Investigation of Glycosaminoglycans with Ion Mobility-Mass Spectrometry and Gas-Phase IR Spectroscopy

Glycosaminoglycans (GAGs) are a family of polydisperse polysaccharides, which are widely distributed at cell surfaces and in the extracellular matrix. Although structurally simple at first glance, with a repeating backbone of alternating hexuronic acid and hexosamine dimers, they display a highly complex structure, which results from their heterogeneous sulfation pattern. Even though structurally poorly understood, there has been increasing evidence that GAGs can transchelate gadolinium-based magnetic resonance imaging (MRI) contrast agents. This unintended release of gadolinium is the leading cause of nephrogenic systemic fibrosis. To date, however, the lack of structurally well-defined GAG samples has hindered a detailed elucidation of the underlying mechanism and only soft evidence of a GAG-induced gadolinium release has been reported. In this work we demonstrate the purification and isolation of GAG fragments from low- molecular-weight-heparin (LMWH) enoxaparin using size-exclusion chromatography. Additionally, we provide the first direct evidence for GAG-gadolinium binding using the synthetic model substance fondaparinux and LMWH enoxaparin. Gas-phase IR spectroscopy and CID-MS/MS experiments revealed possible binding sites of gadolinium on fondaparinux, with surprisingly strong binding contribution from hexuronic acid site chains. This data represent the first direct insights into this complex interaction and will in the future help to unravel the molecular details of GAG-induced gadolinium transchelation from MRI contrast agents

Unveiling Glycerolipid Fragmentation by Cryogenic Infrared Spectroscopy

Mass spectrometry is routinely employed for structure elucidation of molecules. Structural information can be retrieved from intact molecular ions by fragmentation; however, the interpretation of fragment spectra is often hampered by poor understanding of the underlying dissociation mechanisms. For example, neutral headgroup loss from protonated glycerolipids has been postulated to proceed via an intramolecular ring closure but the mechanism and resulting ring size have never been experimentally confirmed. Here we use cryogenic gas-phase infrared (IR) spectroscopy in combination with computational chemistry to unravel the structures of fragment ions and thereby shed light on elusive dissociation mechanisms. Using the example of glycerolipid fragmentation, we study the formation of protonated five-membered dioxolane and six-membered dioxane rings and show that dioxolane rings are predominant throughout different glycerolipid classes and fragmentation channels. For comparison, pure dioxolane and dioxane ions were generated from tailor-made dehydroxyl derivatives inspired by natural 1,2- and 1,3-diacylglycerols and subsequently interrogated using IR spectroscopy. Furthermore, the cyclic structure of an intermediate fragment occurring in the phosphatidylcholine fragmentation pathway was spectroscopically confirmed. Overall, the results contribute substantially to the understanding of glycerolipid fragmentation and showcase the value of vibrational ion spectroscopy to mechanistically elucidate crucial fragmentation pathways in lipidomics

Vertical orientation correction of uav image-based point clouds using statistical modeling of gable roof geometry

4th ISPRS Geospatial Week 2019, Netherlands, 10-14 June 2019201908 bcmaVersion of RecordPublishe

Marker-free coregistration of UAV and backpack LiDAR point clouds in forested areas

202305 bckwAccepted ManuscriptSelf-fundedPublishe

The pattern recognition receptor CD36 is a chondrocyte hypertrophy marker associated with suppression of catabolic responses and promotion of repair responses to inflammatory stimuli

Multiple inflammatory mediators in osteoarthritis (OA) cartilage, including S100/calgranulin ligands of receptor for advanced glycation end products (RAGE), promote chondrocyte hypertrophy, a differentiation state associated with matrix catabolism. In this study, we observed that RAGE knockout was not chondroprotective in instability-induced knee OA in 8-wk-old mice. Hence, we tested the hypothesis that expression of the alternative S100/calgranulin and patterning receptor CD36, identified here as a marker of growth plate chondrocyte hypertrophy, mediates chondrocyte inflammatory and differentiation responses that promote OA. In rat knee joint destabilization-induced OA, RAGE expression was initially sparse throughout cartilage but increased diffusely by 4 wk after surgery. In contrast, CD36 expression focally increased at sites of cartilage injury and colocalized with developing chondrocyte hypertrophy and aggrecan cleavage NITEGE neoepitope formation. However, CD36 transfection in normal human knee-immortalized chondrocytes (CH-8 cells) was associated with decreased capacity of S100A11 and TNF-α to induce chondrocyte hypertrophy and ADAMTS-4 and matrix metalloproteinase 13 expression. S100A11 lost the capacity to inhibit proteoglycans synthesis and gained the capacity to induce proteoglycan synthesis in CD36-transfected CH-8 cells. Moreover, S100A11 required the p38 MAPK pathway kinase MKK3 to induce NITEGE development in mouse articular cartilage explants. However, CH-8 cells transfected with CD36 demonstrated decreased S100A11-induced MKK3 and p38 phosphorylation. Therefore, RAGE and CD36 patterning receptor expression were linked with opposing effects on inflammatory, procatabolic responses to S100A11 and TNF-α in chondrocytes

Ion mobility-tandem mass spectrometry of mucin-type O-glycans

The dense O-glycosylation of mucins plays an important role in the defensive properties of the mucus hydrogel. Aberrant glycosylation is often correlated with inflammation and pathology such as COPD, cancer, and Crohn's disease. The inherent complexity of glycans and the diversity in the O-core structure constitute fundamental challenges for the analysis of mucin-type O-glycans. Due to coexistence of multiple isomers, multidimensional workflows such as LC-MS are required. To separate the highly polar carbohydrates, porous graphitized carbon is often used as a stationary phase. However, LC-MS workflows are time-consuming and lack reproducibility. Here we present a rapid alternative for separating and identifying O-glycans released from mucins based on trapped ion mobility mass spectrometry. Compared to established LC-MS, the acquisition time is reduced from an hour to two minutes. To test the validity, the developed workflow was applied to sputum samples from cystic fibrosis patients to map O-glycosylation features associated with disease