The central nervous system (CNS) is composed of the brain, optic nerve and spinal cord and is responsible for most of the body’s functions and processing external environmental information. Damage to the CNS can develop in different pathological conditions ranging from infection, traumatic injury of the spinal cord (SCI), traumatic brain injury (TBI), and degenerative disorders such as multiple sclerosis (MS). Neural degeneration and demyelination of axons are hallmarks of CNS injury. Demyelination in the CNS occurs due to a variety of pathophysiological conditions, therefore, any repair strategies for demyelination must consider multifactorial pathways including promotion of axonal outgrowth, and remyelination.
Previously, we have demonstrated that low sulphated modified heparin mimetics (LSmHeps) enhance neurite outgrowth and myelination in vitro. Heparin mimetics (mHeps) are a class of glycomolecules with structural similarities to resident heparan sulphates (HS) and are made up of repeating disaccharide units with variable sulphation groups. They are thought to modulate cell signalling by both sequestering ligands and acting as a cofactor in the formation of ligand-receptor complexes. Thus, LS-mHeps have the capacity to represent novel candidates as therapeutics for CNS damage. However, a major hurdle for CNS therapeutics is for molecules and compounds to cross the blood brain barrier (BBB). Large molecular weight is known to prevent molecules crossing the BBB; therefore, we have developed a low molecular weight form of our lead compound LS-mHep7.
This thesis aimed to validate the ability of this low molecular weight form (LS-mHep7L) to maintain the ability to enhance repair in several CNS injury models including in vitro myelinating cultures, during both myelin development (MC-Dev) and demyelination (MCDeMy), and astrocyte injury assays. Additionally, this thesis aimed to optimise an ex vivo slice culture model to further validate LS-mHeps and found that spinal cords from C57BL/6 P1 mice produced healthy myelinating axons for remyelination studies.
It was found that LS-mHep7L enhanced neurite outgrowth in vitro and remyelination both in vitro and ex vivo. LS-mHep7L was found to sequester CCL5 – a negative regulator of myelination both in vitro and ex vivo, and restored CCL5 induced hypomyelination in developing cultures. LS-mHeps also reduced signs of reactive astrocytes with a decrease in nestin expression and appeared to enhance gap closure in the injury model.
Finally, we investigated the use of recombinant heparin mimetics (rHS) as an alternative source for heparin derived therapeutics. Currently 80% of the world’s heparin supply is sourced from China from porcine intestines and having such a reliance on a specific animal source for any new therapeutic comes with an elevated risk. Here we demonstrated that low sulphated recombinant heparan sulphate (rHS10) enhance remyelination in MC-DeMy and SC-DeMy, while rHS09 enhance neurite outgrowth in MC-Inj.
In summary, the results of this thesis demonstrated that low sulphated heparin mimetics have the potential to become novel therapeutics for remyelination and neurite outgrowth for diseases and injury of the CNS. Additionally, low molecular weight LS-mHeps show the same bioactivity as the high molecular weight form, by demonstrating neurite outgrowth in vitro, enhanced remyelination and sequestration of negative regulators of repair both in vitro and ex vivo cultures
