Figure 1A shows a precataractous (21-day old), visibly transparent ICR/f rat lens, and
Figure 1B shows a cataractous (100-day old) ICR/f rat lens in which there is a dense opacification throughout the entirety of the lens. Lenses such as these were used to evaluate changes in protein distribution and solubility during cataract development.
Figure 2 shows the sectioned lens tissues and selected
m/z signals that were obtained from 21-day (
Figs. 2A–D) and 100-day (
Figs. 2E–H) ICR/f rat lenses. In the optical images (
Figs. 2A,
2E), there are obvious regions that are segmented by slight color changes (from white to translucent against the gold background of the MALDI target plate) that appear at various radii from the center of the tissue. These regions represent different stages of lens development, as the nuclear center was developed during embryogenesis with subsequent addition of lens fiber cells.
19 From this, it is plausible to predict regional differences in protein signals, as these regions were formed at different points during lens development. We confirmed this by extracting images from the MSI experiment as indicated in
Figures 2B–D and
2F–H. For this purpose and for reasons discussed later, two of the major protein species present in the MSI experiment were chosen for analysis. The first protein is the N-terminally acetylated, full-length αA-crystallin (residues 1-173) protein, at an average mass of 19,835 Da ± 0.06% (
Figs. 2B,
2D,
2F,
2H, blue). The second protein is an N-terminally acetylated αA-crystallin truncation product (residues 1-53), at an average mass of 6413 Da ± 0.11% (
Fig. 2C,
2D,
2G,
2H, orange). Although other MSI studies have localized low-molecular-weight αA-crystallin truncation products (1-40, 1-50, and 1-58) to the nuclear core of other species,
20–22 this is study is the first to identify the 1-53 truncation product as a major protein present in the lens.