The catalytic subunit of Na+,K+-ATPase has at least four isoforms, α1, α2, α3 and α4, each derived from a different gene. The α-isoform exhibits a tissue-specific pattern of expression (Blanco & Mercer, 1998). α1-isoform is found in nearly every tissue and involved in a housekeeping function in all cells. α2-isoform is predominantly expressed in adipocytes, skeletal muscle, heart and brain. α3-isoform is abundant in brain. α4-isoform is expressed exclusively in testis. It has been reported that no significant change in the expression of α1-isoform was observed in human renal and lung carcinomas (Akopyanz et al. 1991; Rajasekaran et al. 1999).
Herein we investigated whether levels of protein expression of Na+,K+-ATPase α1-, α2- and α3-isoforms could be changed in human colorectal cancers. The specimens of colorectal adenocarcinomas and accompanying normal mucosa were obtained from surgical resection of patients in accordance with the recommendations of the Declaration of Helsinki and with ethics committee approval. Informed consents were obtained from all patients at Toyama Medical and Pharmaceutical University Hospital. Data are shown as means ± S.E.M. Difference between groups were analysed by one-way ANOVA. Comparison between the two groups was made with paired t test.
Western blotting was performed using specific antibodies against α1-, α2- or α3-isoform. Specificity of these antibodies was confirmed using the control antigen peptides. Interestingly, decrease in the expression of α1-isoform was observed in 12 of 12 cancers (100 %) compared with the normal mucosa. No significant expression of α2-isoform was observed in any of the colorectal carcinomas and accompanying normal mucosa. On the other hand, increase in the expression of α3-isoform was observed in 9 of 12 cancers (75 %) compared with the normal mucosa. Simultaneous change of expression of α1-isoform and α3-isoform was found in 6 of 12 cancers, and ouabain (5 µM)-sensitive ATPase activities of these cancer tissues and accompanying normal mucosa were 2.5 ± 0.6 and 3.2 ± 0.4 µmol Pi (mg protein)-1 h-1, respectively (n = 9).
These results suggest that changes in protein expression of α1- and α3-isoforms but not α2-isoform are associated with human colorectal cancers.