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P.R. Cammarata, Z. Wang, Z. Guo, A. Moor; Classification of Estrogen Receptor(s) in Cultured Human Lens Epithelial Cells . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1221.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Epidemiological studies have identified a higher rate of occurrence of cataract in estrogen-deficient postmenopausal women. A plausible explanation might be found in our recent findings that estrogens take action against oxidative damage in human lens epithelial (HLE-B3) cells in culture through protection of mitochondrial membrane potential, intracellular ATP and cell viability (Wang et al., Invest Ophthalmol Vis Sci.2002; 43:a849). Whether estrogen exerts its antioxidant activities in HLE-B3 cells through estrogen receptor-independent mechanisms and/or via genomic interaction remains to be determined. In this study, we document the expression and cellular localization of the estrogen receptors, ERα and ERß, in cultured HLE-B3 cells. Methods: ERα and ERß mRNA expression was evaluated by coupled RT-PCR and Southern blot analysis. Cellular localization of ERα and ERß was determined on formaldehyde-fixed, triton-permeabilized cells using conventional immunofluorescence techniques. Results: Using RT-PCR, estrogen receptor specific primers distinguished mRNA from total RNA extracted from HLE-B3 cells, as well as from human breast carcinoma cells (mcf-7), which provided a positive control. The expected 236-bp (ERα) and 167-bp (ERß) bands were authenticated by sequence analysis. Southern blot analysis using internal oligonucleotides directed to specific primer pairs for ERα and ERß, respectively, further confirmed the specificity of the PCR products and no overlapping recognition of ERα and ERß PCR products was observed with either probe. The level of ERα and ERß PCR product, as determined by RT-PCR or Southern blot analysis in mcf-7 cells, greatly exceeded that detected in HLE-B3 cells, indicative of the relative scarcity of either receptor in the latter cell. Immunofluorescence staining of ERα and ERß corroborated mRNA expression in HLE-B3 cells. While ERα staining tended to appear more perinuclear, ERß staining was more widespread throughout the cytoplasm. Conclusions: The occurrence of ERα and ERß in human lens epithelial cells suggests that gender-specific steroids play a role in the pathophysiology of the lens. The possible difference in the comparative cytoplasmic or nuclear distribution of ERα and ERß raises the interesting probability that the two receptors may have differential cytoprotective potential, and by inference, disparity in their mechanisms of action.
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