Background: Anti-allergic drugs, such as tranilast and ketotifen, inhibit the release of chemokines from mast cells. However, we know little about their direct effects on the exocytotic process of mast cells. Since exocytosis in mast cells can be monitored electrophysiologically by changes in the whole-cell membrane capacitance (Cm), the absence of such changes by these drugs indicates their mast cell-stabilizing properties. Methods: Employing the standard patch-clamp whole-cell recording technique in rat peritoneal mast cells, we examined the effects of tranilast and ketotifen on the Cm during exocytosis. Using confocal imaging of a water-soluble fluorescent dye, lucifer yellow, we also examined their effects on the deformation of the plasma membrane. Results: Relatively lower concentrations of tranilast (100, 250 µM) and ketotifen (1, 10 µM) did not significantly affect the GTP-γ-S-induced increase in the Cm. However, higher concentrations of tranilast (500 µM, 1 mM) and ketotifen (50, 100 µM) almost totally suppressed the increase in the Cm, and washed out the trapping of the dye on the surface of the mast cells. Compared to tranilast, ketotifen required much lower doses to similarly inhibit the degranulation of mast cells or the increase in the Cm. Conclusions: This study provides electrophysiological evidence for the first time that tranilast and ketotifen dose-dependently inhibit the process of exocytosis, and that ketotifen is more potent than tranilast in stabilizing mast cells. The mast cell-stabilizing properties of these drugs may be attributed to their ability to counteract the plasma membrane deformation in degranulating mast cells.

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.

Background: Macrolides, such as clarithromycin, have antiallergic properties. Since exocytosis in mast cells is detected electrophysiologically via changes in membrane capacitance (Cm), the absence of such changes due to the drug indicates its mast cell-stabilizing effect. Methods: Employing the whole-cell patch clamp technique in rat peritoneal mast cells, we examined the effects of clarithromycin on Cm during exocytosis. Using a water-soluble fluorescent dye, we also examined its effect on deformation of the plasma membrane. Results: Clarithromycin (10 and 100 μM) significantly inhibited degranulation from mast cells and almost totally suppressed the GTP-γ-S-induced increase in Cm. It washed out the trapping of the dye on the surface of mast cells. Conclusions: This study provides for the first time electrophysiological evidence that clarithromycin dose-dependently inhibits the process of exocytosis. The mast cell-stabilizing action of clarithromycin may be attributable to its counteractive effect on plasma membrane deformation induced by exocytosis.

Background/Aims: Leukocytes, such as lymphocytes and macrophages, predominantly express delayed rectifier K⁺ channels (Kv1.3) in their plasma membranes. In our previous study, the overexpression of these channels in leukocytes was strongly associated with their proliferation in kidneys and the progression of renal fibrosis in advanced-stage chronic renal failure (CRF). Since benidipine, a long-acting 1,4-dihydropyridine Ca²⁺ channel blocker, is also highly potent as a Kv1.3 channel inhibitor, it could exert therapeutic efficacy in advanced CRF. Methods: Male Sprague-Dawley rats that underwent 5/6 nephrectomy followed by a 14-week recovery period were used as the model of advanced CRF. Benidipine hydrochloride (5 mg/kg) was started at 8 weeks after nephrectomy and orally administered daily for 6 weeks. The histopathological features of the kidneys were examined in vehicle-treated and benidipine-treated CRF rat kidneys. Cellular proliferation of leukocytes and the cortical expression of proinflammatory cytokines were also examined. Results: In CRF rat kidneys, Kv1.3 channels began to be overexpressed in leukocytes as early as 8 weeks after nephrectomy. In the cortical interstitium of benidipine-treated CRF rat kidneys, both immunohistochemistry and real-time PCR demonstrated decreased expression of fibrotic markers. Benidipine treatment significantly reduced the number of proliferating leukocytes within the cortical interstitium and decreased the expression of cell cycle markers and proinflammatory cytokines. Conclusion: This study demonstrated for the first time that benidipine slowed the progression of renal fibrosis in rat kidneys with advanced CRF. Kv1.3 channels overexpressed in leukocytes were thought to be the most likely therapeutic targets of benidipine in decreasing the number of proliferating leukocytes and repressing the production of inflammatory cytokines.

This study demonstrated for the first time that the number of mast cells was significantly increased in the fibrotic peritoneum of CRF rats. The proliferation of mast cells and their increased activity in the peritoneum were thought to be responsible for the progression of peritoneal fibrosis.