Doctor of Philosophy
Chemical and Life Science Engineering
Therapeutic Injectable Iron-Chelating Hydrogels for Improved Central Nervous System Regeneration
Debbie S. Campbell-Rance
A dissertation submitted in partial fulfillment of the requirements of the degree of Doctor of Philosophy at Virginia Commonwealth University 2021.
Director: Xuejun Wen, M.D., Ph.D., AIMBE Fellow, Alice T. and William H. Goodwin Jr. Endowed Chair Professor in Regenerative Medicine, Institute for Engineering and Medicine, Department of Chemical and Life Science Engineering
Severe traumatic brain and spinal cord injuries are major global public health and socioeconomic problems in terms of mortality and morbidity. Currently there are two main lines of treatment under development for traumatic brain injury (TBI) and spinal cord injury (SCI): the use of pharmacological agents and stem cell therapy. Both therapies exhibit potential but fail to restore function of the damaged brain and spinal cord. Pharmacotherapy failed because it focused mainly on neuroprotection of the remaining nervous tissues while ignoring the regeneration of damaged tissues. On the other hand, the stem cell therapies still have not generated consistent results because these studies failed to address the hostile microenvironment at the lesion site and the lack of combinatorial strategies since multiple pathways are involved in the pathophysiology of the secondary damages in TBI and SCI.
To facilitate CNS injury repair, it is essential to transplant stem cells in a favorable microenvironment that is conducive for their long-term survival and integration within the host tissue. Attributes such as three dimensionality of the graft and adhesive support for transplanted cells facilitates integration. Neural tissue engineering (NTE) is a promising strategy for overcoming the limitations of using pharmacological agents or cell therapy alone for the treatment of CNS injuries. It includes the practice of combining biomaterial scaffolds, stem cells and bioactive molecules to create functional constructs that replace, regrow, restore, maintain or improve damaged or diseased tissues. Researchers have shown that they can manipulate the microenvironment at the site of TBI and SCI to show signs of tissue regeneration, but not enough to restore full motor and cognitive functions in the impaired. This suggests that an oversight in the treatment strategy might be playing a pivotal role in preventing full functional recovery.
Biochemical inspection at the CNS injury sites indicates that this microenvironment has an unusually high concentration of iron. Iron is an essential nutrient for cellular processes including electron transport and catalysis. Iron’s ability to redox cycle is an important aspect of its essential functions in the body and controlling iron levels in the body is critical since both over- and under-load of iron can cause cellular dysfunction. Experimental and clinical evidence indicate that excess iron is lethal to cells due to its ability to promote free radical production and the ensuing oxidative stress which is mainly produced by Fenton or/and Haber-Weiss reactions. The free radicals produced induce cellular damage via their interactions with proteins, lipids, carbohydrate and deoxyribonucleic acid (DNA).
This research developed an NTE strategy that mitigates against build-up of iron at the injury site and alleviates the associated deleterious events, improving the survival and functionality of the transplanted stem cells. The end goal of this work was to develop a therapeutic injectable iron-chelating hydrogel to deliver stem cells to the injury site. The hydrogel fills the legion site and forms a microenvironment that mimics that of the brain and spinal cord. The hydrogel will enable the stem cells to survive at the injury site and secrete bioactive molecules that will facilitate neuro-regeneration. Selecting a polymeric material for engineering the injectable hydrogel, involved the design of in vitro experiments that investigated the effect of the polymer on the viability and proliferation of human neural stem cells as well as its ability to mitigate against iron (II)/ascorbic acid-induced oxidative stress and lipid peroxidation. Hyaluronic acid (HA) was the chosen polymeric material because of its known neuroprotection in wound healing and rheological properties tunable to mimics the microenvironment of the CNS. In order to explain the neuroprotective effect of the HA on hNSC when oxidative stress was induced in vitro, we performed iron chelation efficiency studies. Further, we reduced the cytotoxicity of three small molecule iron chelators via chemical modification with HA and PEG. This resulted in three families of iron chelator-polymer conjugates (ICCs) for preliminary studies on hNSC and ultimately to be incorporated into the design of the injectable hydrogel. This project will provide a possible solution to overcome the toxicity caused by high iron concentration at the CNS injury sites.
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Available for download on Friday, May 19, 2023