## 2 The Role(s) of Non-Collagenous Bone Matrix Proteins in Bone
### Effects of Non-Collagenous Proteins on Bone Structure and Mechanics  
Various skeletal disease phenotypes have been associated with a lack of or a mutation in non-collagenous bone matrix proteins, but ascertaining their precise roles in each disease remains a work in progress. Mice are widely used for modeling diseases in man due to certain anatomical, physiological and genetic parallels between the two species, rendering mice models highly effective in recapitulating human disease phenotypes [71]. Since more than 20 years ago, researchers have attempted to isolate the effects of NCPs by using genetically engineered mice with deficiency in a single NCP, i.e., single-knockout models, and compare the resulting bone with that from wild-type (WT) animals. Mice lacking two or more NCPs have also been subsequently investigated [60, 65, 72, 73, 74, 75, 76, 77, 78]. This is done for several reasons: first, studying double- or multiple-knockout animals can help establish whether the effects from single-knockouts are additive, subtractive or neither and whether there exist synergies between two or more NCPs. Secondly, processes in biological systems are almost always interdependent and confounded. A mouse lacking one NCP might and could use another NCP to carry out the same function, and exclusively studying deletions of single NCPs would fail to capture common functional redundancies. Another possibility is that the deficit in one NCP can alter the expression and/or the distribution pattern of other non-collagenous bone matrix proteins [54; 58; 79]; thus, identifying their exact roles would require examining a number of knockout animals. Thirdly, pertinent to studying bone pathologies, a given disease cannot be singly attributed to the absence of one NCP. Studying deficits in _relevant_ pairs of NCPs can thus more closely mimic some pathological conditions, taking, for example, the multiple deficiencies in small leucine-rich proteoglycans (SLRPs) for osteoporosis and osteoarthritis [80]. Table 2 provides an overview of the current knowledge on the deletions of one or more of some NCPs on bone properties. Additionally, associations of these NCPs--such as their mutations, overexpression, and down-expression or deficiencies--to bone diseases in mice models and in man have been summarized. \begin{table}
\begin{tabular}{c c c c c c} \hline \hline
**NCPs and** & **Whole-Bone** & **Ultrastructure** & **Chemistry and** & **Mechanical** & **Related Bone** \\
Out of all NCPs, bone matrix proteins OPN and OC have been subject to more extensive investigations. The first systematic study describing OPN-deficient bone was published in _Nature_ in 1996 [85], and numerous reports have since ensued. Consider OPN-deficient mice, even though the morphological traits of bone at the macroscale remained similar to those of WT bone, ultrastructural and chemical alterations were detected at the nanoscale. Small-angle X-ray scattering analysis confirmed a reduction in the mineral crystal later size and an increase in the mineral misorientation within the mineralized collagen matrix [65; 81]. This was recently corroborated by nanobeam electron diffraction analysis to correlate directly with the disorganized collagen fibrillar structure in OPN\({}^{-/-}\) bone, imaged using TEM (Figure 3a) [36]. In the absence of OPN, the previously mentioned \"glue-like\" mechanism of OPN bridging collagen fibrils, as well as the toughening mechanism by dilatational band formation, would likely be nullified. Together with the disordered texture of mineralized collagen fibrils, OPN-knockout bone possesses impaired mechanics, specifically a reduction in fracture toughness. There is a similar abundance of findings on the chemical changes in both OPN\({}^{-/-}\) and OC\({}^{-/-}\) bone, yet no collagen fibril-level information has been acquired for OC\({}^{-/-}\) bone. Despite this, key insights into how OC-deficiency induces pathological changes in the mineral phase have been plenty (Table 2), validating hypotheses that OC regulates mineralization.