Scott A. Lloyd

Epidermal growth factor messenger RNA production in human lacrimal gland.

Experimental models and clinical investigations have suggested that epidermal growth factor (EGF) may have a role in corneal wound healing. It has been identified as a normal component of human tears. Rabbit and mouse lacrimal glands have recently been shown to synthesize EGF messenger RNA (mRNA). The purpose of the present study was to determine whether the human lacrimal gland synthesizes EGF mRNA. Total cellular RNA was isolated from pathologic specimens of normal human lacrimal glands from two individuals. Reverse transcriptase was used to generate complementary DNA (cDNA) using a human EGF-specific mRNA primer. Amplification of EGF-related cDNA sequences was performed with the polymerase chain reaction (PCR) and human EGF-derived up- and downstream primers. The PCR products from both lacrimal glands contained an amplified product of the expected length of approximately 410 base pairs. The PCR-generated fragment was verified as an EGF-related amplification product with Southern blotting using a synthetic oligonucleotide probe derived from the mature coding sequence of EGF. These results conclusively demonstrate that the human lacrimal gland synthesizes EGF and suggest that the lacrimal gland could have a regulatory role in maintaining the ocular surface and possibly regulating corneal wound healing through the secretion of EGF.

Glucocorticoid receptor and interleukin-1 receptor messenger RNA expression in corneal cells.

Interleukin-1 receptor and glucocorticoid receptor messenger ribonucleic acid (RNA) sequences coding for the corresponding proteins were detected in corneal epithelium, stromal fibroblast, and endothelial cells using the polymerase chain reaction and hot blotting. Identification of interleukin-1 receptor mRNA in each of the three major cell types of the cornea suggests that interleukin-1 α has autocrine and/or paracrine roles in the cornea, since previous studies have found that interleukin-1 α mRNA is produced in corneal epithelial, stromal fibroblast, and endothelial cells. Further investigation is needed to determine the functions regulated by the interleukin-1 receptor and glucocorticoid receptor in the cornea and the role of each in corneal wound healing.

Fibroblast growth factor-1 receptor messenger RNA expression in corneal cells.

The polymerase chain reaction (PCR) and hot-blotting methods were used to identify fibroblast growth factor (FGF) receptor- 1-specific messenger ribonucleic acid (mRNA) sequences in cDNA samples prepared from human corneal endothelial cell cultures with proliferative and senescent morphology, an ex vivo corneal epithelium sample, two primary corneal epithelial cell cultures, two third-passage corneal epithelial cell cultures, and two stromal fibroblast cultures. The PCR primers used in this study distinguished mRNAs coding for three aminoterminal motifs (alpha, beta, and gamma) of the FGF receptor- 1 that are derived by alternative splicing from a single genomic sequence. Messenger RNA molecules coding for FGF receptor-1 amino-terminal motif were detected in corneal endothelial and epithelial cells. The alpha and beta amino-terminal motif, but not the gamma amino-terminal motif, mRNAs were detected in stromal fibroblasts. The gamma motif lacks a known signal sequence for membrane translocation and is thought to represent an intracellular form of the FGF receptor-1. Identification of mRNA coding for FGF receptor-1 along with the previous identification of basic FGF mRNA and protein in corneal endothelial, epithelial, and stromal fibroblast cells suggests an autocrine and/or paracrine role for basic FGF in the physiology of the cornea.