PURPOSE: The effects of retinoic acid in cells are mediated by the nuclear retinoic acid receptors (RARs) and retinoid X receptors (RXRs). Although vitamin A is essential for the normal development and maintenance of the ocular surface, the RARs and RXRs have not been studied in cornea and conjunctiva. The purpose of this study was to identify the mRNA for these receptors in corneal and conjunctival cells in culture and to determine whether all-trans retinoic acid is able to induce expression of RAR mRNA. METHODS: Total RNA was extracted from cultured rabbit corneal stroma and conjunctival fibroblasts and rabbit corneal epithelial cells. RNA was analyzed by Northern blotting using the cDNA probes for RAR alpha, RAR beta, RAR gamma, RXR alpha, RXR beta and RXR gamma mRNA. To investigate induction of retinoid receptors, cells were exposed to 10(-6) M all-trans retinoic acid for 2-48 h before preparation of RNA. Effects of retinoic acid on cell proliferation were also investigated. RESULTS: RAR alpha mRNA transcripts (3.7 kb), RAR beta mRNA transcripts (3.3 kb) and RAR gamma mRNA transcripts (3.3 kb) are expressed by all the cell types studied, as are the RXR alpha mRNA transcripts (5.0 kb) and RXR beta mRNA transcripts (3.3 kb). RXR gamma mRNA is not detectable in corneal and conjunctival cells. All-trans retinoic acid induced RAR beta mRNA expression in corneal and conjunctival fibroblasts. Increased mRNA levels were detectable after 4-8 h and peaked by 24 h. RAR beta mRNA was not induced by retinoic acid in corneal epithelial cells. Retinoic acid also inhibited proliferation of conjunctival and corneal fibroblasts but had no effect on growth of corneal epithelial cells. CONCLUSIONS: The expression of RARs and RXRs in the cornea and conjunctiva is similar to that reported in other tissues. The identification of these receptors may lead to a better understanding of gene transcription pathways in the cornea and conjunctiva and of the mechanisms that control keratinization, differentiation and proliferation of the cells of these tissues. The data suggest a relationship between the induction of RAR beta mRNA expression and inhibition of cell proliferation by retinoic acid.
PURPOSE: The effects of retinoic acid in cells are mediated by the nuclear retinoic acid receptors (RARs) and retinoid X receptors (RXRs). Although vitamin A is essential for the normal development and maintenance of the ocular surface, the RARs and RXRs have not been studied in cornea and conjunctiva. The purpose of this study was to identify the mRNA for these receptors in corneal and conjunctival cells in culture and to determine whether all-trans retinoic acid is able to induce expression of RAR mRNA. METHODS: Total RNA was extracted from cultured rabbit corneal stroma and conjunctival fibroblasts and rabbit corneal epithelial cells. RNA was analyzed by Northern blotting using the cDNA probes for RAR alpha, RAR beta, RAR gamma, RXR alpha, RXR beta and RXR gamma mRNA. To investigate induction of retinoid receptors, cells were exposed to 10(-6) M all-trans retinoic acid for 2-48 h before preparation of RNA. Effects of retinoic acid on cell proliferation were also investigated. RESULTS:RAR alpha mRNA transcripts (3.7 kb), RAR beta mRNA transcripts (3.3 kb) and RAR gamma mRNA transcripts (3.3 kb) are expressed by all the cell types studied, as are the RXR alpha mRNA transcripts (5.0 kb) and RXR beta mRNA transcripts (3.3 kb). RXR gamma mRNA is not detectable in corneal and conjunctival cells. All-trans retinoic acid induced RAR beta mRNA expression in corneal and conjunctival fibroblasts. Increased mRNA levels were detectable after 4-8 h and peaked by 24 h. RAR beta mRNA was not induced by retinoic acid in corneal epithelial cells. Retinoic acid also inhibited proliferation of conjunctival and corneal fibroblasts but had no effect on growth of corneal epithelial cells. CONCLUSIONS: The expression of RARs and RXRs in the cornea and conjunctiva is similar to that reported in other tissues. The identification of these receptors may lead to a better understanding of gene transcription pathways in the cornea and conjunctiva and of the mechanisms that control keratinization, differentiation and proliferation of the cells of these tissues. The data suggest a relationship between the induction of RAR beta mRNA expression and inhibition of cell proliferation by retinoic acid.