Investigating bone resorption in Atlantic herring fish intermuscular bones with solid-state NMR

Bones are connective tissues mainly made of collagen proteins with calcium phosphate deposits. They undergo constant remodeling, including destroying existing bones tissues (known as bone resorption) and rebuilding new ones. Bone remodeling has been well-described in mammals, but it is not the case in fish. Here, we focused on the mobile phase of the bone vascular system by carefully preserving moisture in adult Atlantic herring intermuscular bones. We detected pore water with high ionic strength and soluble degraded peptides whose ¹H-transverse relaxation times, T₂s, exceed 15 milliseconds. With favorable T₂s, we incorporated a solution state spinlock scheme into the INEPT techniques to unequivocally demonstrate collagen degradation. In addition, we detected a substantial amount of inorganic phosphate in solution with ³¹P-NMR in the considerable background of solid hydroxyapatite calcium phosphate by saturation recovery experiment. It is consistent with the idea that bone resorption degrades bone collagen and releases calcium ions and phosphate ions in the pore water with increased ionic strength. Our report is the first to probe the resorption process in the heterogenous bone microstructure with a rigorous characterization of ¹H and ¹³C relaxation behavior and direct assignments. In addition, we contribute to the fish bones literature by investigating fish bone remodeling using NMR for the first time.