ABSTRACTS – ORAL PRESENTATIONS - AMCA, spol. s r.o.

microarrays and high-throughput evaluation by flow cytometry, while detecting also
changes in posttranslational modification and cellular localization.
Acknowledgements
This work was supported by GAUK 596912, IGA NT13462, P302/12/G101, UNCE 204012,
00064203, IGA NT12397.
26. THE EFFECTS OF POTASSIUM CHANNEL INHIBITION ON CALCIUM INFLUX OF
PERIPHERAL T LYMPHOCYTES IN RHEUMATOID ARTHRITIS VERIFIED USING KINETIC
FLOW CYTOMETRY ANALYSIS
Gergely Toldi 1 *, Anna Bajnok 1 , Diána Dobi 2 , Ambrus Kaposi 1 , László Kovács 2 , Barna
Vásárhelyi 3 , Attila Balog 2
*toldigergely@yahoo.com
1
First Department of Pediatrics, Semmelweis University, Budapest, Hungary
2
Department of Rheumatology, University of Szeged, Szeged, Hungary
3
Department of Laboratory Medicine, Semmelweis University, Budapest, Hungary
Background: The transient increase of the cytoplasmic free calcium level plays a key
role in the process of lymphocyte activation. Kv1.3 and IKCa1 potassium channels are
important regulators of the maintenance of calcium influx during lymphocyte activation
and present a possible target for selective immunomodulation. We aimed to characterize
the effects of lymphocyte potassium channel inhibition on peripheral blood T lymphocyte
activation in rheumatoid arthritis (RA) compared to healthy individuals.
Methods: We isolated peripheral lymphocytes from 10 healthy controls and 9 recently
diagnosed RA patients receiving no anti-rheumatic treatment. We evaluated calcium
influx kinetics following activation in CD4, Th1, Th2 and CD8 cells. We also assessed the
sensitivity of the above subsets to specific inhibition of the Kv1.3 and IKCa1 potassium
channels.
Cells were stained with intracellular calcium binding dyes (Fluo-3 and Fura Red) and flow
cytometry analysis was performed in a kinetic manner (BD FACSAria). Data acquired from
the measurements were evaluated using a novel algorithm based on the calculation of
a double-logistic function for each recording (www.facskin.com). Specific parameter
values describing each function were also calculated and used to compare individual
measurements in an objective manner.
Results: The peak of calcium influx in lymphocytes isolated from RA patients is reached
more rapidly, indicating that they respond more quickly to stimulation compared to
controls. In healthy individuals, the inhibition of the IKCa1 channel decreased calcium
influx in Th2 and CD4 cells to a lower extent than in Th1 and CD8 cells. On the contrary,
the inhibition of Kv1.3 channels resulted in a larger decrease of calcium entry in Th2 and
CD4 than in Th1 and CD8 cells. No difference was detected between Th1 and Th2 or CD4
and CD8 cells in the sensitivity to IKCa1 channel inhibition among lymphocytes of RA
patients. However, specific inhibition of the Kv1.3 channel acts differentially on calcium
influx kinetics in RA lymphocyte subsets. Th2 and particularly CD8 cells are inhibited
Analytical Cytometry VII 51