X–Ray Diffraction Patterns and Protein Concentrations in Developing Rat Lens

Abstract

Purpose: : It has been reported that the small–angle X–ray diffraction pattern analysis could evaluate the short–range order of crystallins that was necessary for lens transparency. Developmentally, lens fiber cells continue differentiating and the rate of protein synthesis changes dramatically. Therefore, we investigated the time course of X–ray diffraction patterns and protein concentrations in developing rat lens to reach a better understanding of the required conditions to keep transparency even with high protein concentration.

Methods: : We obtained X–ray diffraction patterns from excised whole rat lens and measured protein concentrations at postnatal day 5 (P5), P10, and P15. The X–ray experiments were performed at 37°C. The X–ray beam passed lens–center in a direction parallel to optic axis. Protein concentrations of lenses were measured by Bicinchonianate method, using bovine serum albumin as standard. Lens volume was obtained from weight and specific gravity.

Results: : Although scattering patterns of X–ray were observed at P5 and P10, the diffraction pattern of broad ring with a characteristic spacing of ∼15 nm appeared at P15. Total protein concentration of the lens fiber cell cytoplasm significantly increased especially from P10 to P15 (274.4 ± 39.6, 347.4 ± 41.6, and 569.0 ± 60.7 [mg/ml lens volume] at P5, P10, and P15, respectively, mean ± SD, each group: n = 5). All lenses at each postnatal day were transparent at 37°C and therefore short–range order of crystallins may not necessary to keep transparent at least with relatively lower protein concentration at physiological temperature.

Conclusions: : Short–range order of crystalline developed with a marked increase of protein concentration of the lens fiber cell cytoplasm in short time.

Keywords: crystalline lens • development