ALBERT

Description: Skeletal unloading induces loss of bone mineral density in weight-bearing bones. The objectives of this study were to characterize the post-translational modifications of collagen of weight-bearing bones subjected to hindlimb unloading for 8 weeks. In unloaded bones, tibiae and femurs, while the overall amino acid composition was essentially identical in the unloaded and control tibiae and femurs, the collagen cross-link profile showed significant differences. Two major reducible cross-links (analyzed as dihydroxylysinonorleucine and hydroxylysinonorleucine) were increased in the unloaded bones. In addition, the ratios of the former to the latter as well as pyridinoline to deoxypyridinoline were significantly decreased in the unloaded bones indicating a difference in the extent of lysine hydroxylation at the cross-linking sites between these two groups. These results indicate that upon skeletal unloading the relative pool of newly synthesized collagen is increased and it is post-translationally altered. The alteration could be associated with impaired osteoblastic differentiation induced by skeletal unloading that results in a mineralization defect.

Description: The osteopetrotic mib mutation in rats causes defects in the skeletal bone tissue in young animals. These defects, i.e. slow bone remodelling, changes in both crystallinity and mineral content, are transient and undergo normalization, even without any treatment in 6-wk-old animals. Treatment with CSF-1 (colony stimulating factor-1) accelerates the normalization process in skeletal bones. The periodontal tissues around the apices of incisors show abnormalities caused by the slow remodelling process of the mandible bone tissue, the deficiency of osteoclasts and their abnormal morphology, as well as the disorganization of periodontal ligament fibres. In contrast to the skeletal tissues, these abnormalities would not undergo spontaneous normalization. Under treatment with colony stimulating factor 1 (CSF-1), the primitive bone trabeculae of mandible are resorbed and the normalization of the number of osteoclasts and their cytology occurs. The organization of the periodontal ligament fibres is partially restored, resembling the histological structure of the normal one.

Description: The mechanical integrity of bone is dependent on the bone matrix, which is believed to account for the plastic deformation of the tissue, and the mineral, which is believed to account for the elastic deformation. The validity of this model is shown in this study based on analysis of the bones of vitamin B6-deficient and vitamin B6-replete chick bones. In this model, when B6-deficient and control animals are compared, vitamin B6 deficiency has no effect on the mineral content or composition of cortical bone as measured by ash weight (63 +/- 6 vs. 58 +/- 3); mineral to matrix ratio of the FTIR spectra (4.2 +/- 0.6 vs. 4.5 +/- 0.2), line-broadening analyses of the X-ray diffraction 002 peak (beta 002 = 0.50 +/- 0.1 vs. 0.49 +/- 0.01), or other features of the infrared spectra. In contrast, collagen was significantly more extractable from vitamin B6-deficient chick bones (20 +/- 2% of total hydroxyproline extracted vs. 10 +/- 3% p 〈 or = 0.001). The B6-deficient bones also contained an increased amount of the reducible cross-links DHLNL, dehydro-dihydroxylysinonorleucine, (1.03 +/- 0.07 vs. 0.84 +/- 0.13 p 〈 or = 0.001); and a nonsignificant increase in HLNL, dehydro-hydroxylysinonorleucine, (0.51 +/- 0.03 vs. 0.43 +/- 0.03, p 〈 or = 0.10). There were no significant changes in bone length, bone diameter, or area moment of inertia. In four-point bending, no significant changes in elastic modulus, stiffness, offset yield deflection, or fracture deflection were detected. However, fracture load in the B6-deficient animals was decreased from 203 +/- 35 MPa to 151 +/- 23 MPa, p 〈 or = 0.01, and offset yield load was decreased from 165 +/- 9 MPa to 125 +/- 14 MPa, p 〈 or = 0.05. Since earlier histomorphometric studies had demonstrated that the B6-deficient bones were osteopenic, these data suggest that although proper cortical bone mineralization occurred, the alterations of the collagen resulted in changes to bone mechanical performance.

Description: Skeletal adaptability to mechanical loads is well known since the last century. Disuse osteopenia due to the microgravity environment is one of the major concerns for space travelers. Several studies have indicated that a retardation of the mineralization process and a delay in matrix maturation occur during the space flight. Mineralizing fibrillar type I collagen possesses distinct cross-linking chemistries and their dynamic changes during mineralization correlate well with its function as a mineral organizer. Our previous studies suggested that a certain group of matrix proteoglycans in bone play an inhibitory role in the mineralization process through their interaction with collagen. Based on these studies, we hypothesized that the altered mineralization during spaceflight is due in part to changes in matrix components secreted by cells in response to microgravity. In this study, we employed hindlimb elevation (tail suspension) rat model to study the effects of skeletal unloading on matrix proteoglycans in bone.

Description: A common feature of various types of mammalian osteopetroses is a marked increase in bone mass accompanied by spontaneous bone fractures. The toothless (tl/tl) rat osteopetrotic mutation is characterized by drastically reduced bone resorption due to a profound deficiency of osteoclasts and their precursors. An altered bone morphology has also been observed. The mutants cannot be cured by bone marrow transplantation, but skeletal defects are greatly reduced after treatment with colony stimulating factor 1 (CSF-1). The objectives of this study were to characterize mineral and collagen matrices in cancellous and compact bone isolated from long bones of 6-week-old normal littermates, tl/tl osteopetrotic mutants and mutants (tl/tl) treated with CSF-1. There were no differences in bone mineral content, but a significant decrease in the crystallinity of mineral evaluated by the method based on electron paramagnetic resonance spectrometry was observed in all bones of tl/tl mutants as compared to that of controls. Within the collagen matrix, slight decreases in the labile cross-links, but significant increases in the content of the stable cross-links, pyridinoline, and deoxypyridinoline, were observed in both cancellous and compact bone of osteopetrotic mutants. In tl/tl mutants treated with human recombinant CSF-1, the normalization of the crystallinity of bone mineral as well as collagen cross-links was found. Our results indicate that remodeling of bone matrix in tl/tl mutants is highly suppressed, but that after treatment with CSF-1, this activity recovers significantly. Taken together, these data provide further support for the hypothesis that CSF-1 is an essential factor for normal osteoclast differentiation and bone remodelling.

Description: A calcium ion precipitable, trypsin-generated proteoglycan fragment has been isolated from the demineralized, EDTA-insoluble matrices of bone. The demineralized matrix was completely digested with trypsin, increasing concentrations of CaCl2 were added to the supernatant, and the resulting precipitates were analyzed. The amount of precipitate gradually increased with higher concentrations of calcium and was reversibly solubilized by EDTA. After molecular sieve and anion exchange chromatography, a proteoglycan-containing peak was obtained. Immunochemical analysis showed that this peak contained chondroitin 4-sulfate and possibly keratan sulfate. Amino acid analysis showed that this proteoglycan contained high amounts of aspartic acid/asparagine (Asx), serine (Ser), glutamic acid/glutamine (Glx), proline (Pro), and glycine (Gly); however, it contained little leucine (Leu) which suggests that it is not a member of the leucine-rich small proteoglycan family. In addition, significant amounts of phosphoserine (P-Ser) and hydroxyproline (Hyp) were identified in hydrolysates of this fraction. A single band (M(r) 59 kDa) was obtained on SDS-PAGE that stained with Stains-all but not with Coomassie Brilliant Blue R-250. If bone powder was trypsinized prior to demineralization, this proteoglycan-containing fraction was not liberated. Collectively, these results indicate that a proteoglycan occurs in the demineralized matrix that is precipitated with CaCl2 and is closely associated with both mineral and collagen matrices. Such a molecule might facilitate the structural network for the induction of mineralization in bone.