Calcium-induced changes on crystallins in organ-cultured porcine lens

In this study, intact porcine lenses were cultured in vitro for 7 days supplemented with commercial balanced salt solution (BSS) which is usually used as an irrigation solution during intraocular surgery, and the lenses were maintained under various culture conditions, e.g. temperature and CO2 concentration. The intact porcine lenses after 7 days culture were analyzed with optical density scanner, gel permeation chromatography on TSK HM-55 column and SDS-PAGE (polyacrylamide gel electrophoresis). It was found that lenses exhibited the least opacity when lenses were cultured with Ca(+2)-free BSS buffer, CO2-free incubator and maintained at a temperature of 25 degrees C. After the lenses were cultured with Ca(+2)-free BSS or BSS medium, the composition of crystallins in lenses was separated with TSK HM-55 column. It was indicated that the percentage of high molecular weight (HMW) protein and (alpha-crystallin increased, and gamma-crystallin decreased in lenses incubated with BSS medium compared with lenses incubated with Ca(+2)-free BSS medium. Following an increase in the concentration of calcium in the medium from 4.3 mM, 20 mM, 50 mM, 100 mM to 200 mM, the opacity of the lens was measured with a densitometer. The changed percentage of various crystallins was similar to lenses with BSS media that increased in HMW protein and alpha-crystallin, decreasing in gamma-crystallin. In the case of lens protein pattern, the crystallin washed from TSK HM-55 gel was separated with SDS-PAGE (polyacrylamide gel electrophoresis). It was indicated that some of proteins disappeared when lenses were incubated with various concentrations of calcium. The vanished pH proteins were 20.5 kDa at 50 mM calcium, 20.5 kDa and 21 kDa at 100 mM, 20.5 kDa, 21 kDa, 22 kDa and 23 kDa at 200 mM which were compared with the protein bands in the presence of 20 mM calcium in BSS medium. This study indicates that the commercial balanced salt solution (BSS) which is usually used as an irrigating solution during intraocular operations may increase the risk for lens opacity because of the calcium contained in the solution.