Effects of transforming growth factor-β deficiency on bone development: A Fourier Transform-Infrared imaging analysis

Transforming growth factor-beta 1 (TGF-β1) is a cytokine member of the TGF-β superfamily involved in the control of proliferation and differentiation of various cell types. TGF-β1 plays an important role in bone formation and resorption. To determine the effect of TGF-β1 deficiency on bone mineral and matrix, tibias from mice in which TGF-β1 expression had been ablated (TGF-β1 null) were analyzed and compared with background- and age-matched wild-type (WT) control animals by Fourier transform-infrared imaging (FTIRI) and histochemistry. FTIRI allows the characterization of nondemineralized thin tissue sections at the ultrastructural level with a spatial resolution of 7 μm. The spectroscopic parameters calculated were: mineral-to-matrix ratio (previously shown to correspond to ash weight); mineral crystallinity (related to the crystallographically determined crystallite size and perfection in the apatite c-axis direction); and collagen maturity (related to the ratio of pyridinoline:deH-DHLNL collagen cross-links). Several fields were selected to represent different stages of bone development within the same specimen from the secondary ossification center to the distal diaphysis. Anatomically equivalent areas were compared as a function of age and genotype. The spectroscopic results were expressed both as color-coded images and as pixel population distributions for each of the three parameters monitored. Based on comparisons of histochemistry and FTIRI, there were distinctive age and genotype variations. At all ages examined, in the TGF-β1 null mice growth plates, alkaline phosphatase (ALP) activity and collagen maturity were reduced, but no effect on mineral content or crystallinity was noted. In the TGF-β1 null mice metaphyses, there was a persistence of trabeculae, but no significant alterations in mineral content or crystallinity. In contrast, mineral content, mineral crystallinity, and collagen maturity were reduced in the secondary ossification center and cortical bone of the TGF-β1 null mice. These results, consistent with a mechanism of impaired bone maturation in the TGF-β1 null mice, may be directly related to TGF-β1 deficiency and indirectly to increased expression of inflammatory cytokines in the TGFβ1 null mice.