Eluate derived by extracorporal antibody-based immunoadsorption elevates the cytosolic Ca2+ concentration in podocytes via B2 kinin receptors.

Background/aim: Patients with idiopathic focal segmental glomerulosclerosis (FSGS) often develop a recurrence of the disease after kidney transplantation. In a number of FSGS patients, plasmapheresis and immunoadsorption procedures have been shown to transiently reduce proteinuria and are thought to do this by eliminating a circulating factor. Direct cellular effects of eluates from immunoadsorption procedures on podocytes, the primary target of injury in FSGS, have not yet been reported.

Methods: Eluates were derived from antibody-based immunoadsorption of a patient suffering from primary FSGS, a patient with systemic lupus erythematosus, and a healthy volunteer. The cytosolic free Ca(2+) concentration ([Ca(2+)](i)) of differentiated podocytes was measured by single-cell fura-2 microfluorescence measurements. Free and total immunoreactive kinin levels were measured by radioimmunoassay.

Results: FSGS eluates increased the [Ca(2+)](i) levels concentration dependently (EC(50) 0.14 mg/ml; n = 3-19). 1 mg/ml eluate increased the [Ca(2+)](i) values reversibly from 82 +/- 12 to 1,462 +/- 370 nmol/l, and then they returned back to 100 +/- 16 nmol/l (n = 19). The eluate-induced increase of [Ca(2+)](i) consisted of an initial Ca(2+) peak followed by a Ca(2+) plateau which depended on the extracellular Ca(2+) concentration. The eluate-induced increase of [Ca(2+)](i) was inhibited by the specific B(2) kinin receptor antagonist Hoe 140 in a concentration-dependent manner (IC(50) 2.47 nmol/l). In addition, prior repetitive application of bradykinin desensitized the effect of eluate on [Ca(2+)](i). A colonic epithelial cell line not reacting to bradykinin did not respond to eluate either (n = 6). Similar to FSGS eluates, the eluate preparations of both the systemic lupus patient and the healthy volunteer led to a biphasic, concentration-dependent [Ca(2+)](i) increase in podocytes which again was inhibited by Hoe 140. Free kinins were detected in all eluate preparations.

Conclusion: The procedure of antibody-based immunoadsorption leads to kinin in the eluate which elevates the [Ca(2+)](i) level of podocytes via B(2) kinin receptors.