title: Assessment of collagen quality associated with non-enzymatic cross-links in human bone using Fourier-transform infrared imaging.
abstract: Aging and many disease conditions, most notably diabetes, are associated with the accumulation of non-enzymatic cross-links in the bone matrix. The non-enzymatic cross-links, also known as advanced glycation end products (AGEs), occur at the collagen tissue level, where they are associated with reduced plasticity and increased fracture risk. In this study, Fourier-transform infrared (FTIR) imaging was used to detect spectroscopic changes associated with the formation of non-enzymatic cross-links in human bone collagen. Here, the non-enzymatic cross-link profile was investigated in one cohort with an in vitro ribose treatment as well as another cohort with an in vivo bisphosphonate treatment. With FTIR imaging, the two-dimensional (2D) spatial distribution of collagen quality associated with non-enzymatic cross-links was measured through the area ratio of the 1678/1692cm-1subbands within the amide I peak, termed the non-enzymatic crosslink-ratio (NE-xLR). The NE-xLR increased by 35% in the ribation treatment group in comparison to controls (p<0.005), with interstitial bone tissue being more susceptible to the formation of non-enzymatic cross-links. Ultra high-performance liquid chromatography, fluorescence microscopy, and fluorometric assay confirm a correlation between the non-enzymatic cross-link content and the NE-xLR ratio in the control and ribated groups. High resolution FTIR imaging of the 2D bone microstructure revealed enhanced accumulation of non-enzymatic cross-links in bone regions with higher tissue age (i.e., interstitial bone). This non-enzymatic cross-link ratio (NE-xLR) enables researchers to study not only the overall content of AGEs in the bone but also its spatial distribution, which varies with skeletal aging and diabetes mellitus and provides an additional measure of bone's propensity to fracture.
authors.affiliation: Department of Osteology and Biomechanics, University Medical Center, 22529 Hamburg, Germany. Electronic address: fel.schmidt@uke.de.
authors.forename: F N
authors.initials: FN
authors.lastname: Schmidt
authors.affiliation: Department of Osteology and Biomechanics, University Medical Center, 22529 Hamburg, Germany. Electronic address: e.zimmermann@uke.de.
authors.forename: E A
authors.initials: EA
authors.lastname: Zimmermann
authors.affiliation: Institute of Biomechanics, Hamburg University of Technology, 21073 Hamburg, Germany. Electronic address: graeme.campbell@tuhh.de.
authors.forename: G M
authors.initials: GM
authors.lastname: Campbell
authors.affiliation: Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA. Electronic address: srogag@rpi.edu.
authors.forename: G E
authors.initials: GE
authors.lastname: Sroga
authors.affiliation: Department of Forensic Medicine, University Medical Center, 22529 Hamburg, Germany. Electronic address: pueschel@uke.de.
authors.forename: K
authors.initials: K
authors.lastname: Püschel
authors.affiliation: Department of Osteology and Biomechanics, University Medical Center, 22529 Hamburg, Germany. Electronic address: amling@uke.de.
authors.forename: M
authors.initials: M
authors.lastname: Amling
authors.affiliation: Department of Orthopaedics, Washington University in St. Louis, St. Louis, MO, USA. Electronic address: tangs@wudosis.wustl.edu.
authors.forename: S Y
authors.initials: SY
authors.lastname: Tang
authors.affiliation: Department of Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA. Electronic address: vashid@rpi.edu.
authors.forename: D
authors.initials: D
authors.lastname: Vashishth
authors.affiliation: Department of Osteology and Biomechanics, University Medical Center, 22529 Hamburg, Germany. Electronic address: b.busse@uke.uni-hamburg.de.
authors.forename: B
authors.initials: B
authors.lastname: Busse
mesh_terms: D000293:Adolescent; D001120:Arginine / Q000031:analogs & derivatives / Q000378:metabolism; D001842:Bone and Bones / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D003094:Collagen / Q000378:metabolism*; D003432:Cross-Linking Reagents / Q000378:metabolism*; D004164:Diphosphonates / Q000494:pharmacology / Q000627:therapeutic use; D006801:Humans; D008239:Lysine / Q000031:analogs & derivatives / Q000378:metabolism; D008875:Middle Aged; D010024:Osteoporosis / Q000188:drug therapy / Q000473:pathology; D012266:Ribose / Q000494:pharmacology; D017550:Spectroscopy, Fourier Transform Infrared
chemical_list: D003432:Cross-Linking Reagents; D004164:Diphosphonates; D012266:Ribose; D003094:Collagen; D001120:Arginine; C062187:pentosidine; D008239:Lysine
keywords: Advanced glycation end products; Bisphosphonates; Bone quality; Collagen; Fourier transform infrared spectroscopy; Non-enzymatic cross-links