RECOMBINANT VGR-1 BMP-6 EXPRESSING TUMORS INDUCE FIBROSIS AND ENDOCHONDRAL BONE-FORMATION IN-VIVO

Members of the TGF-beta superfamily appear to modulate mesenchymal dif ferentiation, including the processes of cartilage and bone formation. Nothing is yet known about the function of the TGF beta-related facto r vgr-1, also called bone morphogenetic protein-6 (BMP-6), and only li mited studies have been conducted on the most closely related factors BMP-5, osteogenic protein-1 (OP-1) or BMP-7, and OP-2. Because vgr-1 m RNA has been localized in hypertrophic cartilage, this factor may play a vital role in endochondral bone formation. We developed antibodies to vgr-1, and documented that vgr-1 protein was expressed in hypertrop hic cartilage of mice. To further characterize the role of this protei n in bone differentiation, we generated CHO cells that overexpressed r ecombinant murine vgr-1 protein. Western blot analysis documented that recombinant vgr-1 protein was secreted into the media and was proteol ytically processed to yield the mature vgr-1 molecule. To assess the b iological activity of recombinant vgr-1 in vive, we introduced the vgr -1-expressing CHO cells directly into the subcutaneous tissue of athym ic nude mice. CHO-vgr-1 cells produced localized tumors, and the conti nuous secretion of vgr-1 resulted in tumors with a strikingly differen t gross and histological appearance as compared to the parental CHO ce lls. The tumors of control CHO cells were hemorrhagic, necrotic, and f riable, whereas the CHO-vgr-1 tumors were dense, firm, and fibrotic. I n contrast with control CHO tumors, the nests of CHO-vgr-1 tumor cells were surrounded by extensive connective tissue, which contained large regions of cartilage and bone. Further analysis indicated that secret ion of vgr-1 from the transfected CHO tumor cells induced the surround ing host mesenchymal cells to develop along the endochondral bone path way. These findings suggest that endogenous vgr-1 acts as an osteoindu ctive factor during endochondral bone formation.