Phorbol ester preferentially stimulates mouse fornical conjunctival and limbal epithelial cells to proliferate in vivo

PURPOSE. The authors investigated whether fornical epithelium displays a differential in vivo response to acute and chronic stimulation when compared with bulbar and palpebral epithelia. METHODS. To induce an increase in epithelial proliferation, 0.5% phorbol myristate (TPA) was topically applied in petrolatum daily to both eyes of SENCAR mice for 12 days. Control mice (three per group) received petrolatum only. After 6, 12, 18, and 24 hours (acute) and 2, 3, 4, 5, 7, 9, and 12 days (chronic) of TPA treatment, mice (three per group) were administered intraperitoneally 0.1 ml 40 μCi [³H]thymidine ([³H]TdR) 1 hour before they were killed. Conjunctival epithelium was fixed and processed for autoradiography, and the labeling index (LI; number of [³H]TdR-labeled nuclei per 1000 basal keratinocytes) was determined for each of the epithelial zones. RESULTS. Under normal situations, the LI was lowest in fornical epithelium (1.9 ± 0.5) compared with bulbar (4.4 ± 0.9) and palpebral (5.5 ± 0.5) epithelia. Within 24 hours of TPA treatment, a 12-fold increase in fornical basal cell labeling was noted compared with a 2.5- and 5-fold increase in bulbar and palpebral basal cell labeling, respectively. Fornical epithelium maintained a significantly greater proliferative response (4.5-fold increase) during chronic stimulation than either bulbar or palpebral epithelia (0.5- and 1.5- fold increase, respectively). CONCLUSIONS. The more vigorous response of the fornical epithelium to acute and chronic stimulation is strong evidence that this epithelium has a greater proliferative capacity than the other two epithelia, which is consistent with the authors' hypothesis that conjunctival epithelial stem cells are primarily located in the fornical region.