Developmental expression of mucin genes ASGP (rMuc4) and rMuc5ac by the rat ocular surface epithelium

PURPOSE. To determine site and time of initiation of expression of the memb rane-spanning mucin ASGP (rMuc4) and the goblet cell-specific, gel-forming mucin rMuc5AC by the developing rat ocular surface epithelium. METHODS. Newborn Sprague-Dawley rat pups were killed at 1, 7, and 14 days a fter birth. Adult rats (weight, 200 g) were used as controls. Reverse trans cription-polymerase chain reaction (RT-PCR) was performed to detect ASGP mR NA using beta-actin as an internal control. Competitive RT-PCR was performe d to quantitate rMuc5AC: mRNA using an rMuc5AC-competitive reference standa rd (CRS) as an internal control. In situ hybridization was performed to loc alize ASGP and rMuc5AC mRNA. Goblet cells were detected by staining with pe riodic acid-Schiff (PAS) reagent. RESULTS. ASGP mRNA was detected by RT-PCR at 1 day after birth. Compared wi th beta-actin, the amount of ASGP mRNA should a progressive increase from 1 to 14 days of postnatal development. By in situ hybridization, the express ion of ASGP was first clearly detected at 14 days after birth at the lid ma rgin, where the most stratification of epithelium was seen, and along the a djacent palpebral conjunctiva. This pattern was seen in rat eyelids that we re not yet open but appeared about to open. In rat eyelids already open at 14 days after birth, ASGP mRNA was difficulty spread in the apical cell lay er of both conjunctival and corneal epithelia. The expression of rMuc5AC wa s detected by RT-PCR in ocular surface epithelium in rat pups 1 day after b irth. Quantitative RT-PCR showed a low level of rMuc5AC RNA expression in c onjunctiva of 1-, 7-, and 14-day-old rats followed by a large increase in e xpression between 14 days and adulthood. The expression of rMuc5AC was firs t detected by in situ hybridization in a few goblet cells at 7 days after b irth. One or two labeled cells were present in the fornical area: some mere on the palpebral side of the fornix; others were present. on the bulbar si de. The distribution and time of appearance of rMuc5AC correlated with that of PAS staining of goblet cells. CONCLUSIONS. The developmental expression of the membrane-spanning mucin AS GP (rMuc4) and the gel-forming mucin rMuc5AC are regionally and temporally separated. Expression of the gel-forming mucin begins at the fornix at 7 da ys after birth and is correlated with the appearance of goblet cells, where as, expression of the membrane-spanning mucin begins later at the lid margi n at day 14. Expression of the membrane-spanning mucin correlates to eyelid opening.