A bone-derived mixture of TGF beta-superfamily members forms a more mature vascular network than bFGF or TGF-beta 2 in vivo. 
Clinical trials of therapeutic angiogenesis for the treatment of cardiovascular ischemia have failed to meet the expectations with the use of single growth factors, namely VEGF and bFGF.  We show here that a bovine bone-derived growth factor mixture (GFM) of TGFbetas, BMPs, and no more than 0.1% aFGF can initiate a dose-dependent angiogenic response in subcutaneously implanted Growth Factor Reduced Matrigel plugs that includes abundant smooth muscle actin positive (SMA+) tubes and functional CD31+, red blood cell filled, capillaries.  Tube forming activity of the single factors, recombinant bFGF and bone-derived TGF-beta2, were comparable to GFM, but only the bone-derived factors were able to create a larger fraction of SMA+ tubes than Matrigel alone at an equal dose.  Basic FGF formed a greater number of RBC-filled capillaries within the plugs than GFM or TGF-beta2 at the highest doses, although GFM created RBC-filled capillaries that penetrated deeper into the plugs than bFGF.  However, bFGF showed the greatest number of non-cell-lined, RBC-filled pools, suggestive of vessel rupture, and the largest number of plugs showing signs of fluid accumulation in the form of large, cell-lined clefts in the implants.  TGF-beta2 showed less RBC-filled pools, but a significant number of implants with signs of fluid accumulation.  At high doses of GFM penetration by blood vessels and mesenchymal cells was obstructed by cartilage development within the plugs accompanied by a prominent band of SMA+ granulation tissue with abundant RBC-filled capillaries encapsulating the implants.  Thus, GFM is also capable of dramatically remodeling the vascular system in the interstitial space surrounding the plug.  These results show that GFM is capable of inducing the formation of a more mature vascular system than that formed by the single factors bFGF and TGFbeta-2.  Natural mixtures of TGFbetas, BMPs, and FGFs may have superior clinical utility in therapeutic angiogenesis applications.