Haddad, Oualid

The purpose of this study was to characterize the effect of TIEG1 on the molecular structure of collagen within tail tendon fibers using 3-mo-old female C57BL/6 wild-type (WT) and TIEG1 KO mice. Synchrotron X-ray microdiffraction experiments were carried out on single tendon fibers extracted from the WT and TIEG1 KO dorsal tail tendon. The fibers were scanned in the radial direction, and X-ray patterns were obtained. From these patterns, the meridional direction was analyzed through X-ray intensity profile. In addition, collagen content was investigated using hydroxyproline assays, and qualitative real-time PCR experiments were performed on RNA isolated from fibroblasts to examine specific gene expression changes. The results showed different X-ray diffraction patterns between WT and TIEG1 KO tendon fibers, indicating a disorganization of the collagen structure for the TIEG1 KO compared with WT mice. Furthermore, the analyses of the X-ray intensity profiles exhibited a higher (23 Å) period of collagen for the TIEG1 KO compared with the WT mice. The results of the hydroxyproline assays revealed a significant decrease in the TIEG1 KO compared with WT mice, leading to a decrease in the total amount of collagen present within the TIEG1 KO tendons. Moreover, qualitative real-time PCR results showed differences in the expression profiles of specific genes known to play important roles in tendon fiber development. These data further elucidate the role of TIEG1 on tendon structure and could explain the previous defects in the structure-function relationship found for TIEG1 KO tendon fibers.

Through the development of TGFβ-inducible early gene-1 (TIEG1) knockout (KO) mice, we have demonstrated that TIEG1 plays an important role in osteoblast-mediated bone mineralization, and in bone resistance to mechanical strain. To further investigate the influence of TIEG1 in skeletal maintenance, osteocytes were analyzed by transmission electron microscopy using TIEG1 KO and wild-type mouse femurs at one, three and eight months of age. The results revealed an age-dependent change in osteocyte surface and density, suggesting a role for TIEG1 in osteocyte development. Moreover, there was a decrease in the amount of hypomineralized bone matrix surrounding the osteocytes in TIEG1 KO mice relative to wild-type controls. While little is known about the function or importance of this hypomineralized bone matrix immediately adjacent to osteocytes, this study reveals significant differences in this bone microenvironment and suggests that osteocyte function may be compromised in the absence of TIEG1 expression.