Overexpression of Delayed Rectifier K(+) Channels Promotes In situ Proliferation of Leukocytes in Rat Kidneys with Advanced Chronic Renal Failure

Abstract

Leukocytes, such as lymphocytes and macrophages, predominantly express delayed rectifier K(+) channels (Kv1.3), and the channels play crucial roles in the activation and proliferation of the cells. Since lymphocytes are activated in patients with end-stage renal disease (ESRD), the channels expressed in those cells would contribute to the progression of renal fibrosis in advanced-stage chronic renal failure (CRF). In the present study, using a rat model with advanced CRF that underwent 5/6 nephrectomy followed by a 14-week recovery period, we examined the histopathological features of the kidneys and the leukocyte expression of Kv1.3-channels and cell cycle markers. Age-matched sham-operated rats were used as controls. In the cortical interstitium of advanced CRF rat kidneys, leukocytes proliferated in situ and overexpressed Kv1.3 channel protein in their cytoplasm. Treatment with margatoxin, a selective Kv1.3-channel inhibitor, significantly suppressed the number of leukocytes and the progression of renal fibrosis with a significant decrease in the cortical cell cycle marker expression. This study demonstrated for the first time that the number of leukocytes was dramatically increased in rat kidneys with advanced CRF. The overexpression of Kv1.3 channels in the leukocytes was thought to contribute to the progression of renal fibrosis by stimulating cell cycling and promoting cellular proliferation.

Figure 1

Histological features of sham operated (sham) and advanced CRF rat kidneys. (A) Hematoxylin and eosin staining (H&E) in sham operated (sham) and advanced CRF rat kidneys. (a) and (b) Low-power views of cortex. Magnification, ×20. (c) and (d) High-power views of cortical interstitium. Magnification, ×60. (B) The mRNA abundance of CD3 (left) and ED-1 (right) in the renal cortex of sham operated and advanced CRF (CRF) rat kidneys. ^# P < 0.05 versus sham operated rats. Values are means ± SEM (n = 6). Differences were analyzed by ANOVA followed by Dunnett's or Student's t-test. (C) Immunohistochemistry using antibody for Ki-67 (brown) in sham operated and advanced CRF rat kidneys. (a) and (b) High-power views of cortical interstitium. Magnification, ×60.

Figure 2

Kv1.3 and cell cycle marker expression in sham operated (sham) and advanced CRF rat kidneys. (A) Kv1.3 expression. (a) KCNA3 mRNA abundance in the renal cortex of sham operated (sham) and advanced CRF (CRF) rat kidneys. (b) and (c) Immunohistochemistry using antibody for Kv1.3 (brown) in sham operated and advanced CRF rat kidneys. High-power views of cortical interstitium. Magnification, ×60. (B) Cell cycle marker expression. The mRNA abundance of cyclin-dependent kinase 4 (Cdk4) (left) and p21 (right) in the renal cortex of sham operated and advanced CRF rat kidneys. ^# P < 0.05 versus sham operated rats. Values are means ± SEM (n = 6). Differences were analyzed by ANOVA followed by Dunnett's or Student's t-test.

Figure 3

Collagen III and cell cycle marker expression in advanced CRF rat kidneys after margatoxin (MgTX) treatment. (A) Hematoxylin and eosin staining (H&E) in advanced CRF rat kidneys after margatoxin (MgTX) treatment. (a) A low-power view of cortex. Magnification, ×20. (b) A high-power view of cortical interstitium. Magnification, ×60. (B) Immunohistochemistry using antibody for collagen III (brown) in advanced CRF rat kidneys with and without MgTX treatment. (a) and (b) Low-power views of cortex. Magnification, ×20. (C) Cell cycle marker expression. The mRNA abundance of Cdk4 (left) and p21 (right) in the renal cortex of advanced CRF rat kidneys with and without MgTX treatment. *P < 0.05 versus advanced CRF rats without treatment. Values are means ± SEM (n = 6). Differences were analyzed by ANOVA followed by Dunnett's or Student's t-test.