显著增加，第 5 天（盐度 25）检测到广泛的 TUNEL 阳性凋亡，表明盐度诱导的凋亡已经发生。这进一步得到了以下证据的支持：从第 2 天（盐度 10）到第 6 天（海水），鳃中 p53 的 mRNA 表达上调，p53 是细胞周期停滞和凋亡的关键调节因子，以及第 6 天鳃中 p53 蛋白丰度增加。海水适应显然激活了外在和内在途径，这反映在鳃 caspase-8 和 caspase-9 活性的显著增加上。内在途径的参与通过第 4 天（盐度 20）和第 6 天（海水）之间鳃 bax mRNA 表达的显著增加得到了证实。Western blot 结果显示存在淡水 Na(+)/K(+)-ATPase (Nka) α-同工型，Nka α1a，和海水同工型，Nka α1b，其蛋白丰度在海水适应过程中分别降低和增加。免疫荧光显微镜显示存在两种大小明显不同的 MRC 类型，并证实 Nka α1a 表达的减少，以及 Nka α1b、Na(+):K(+):2Cl(-)共转运蛋白 1 和囊性纤维化跨膜电导调节因子 Cl(-)通道表达的显著增加，与盐度诱导的凋亡事件相吻合。由于仅调节现有的 MRC 不可能导致广泛的盐度诱导凋亡，因此，在攀鲈的鳃中，一些（如果不是全部）淡水型 MRC 可能通过凋亡增加而被去除，随后在海水适应过程中被海水型 MRC 取代。
