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Increases in apoptosis, caspase activity and expression of p53 and bax, and the transition between two types of mitochondrion-rich cells, in the gills of the climbing perch, Anabas testudineus, during a progressive acclimation from freshwater to seawater.

Ching B, Chen XL, Yong JH, Wilson JM, Hiong KC, Sim EW, Wong WP, Lam SH, Chew SF, Ip YK - Front Physiol (2013)

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Science, National University of Singapore Kent Ridge, Singapore, Singapore.

ABSTRACT
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.

No MeSH data available.


Related in: MedlinePlus

Protein abundance of Na+/K+-ATPase α1a (Nka α1a) in the gills of Anabas testudineus kept in freshwater on day 1 (FW; control), or after 24 h of exposure to salinity 15 on day 3, salinity 25 on day 5, or salinity 30 (seawater) on day 6 during a progressive acclimation from freshwater to seawater. (A) Immunoblots of Nka α 1a and actin. (B) The intensity of the Nka α 1a protein band normalized with respect to actin. Results represent mean + S.E.M. (N = 4). Means not sharing the same letter are significantly different (P < 0.05).
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Figure 8: Protein abundance of Na+/K+-ATPase α1a (Nka α1a) in the gills of Anabas testudineus kept in freshwater on day 1 (FW; control), or after 24 h of exposure to salinity 15 on day 3, salinity 25 on day 5, or salinity 30 (seawater) on day 6 during a progressive acclimation from freshwater to seawater. (A) Immunoblots of Nka α 1a and actin. (B) The intensity of the Nka α 1a protein band normalized with respect to actin. Results represent mean + S.E.M. (N = 4). Means not sharing the same letter are significantly different (P < 0.05).

Mentions: Immunoblotting of gill samples from A. testudineus exposed to a progressive increase in salinity from freshwater to seawater using custom-made anti-Nka α1a or anti-Nka α1b monoclonal antibodies confirmed that Nka α 1a was a freshwater isoform while Nka α 1b was a seawater isoform. There was a significant decrease in the protein abundance of Nka α 1a in the gills of fish exposed to salinity 25 (day 5) and to seawater (day 6; Figure 8). By contrast, there was a significant increase in the protein abundance of Nka α 1b in the gills of these experimental fish (Figure 9). Overall, there was a significant increase in the protein abundance of Nka α-subunit, as demonstrated by the pan-specific anti-NKA (α 5) antibody, in gills of A. testudineus acclimated progressively from freshwater to seawater (Figure 10).


Increases in apoptosis, caspase activity and expression of p53 and bax, and the transition between two types of mitochondrion-rich cells, in the gills of the climbing perch, Anabas testudineus, during a progressive acclimation from freshwater to seawater.

Ching B, Chen XL, Yong JH, Wilson JM, Hiong KC, Sim EW, Wong WP, Lam SH, Chew SF, Ip YK - Front Physiol (2013)

Protein abundance of Na+/K+-ATPase α1a (Nka α1a) in the gills of Anabas testudineus kept in freshwater on day 1 (FW; control), or after 24 h of exposure to salinity 15 on day 3, salinity 25 on day 5, or salinity 30 (seawater) on day 6 during a progressive acclimation from freshwater to seawater. (A) Immunoblots of Nka α 1a and actin. (B) The intensity of the Nka α 1a protein band normalized with respect to actin. Results represent mean + S.E.M. (N = 4). Means not sharing the same letter are significantly different (P < 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3675322&req=5

Figure 8: Protein abundance of Na+/K+-ATPase α1a (Nka α1a) in the gills of Anabas testudineus kept in freshwater on day 1 (FW; control), or after 24 h of exposure to salinity 15 on day 3, salinity 25 on day 5, or salinity 30 (seawater) on day 6 during a progressive acclimation from freshwater to seawater. (A) Immunoblots of Nka α 1a and actin. (B) The intensity of the Nka α 1a protein band normalized with respect to actin. Results represent mean + S.E.M. (N = 4). Means not sharing the same letter are significantly different (P < 0.05).
Mentions: Immunoblotting of gill samples from A. testudineus exposed to a progressive increase in salinity from freshwater to seawater using custom-made anti-Nka α1a or anti-Nka α1b monoclonal antibodies confirmed that Nka α 1a was a freshwater isoform while Nka α 1b was a seawater isoform. There was a significant decrease in the protein abundance of Nka α 1a in the gills of fish exposed to salinity 25 (day 5) and to seawater (day 6; Figure 8). By contrast, there was a significant increase in the protein abundance of Nka α 1b in the gills of these experimental fish (Figure 9). Overall, there was a significant increase in the protein abundance of Nka α-subunit, as demonstrated by the pan-specific anti-NKA (α 5) antibody, in gills of A. testudineus acclimated progressively from freshwater to seawater (Figure 10).

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Science, National University of Singapore Kent Ridge, Singapore, Singapore.

ABSTRACT
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.

No MeSH data available.


Related in: MedlinePlus