Isolation of growth factors from adult bovine bone

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Abstract
Fetal rat calvariae synthesize transforming growth factor β (TGFβ), β2 microglobulin (β2 m), and insulinlike growth factor I (IGF I), but, except for TGFβ, it is not known if these polypeptides are also present in adult bone tissue. Pulverized bovine bone, extracted with 0.5 N HCl and 4 M guanidine HCl and fractionated by gel filtration, was found to contain several biologically active components when tested for its effects on DNA synthesis in osteoblast-rich cell cultures. TGFβ, β2 m, and IGF I were identified and further purified using high performance liquid chromatography (HPLC). TGFβ, identified by a standard TGFβ bioassay or by immunoreactivity, was purified by μBondapak C18 and μBondapak CN reversed phase HPLC. β2 m, identified by immunoreactivity, required an additional fractionation step on a DEAE-HPLC column for complete purification. IGF I, identified by immunoreactivity, was purified by HPLC using a μBondapak C18 and a DEAE-HPLC column. Purified TGFβ, β2 m, and IGF I migrated as single bands on polyacrylamide gel electrophoresis with respective molecular masses of 24,000, 10,000, and 7,500. In conclusion, adult bone matrix, like fetal bone cultures, contains TGFβ, β2 m, and IGF I and these factors may play a role in adult skeletal remodeling.