Three-dimensional organization of primary lens fiber cells

RESULTS. Injection of either vector resulted in GFP expression in primary fiber cells. GFP-expressing cells were heterogeneous in shape and length. Some regions of the fibers were varicose, with diameters .10 mm; regions between the varicosities were often extremely thin, with diameters of ,2 mm. No differences in the morphologies of GFP-expressing cells were noted between adenovirus- and plasmid-injected lenses, suggesting that the irregular, undulating, appearance of the primary fibers was not the result of viral infection. Three-dimensional reconstruction of primary fiber cells revealed that, by E6, the posterior tips of the fibers had detached from the lens capsule. The anterior fiber tips remained in contact with the overlying epithelium for 1 to 2 additional days, demonstrating that the formation of the anterior and posterior sutures was asynchronous. CONCLUSIONS. The three-dimensional cellular organization of GFP-expressing cells is consistent with previous analyses of fiber cell morphology in the embryonic nucleus of adult human and bovine lenses. The present data confirm that the disorganized appearance of primary fiber cells observed in adult lenses is largely a reflection of developmental processes rather than a consequence of aging. (Invest Ophthalmol Vis Sci. 2000;41:859 ‐ 863) P rimary fiber cells are formed early in embryonic development by elongation of cells at the posterior of the lens vesicle. The elongating primary fiber cells rapidly obliterate the lumen of the lens vesicle. Subsequently, differentiation of cells at the lens equator leads to the formation of secondary fiber cells that come to overlie the primary fibers. Formation of secondary fiber cells occurs continuously throughout life with the original primary fiber cells becoming progressively buried. In the adult lens, the primary fiber cells constitute only a small fraction of the total number of lens cells and are located in the center of the tissue in a region termed the embryonic nucleus. Compared to the regular and well-ordered layers of secondary fiber cells, the primary fiber cells are known to be a rather heterogeneous cell population. In human lenses, for example, electron microscopic analysis of lenses revealed that the cross-sectional area of primary fiber cells was 80 6 68 mm 2 (mean 6 SD) with a range of 7 to 308 mm 2 . 1 In contrast, the cross-sectional area of secondary fiber cells in the cortex of the lens was found to be much more uniform (24 6 9 mm 2 ). In another study, light microscopic analysis of bovine lenses also indicated that the cross-sectional profiles of primary fiber cells were very heterogeneous, with an average value of 63 6 61 mm 2 . 2 These data suggest that the cellular morphology and organization of primary fiber cells is very different from that of the secondary fiber cells, which make up the majority of the tissue volume. Although the architecture of the lens nucleus has been studied extensively by scanning electron microscopy, 3‐5 the three-dimensional organization of individual primary fiber cells has not been fully elucidated. Consequently, the cellular features that give rise to the large, observed variations in cross-sectional area have yet to be determined. Here we report a novel approach for visualizing the three-dimensional structural organization of the primary fiber cell population in intact or sliced embryonic lens preparations. The technique involves the use of adenoviral and plasmid expression vectors carrying the gene for Green Fluorescent Protein (GFP), an autofluorescent protein isolated from the jellyfish Aquorea victoria. 6