Lathyrism-induced alterations in collagen cross-links influence the mechanical properties of bone material without affecting the mineral

Abstract

In the present study a rat animal model of lathyrism was employed to decipher whether anatomically confined alterations in collagen cross-links are sufficient to influence the mechanical properties of whole bone.Animal experiments were performed under an ethics committee approved protocol. Sixty-four female (47 day old) rats of equivalent weights were divided into four groups (16 per group): Controls were fed a semi-synthetic diet containing 0.6% calcium and 0.6% phosphorus for 2 or 4 weeks and β-APN treated animals were fed additionally with β-aminopropionitrile (0.1% dry weight). At the end of this period the rats in the four groups were sacrificed, and L2–L6 vertebra were collected. Collagen cross-links were determined by both biochemical and spectroscopic (Fourier transform infrared imaging (FTIRI)) analyses. Mineral content and distribution (BMDD) were determined by quantitative backscattered electron imaging (qBEI), and mineral maturity/crystallinity by FTIRI techniques. Micro-CT was used to describe the architectural properties. Mechanical performance of whole bone as well as of bone matrix material was tested by vertebral compression tests and by nano-indentation, respectively.The data of the present study indicate that β-APN treatment changed whole vertebra properties compared to non-treated rats, including collagen cross-links pattern, trabecular bone volume to tissue ratio and trabecular thickness, which were all decreased (p < 0.05). Further, compression tests revealed a significant negative impact of β-APN treatment on maximal force to failure and energy to failure, while stiffness was not influenced. Bone mineral density distribution (BMDD) was not altered either. At the material level, β-APN treated rats exhibited increased Pyd/Divalent cross-link ratios in areas confined to a newly formed bone. Moreover, nano-indentation experiments showed that the E-modulus and hardness were reduced only in newly formed bone areas under the influence of β-APN, despite a similar mineral content.In conclusion the results emphasize the pivotal role of collagen cross-links in the determination of bone quality and mechanical integrity. However, in this rat animal model of lathyrism, the coupled alterations of tissue structural properties make it difficult to weigh the contribution of the anatomically confined material changes to the overall mechanical performance of whole bone. Interestingly, the collagen cross-link ratio in bone forming areas had the same profile as seen in actively bone forming trabecular surfaces in human iliac crest biopsies of osteoporotic patients.

Effects of alendronate and risedronate on bone material properties in actively forming trabecular bone surfaces

“…(38,57) It has recently been shown in animals that alterations in collagen cross-links (most notably the pyr/divalent cross-link ratio), coupled with changes in microarchitecture, are sufficient to adversely affect bone biomechanical performance in the absence of any concomitant changes in mineral quantity and quality, even when these changes are anatomically confined to bone-forming surfaces. (61) Both shorter-term and longer-term RIS use resulted in significantly lower pyr/divalent collagen cross-link ratios compared to ALN, in agreement with a recent publication. (62) It is also in general agreement with previously published reports showing that although RIS administration in postmenopausal women lowered this bone quality index compared to placebo, ALN did not.…”

“…Although these differences may be anatomically restricted to bone-forming surfaces, recent data from animals indicated that such changes are still sufficient to influence bone strength even with no change in the amount and quality of mineral. (61) A limitation of the present study is that end-point biopsies were collected from the patients in the general population, thus compliance with therapy is uncertain. Another limitation is the low sample number of biopsies from patients on RIS for >5 years compared to the other groups.…”

Exercise prevents β-aminopropionitrile-induced morphological changes to type I collagen in murine bone

“…Although the specific mechanisms responsible are not clear, changes in crystallinity because of BAPN treatment is an interesting finding that warrants further investigation using more direct methods of evaluating changes to the crystalline structure. 27 Another study demonstrated no effects on crystallinity/maturity with BAPN treatment, 13 but these conflicting data may be due to differences in dose (B780 vs 300 mg kg À 1 ), method of delivery (dietary vs subcutaneous injection), animal model (rat vs mouse) and anatomical site (vertebral crosssection vs native surface of femur).…”

“…9 The sweet pea (Lathyrus odoratus) contains b-aminopropionitrile (BAPN), 10 which irreversibly binds to the LOX active site, 11 and is a commonly used lathyrogen to experimentally replicate osteolathyrism. [12][13][14][15] Understanding the implications of reduced enzymatic crosslinking is important because crosslinks are reduced with aging and other diseases such as diabetes and osteoporosis. 16 Therefore, treatment with BAPN can serve as a model of osteolathyrism and further our understanding of how nanoscale changes in collagen impact bone strength and fracture resistance.…”

The contribution of collagen crosslinks to bone strength

“…Using the same animal model, it was recently reported that beta aminoproprionitrile treatment induced a decrease in divalent and trivalent crosslinks, which was associated with a decrease of trabecular thickness and compressive energy to failure, while stiffness was not affected. 32 Another rat study showed that increased homocysteinemia induced by a diet enriched in methiononine or homocysteine -which is likely to interfere with crosslink formation -for 3 months resulted in decreased bone formation, bone volume and bone strength at the distal femur. 33 The impact of an increase of enzymatic crosslinks on bone strength can also be investigated using fetal bovine cortical bone, which is characterized by a low level of these molecules.…”