Synthesis of Sulfated Glycosaminoglycan in Newt Iris during Lens Regeneration

Abstract

Lens regeneration in newts involves the dedifferentiation of pigmented iris epithelial cells and their conversion into lens fibers at the dorsal pupillary margin. We have investigated the pattern of incorporation of 35SO 4 into sulfated glycosaminoglycan (GAG) within the iris during this process. Newts were injected with [ 35S]H 2SO 4 at varying intervals after lens removal. Iris and lens tissues were excised and the accumulation of label into total sulfated GAG was measured from the radioactivity incorporated in the ethanol-insoluble, cetylpyridinium chloride-precipitable fractions of pooled tissue homogenates. The proportions of label in GAG of the chondroitin sulfate, dermatan sulfate, and heparin/heparan sulfate types were determined from the reduction of label after chondroitinase AC, chondroitinase ABC, and nitrous acid treatments. Compared with the normal iris of the unoperated newt, uptake of label into sulfated GAG was increased dramatically after lens removal. Labelling rose to a peak at 15 days after lentectomy and declined slightly from its maximum by day 30. At all times heparin/heparan sulfate accounted for approximately 60% of total label uptake while chondroitin sulfate and dermatan sulfate each contributed 15–25%. Incorporation of 35SO 4 was nearly equivalent in dorsal (lens-forming) and ventral (non-lens-forming) halves of the iris at all stages examined. Only negligible quantities of label were associated with either mature (normal) or developing (30 day) lenses. It is concluded that lentectomy stimulates an elevation in the total rate of incorporation of 35SO 4 into newly synthesized sulfated GAG within the newt iris, while preserving the relative labelling rates of the different types of GAG produced. We suggest that an enhanced synthesis of GAG may be associated with the dedifferentiation of iris epithelial cells following lentectomy and the retrieval pathway of redifferentiation into pigment cells.