Effects of Transforming Growth Factor β1 on Bonelike Tissue Formation in Three-Dimensional Cell Culture. I. Culture Conditions and Tissue Formation

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Abstract
Bone tissue engineering based on growing bone marrow stromal cells on poly(L-lactic-co-glycolic acid) fiber meshes suffers from limited matrix production and mineralization when the cells are cultured with the standard differentiation supplements (dexamethasone, β-glycerophosphate, and ascorbic acid). To overcome this problem we included transforming growth factor β1 (TGF-β1), which is described as playing a key role in collagen type I formation, although its effect on mineralization is controversially discussed. The investigations focused on establishing culture conditions for the application of TGF-β1 in three-dimensional cell culture and on the effects of different doses of TGF-β1 (1–20 ng/mL) on bonelike extracellular matrix formation. Immunohistochemical staining showed that TGF-β1 enhanced the formation of procollagen type I, collagen type I, and collagen type V, especially under dynamic culture conditions (orbital shaker). A long-term study confirmed positive effects on the formation of extracellular matrix, which penetrated the scaffold to a depth of 250 to 300 µm. Mineralization, qualified by scanning electron microscopy in combination with energy-dispersive X-ray analysis and evaluated by determination of the Ca2+ content per scaffold, was up to 1.7-fold increased by TGF-β1 compared with the control. In conclusion, the growth factor TGF-β1 seems to be effective in improving extracellular bonelike matrix formation in vitro.