Characterization of heparan sulfate on hepatocytes in regenerating rat liver

Liver regeneration occurs through interactions between the receptors on hepatocytes, including proteoglycans (PGs) and glycosaminoglycans (GAGs), and various growth factors. We investigated serial changes in GAGs, particularly heparan sulfate (HS), in proliferating hepatocytes. We performed 70% hepatectomy in male Wistar rats, and we then isolated hepatocytes by a collagenase perfusion method after each surgery. DNA synthesis was evaluated by measuring proliferating cell nuclear antigen (PCNA). After we had treated the hepatocytes by delipidation and digestion with actinase E, endo-beta-xylosidase, and alpha-amylase, we quantified GAGs by a carbazole-sulfuric acid method. GAGs were analyzed by ion-exchange chromatography, and changes in molecular weight of the HS component were investigated by size-fractionation HPLC. Hepatocyte mitosis peaked at 24 h after the amount of GAGs was increased at 24 and 72 h after surgery. The amount of HS was slightly increased at 3 to 12 h after surgery, and then peaked at 24 h. The molecular weight of the HS declined by 12 h, but had recovered to the preoperative level by 24 h. These results suggested that this HS molecule, which contained about ten disaccharide units during proliferation, may be an initiator of hepatocyte proliferation.