Synthesis of chondroitin sulfate by fibrotic vitreous induced by monocytes and lymphocytes

Vitreous fibrosis was induced in rabbit eyes by intravitreal injection of monocytes and lymphocytes. The fibrotic vitreous and normal vitreous removed from experimental animals were then incubated with 3H]-glucosamine at 37 degrees C for 24 hr. The newly synthesized 3H-labeled glycosaminoglycans (GAGs) were isolated by 4 M GuHCl extraction followed by pronase digestion. The 3H-labeled GAGs were then characterized by gel-filtration column chromatography and by specific enzymatic degradation, i.e. hyaluronidase, chondroitinase AC and/or chondroitinase ABC. The disaccahrides derived from chondroitinase ABC degradation were identified by thin-layer chromatography. Our results indicated that 91% of the total glycosaminoglycan synthesized by normal vitreous was hyaluronic acid. In contrast, in the fibrotic vitreous, the synthesis of hyaluronic acid was decreased to 30% whereas the synthesis of chondroitin sulfate increased to 47% of the total newly synthesized glycosaminoglycans. Control vitreous which was injected with freeze-thawed monocytes and lymphocytes synthesized 70% hyaluronic acid and 12% chondroitin sulfate although no fibrosis was observed.