Potassium Transport in Lens Cells Derived From Myotonic Dystrophy Cataracts

Abstract

Abstract: : Purpose: Previous work has identified a role for calcium–activated potassium channels in myotonic dystrophy cataract. We have, therefore, investigated potassium transport in lens cell lines derived from cataract patients with and without myotonic dystrophy (DM). Methods: A total of 4 cell lines were derived by SV40 transformation of lens capsulorhexis specimens obtained immediately following cataract surgery. Rhexis samples were collected in culture medium and cells were infected with adenovirus carrying SV40 large T. Two age matched lines from routine cataracts (CCat1 and 2) and two from DM cataracts (DMCat1 and 2) were used in this study. The CTGn triplet repeat expansion was measured using Small Pool Polymerase Chain Reaction. ⁸⁶Rubidium (⁸⁶Rb) was used as a tracer ion to measure potassium transport and ionomycin to stimulate internal calcium in efflux experiments. Results: All the DM cell lines had characteristic CTG repeat expansions ranging from ∼450 to 2950. The DM cells also displayed much slower growth and exhibited senescence at much lower passage numbers than the non–DM cells. The ⁸⁶Rb uptake kinetics were, however, similar in all cell lines and could be fitted by a single exponential with a rate constant 0.007 min^–1. The uptake reached a steady value of approximately 875 nmol Rb mg^–1 protein. The influx rate was 6.75 nmol min^–1 mg^–1 for all cell lines. Ouabain (1 mM) reduced the influxes by 50% and 53% in CCat and DMCat cells respectively. The efflux rate constant in control conditions was 0.007 min^–1 in both types of cell line showing that the cells were in steady state with respect to potassium transport during the course of the experiment. Ionomycin increased the rate 4 fold for CCat1 and CCat2 and >10 fold for DMCat1 and 2. Quinine (300 µM) significantly inhibited the ionomycin stimulated efflux in all cases indicating the involvement of calcium activated potassium channels. Conclusions: The DM cells exhibited considerable CTG repeat expansions that are characteristic of myotonic dystrophy. They exhibited slower growth characteristics and the onset of senescence was greatly accelerated. Calcium stimulated potassium efflux was much greater in the DM cells and we have previously suggested a role for calcium activated potassium channels in DM cataract. DM derived lens cell lines appear to provide an excellent model for the investigation of the molecular mechanisms of DM cataract.

Keywords: ion channels • cataract • calcium