Basement membrane proteoglycan in various tissues: characterization using monoclonal antibodies to the Engelbreth-Holm-Swarm mouse tumor low density heparan sulfate proteoglycan

Abstract

The Engelbreth-Holm-Swarm mouse tumor has been found to produce at least two molecular species of heparan sulfate proteoglycan, a low density one (LD) and a high density one, which differ not only in core proteins but also in glycosaminoglycan structures (Kato, M., Y. Koike, Y. Ito, S. Suzuki, and K. Kimata. 1987. J. Biol. Chem. 262:7180-7188). With aim at investigating their distribution and possible functions in tissues, monoclonal antibodies were produced. Hybridomas obtained by fusion of NS-1 mouse myeloma cells with spleen cells from the rat immunized with a mixture of these proteoglycans were selected by their ability to react with the antigen. Two of them secreted monoclonal antibodies (IgG2a), designated HK-84 and HK-102, that recognize specifically the core protein moiety of LD. Immunofluorescent staining of various tissues (skeletal muscle, cardiac muscle, lung, brain, and kidney) with these monoclonal antibodies has demonstrated that the antigen molecules were present in all basement membranes of these tissues. SDS-PAGE of heparitinase-treated proteoglycan fractions prepared from these tissues and subsequent immunoblotting using these monoclonal antibodies have confirmed that the antigen molecule was LD, and further suggested that there was a tissue-specific variation in the core molecular size. Based on these results, we propose that LD may be an essential component in all basement membranes.