Molecular Mimicries of Glycosaminoglycans Unraveling the Mechanisms of Uptake and Plasticity in Neuronal Cell Types

Abstract

Glycosaminoglycans are negatively-charged polysaccharide compounds, composed of repeating disaccharide units and found in a majority of metazoan and mammalian tissues. The molecular structure of the GAGs and the pattern of sulfation that they carry largely determine their specific function in the body of an organism, which includes vital processes such as regulation of cell proliferation, anti-coagulation, anti-inflammatory response, cellular signaling, synaptic development, and many more. Heparan and Chondroitin Sulfate (HS and CS) are the major components of the proteoglycans in the ECM, including glypican, syndecan, versican, and neurocan. In the CNS, CSPGs bind to Type IIa receptor protein tyrosine phosphatases, CD44, and a wide range of neuronal growth factors (midkine, BDNF, NT-4), resulting in the regulation of neuronal morphology: promoting or inhibiting neuronal outgrowth depending on the context. HSPGs are important components of the synapse-organizing protein complexes and are central to the development and function of neural networks. Due to the varied roles of HS and CS, there is a great deal of interest in synthesizing heterogeneously sulfated analogs of these GAGs and studying their binding preferences and structure-activity relationships in biological systems. The work this thesis aimed to describe the methods undertaken to synthesize basic GAG monosaccharide building blocks, functionalizing GAG oligomers to gold nanoparticles and synthetics peptides for experiments, and the subsequent assays that were performed to gauge their uptake/binding preferences in murine primary neuronal cultures and secondary cell lines. Our study also aimed to characterize the effect of increasing the concentration of an analytically selected GAG oligomer (CS-E) on the cell surface or in the ECM of primary neurons, in order to understand the effects on neurite outgrowth and plasticity elicited by the molecule. Mapping the specific binding and interactions of the HS and CS oligomers and revealing their effect on in-vitro neural plasticity will provide useful insights to develop specialized therapies and potential cellular targeting mechanisms for novel drugs.