Different effects of cyclosporine A and FK506 on potassium transport systems in MDCK cells

Background. Hyperkalemia and metabolic acidosis are common manifestations i n patients receiving the immunosuppressive agent cyclosporine A (CsA) and t he recently introduced FK506. We compared the acute toxic and antiprolifera tive effects as well as the effects on the transport activity of Na+/K+-ATP ase and Na+/K+/2Cl(-) cotransporter of CsA and FK506 in an established cell line of distal/collecting tubule origin (MDCK cells). Methods: MDCK cells were exposed to various concentrations of CsA or FK506 and the effects on c ell viability (MTT test and neutral red uptake), plasma membrane integrity (lactate dehydrogenase (LDH) release) and cell proliferation (bromodeoxyuri dine (BrdU) incorporation) were compared. For transport studies, after conf luence, MDCK cells were exposed to CsA or FK506 for 48 h in the presence an d absence of aldosterone. Ouabain- and bumetanide-sensitive (86)Rubidium up take measurements were used to study the activity of the Na+/K+-ATPase and Na+/K+/2Cl(-) cotransporter at the surface of intact cells. Results: After 24 h of exposure CsA reduced the number of viable cells to 50% at 30 muM, w hereas for FK506 2-3 times higher concentrations had to be employed. Simila rly, LDH release was stimulated tenfold by 30 muM CsA but only fourfold by 70 muM FK506. In contrast, DNA synthesis was affected at lower concentratio ns of FK506 than of CsA. In cells treated for 24 h BrdU incorporation was s ignificantly inhibited by 3 muM FK506, whereas a similar inhibition require d 10 muM CsA. The transport activity of Na+/K+-ATPase and of Na+/K+/2Cl(-) cotransporter were significantly decreased (37 and 63%, respectively) on Cs A administration (8 muM). In CsA-treated cells the K+ channel blockers bari um (1 mM), TEA (10 mM) and quinine (1 mM) did not further inhibit the trans port activities suggesting that CsA might also act via inhibition of K+ cha nnels. FK506 at 8 muM had no effect on Na+/K+-ATPase transport activity but stimulated Na+/K+/2Cl(-) cotransporter activity by 59%. The stimulatory ef fect was abolished by K+ channel blockers indicating that recycling of K+ m ight increase by FK506. The simultaneous presence of aldosterone (5 muM) pr otected the cells from the inhibitory effect of CsA on Na+/K+-ATPase and Na +/K+/2Cl(-) cotransporter activity. The stimulatory effect of FK506 on the Na+/K+/2Cl(-)cotransporter activity was completely abolished in the presenc e of aldosterone. Conclusions: Both CsA and FK506 showed acute toxicity in MDCK cells in vitro with the effects of FK506 being less pronounced. CsA an d FK506 had different effects on the in vivo transport rates of the Na+/K+- ATPase and the Na+/K+/2Cl(-) cotransporter; CsA inhibited the activity of t he Na+/K+-ATPase and the Na+/K+/2Cl(-) cotransporter whereas FK506 stimulat ed the activity of Na+/K+/2Cl(-) cotransporter. These effects were abolishe d by the application of aldosterone. Copyright (C) 2001 S. Karger AG, Basel .