Development and use of the lens epithelial explant system to study lens differentiation and cataractogenesis

Abstract

Over the last two decades much progress has been made in identifying and characterizing many of the molecules involved in understanding normal lens biology and its pathology. Much of this has been made possible through the establishment and use of the lens epithelial explant system. This simplistic tissue culture model, comprised of a sheet of lens epithelium on its native substratum, has been used effectively to study many cellular processes, including lens epithelial cell proliferation, fiber cell differentiation, cell apoptosis as well as epithelial-to-mesenchymal transformation of cells. In doing so, a number of key growth factors and cytokines, including members of the FGF, Wnt and TGFbeta family have been shown to play essential roles in many of these cellular events. This has led to further studies exploring the signaling pathways downstream of these molecules in the lens, paving the way for the development of a number of in situ models (primarily transgenic mouse lines) to further explore in more detail the nature of these molecular and cellular interactions. To reciprocate, the lens epithelial explant system is increasingly being used to further characterize the nature of many complex phenotypes and pathologies observed in these in situ models, allowing us to selectively isolate and examine the direct impact of an individual molecule on a specific cellular response in lens cells. There is no question that the lens epithelial explant system has served as a powerful tool to further our understanding of lens biology and pathology, and there is no doubt that it will continue to serve in such a capacity, as new developments are realized and putative treatments for aberrant lens cell behavior are to be trialed.

Figure 1

Schematic diagram demonstrating how the gradient of FGF stimulation affects the anteroposterior patterns of differentiation in the lens. The lens epithelium is frequently divided into two zones, the central lens epithelium (CE) and the germinative zone (GZ) which extends to the equator of the lens (EQ). The transitional zone (TZ), just posterior to the equator is where lens epithelial cells differentiate into fiber cells. Modified from (Lang and McAvoy 2004).

Figure 2

Representative figures of a rat lens epithelial explant. A. Bright field image demonstrating how an explant is pinned to the base of a tissue culture dish using fine forceps. B. Scanning electron micrograph of a rat lens epithelial explant pinned to the base of a tissue culture dish. C. Higher magnification view of explant (taken from box in B), demonstrating tightly packed sheet of lens epithelial cells which cover the surface of the lens epithelial explant. Modified from Lovicu and McAvoy, 2008.

Figure 3

Schematic diagram illustrating FGF and TGFβ induction of lens epithelial cells into lens fiber cells or myofibroblasts cells, respectively. Modified from (Lang and McAvoy 2004)

Figure 4

Lens epithelial explant culture of 21 day old rats, immunostained for α-SMA. Untreated explants do not exhibit αSMA immunoreactivity (A), whereas explants treated with 4ng/ml of TGFβ-2 for 4 days exhibit α-SMA accumulation (green) (B). The co-treatment of explants with TGFβ-2 (4ng/ml) and 25μM of MMPI-2/-9 (C), resulted in the suppression of α-SMA accumulation. Blue represents 4′,6-diamidino-2-phenylindole (DAPI) staining for nuclei.