Activation of endogenously expressed ion channels by active complement in the retinal pigment epithelium

作者：Genewsky Andreas; Jost Ingmar; Busch Catharina; Huber Christian; Stindl Julia; Skerka Christine; Zipfel Peter F; Rohrer Baerbel; Strauss Olaf^*

来源：Pflugers Archiv-European Journal of Physiology, 2015, 467(10): 2179-2191.

DOI：10.1007/s00424-014-1656-2

• (4-6): complement system
• Retinal pigment epithelium
• RPE
• L-type channels
• BK channels

Defective regulation of the alternative pathway of the complement system is believed to contribute to damage of retinal pigment epithelial (RPE) cells in age-related macular degeneration. Thus we investigated the effect of complement activation on the RPE cell membrane by analyzing changes in membrane conductance via patch-clamp techniques and Ca2+ imaging. Exposure of human ARPE-19 cells to complement-sufficient normal human serum (NHS) (25 %) resulted in a biphasic increase in intracellular free Ca2+ ([Ca2+](i)); an initial peak followed by sustained Ca2+ increase. C5- or C7-depleted sera did not fully reproduce the signal generated by NHS. The initial peak of the Ca2+ response was reduced by sarcoplasmic Ca2+-ATPase inhibitor thapsigargin, L-type channel blockers (R)-(+)-BayK8644 and isradipine, transient-receptor-potential (TRP) channel blocker ruthenium-red and ryanodine receptor blocker dantrolene. The sustained phase was carried by Ca(V)1.3 L-type channels via tyrosine-phosphorylation. Changes in [Ca2+](I) were accompanied by an abrupt hyperpolarization, resulting from a transient increase in membrane conductance, which was absent under extracellular Ca2+- or K+-free conditions and blocked by (R)-(+)-BayK8644 or paxilline, a maxiK channel inhibitor. Single-channel recordings confirmed the contribution of maxiK channels. Primary porcine RPE cells responded to NHS in a comparable manner. Pre-incubation with NHS reduced H2O2-induced cell death. In summary, in a concerted manner, C3a, C5a and sC5b-9 increased [Ca2+](i) by ryanodine-receptor-dependent activation of L-type channels in addition to maxi-K channels and TRP channels absent from any insertion of a lytic pore.

• 出版日期2015-10