Over the last 12 years, I have conducted research or participated as a student intern in twelve different neuroscience research laboratories across three countries. Four of these experiences have resulted in five publications, available below:
Neurotrophic factors in combinatorial approaches for spinal cord regeneration
Optimization of adult sensory neuron electroporation to study mechanisms of neurite growth
p75 neurotrophin receptor regulates tissue fibrosis through inhibition of plasminogen activation via a PDE4 cAMP PKA pathway
Photoreceptor rescue and toxicity induced by different calpain inhibitors
Structural and Functional Changes in Regenerating Antennules in the Crayfish Orconectes sanborni
Currently at the Heidelberg University Spinal Cord Injury Center, under the direction of Dr. Armin Blesch and a student of the HBIGS Molecular and Cellular Biology Program, my PhD thesis targets intrinsic neuronal mechanisms in adult mammalian regenerative processes. A brief introduction to my current work follows:
Injuries in the peripheral nervous system (PNS) lead to a complex regenerative program at the molecular and cellular level to restore the function and organization of axons . Regenerating axons respond to chemotropic cues composed of growth-promoting substances, such as extracellular matrix proteins and growth factors that are upregulated along axonal tracks, which support regeneration and the formation of new connections. In contrast, axons in the central nervous system (CNS) regenerate very poorly or not at all due to growth inhibition by extracellular matrix, myelin-based inhibitors [2-4], insufficient growth stimulation [5, 6], and inflammatory responses [7, 8]. These factors in the environment of injured axons are only partly responsible for the limited regenerative capacity, since axons from transplanted neural progenitor cells and embryonic and adult dorsal root ganglion (DRG) neurons grow surprisingly well in an environment host to many regeneration-limiting, negative factors [9-11]. These results suggest that neuron-intrinsic factors equally contribute to the limited regeneration capacity in the CNS.
In response to injuries in the PNS, significant changes in gene expression take place, resulting in effective use of resources for robust growth and repair. Neurons of the CNS fail to elicit such a program and consequently do not regenerate. It therefore seems necessary to both influence the environment and manipulate cell-autonomous processes to induce effective regeneration in the CNS. The aim of my thesis entails identifying mechanisms specifically related to the genetic programs that contribute to the regeneration process and further manipulating these via a gene therapy approach to improve regeneration in the injured adult mammalian spinal cord. The ultimate goal of the work is to build upon our established combinatorial paradigm to increase the number of regenerating axons and the distance of axonal elongation in vivo [12-14].
DRG neurons serve as an excellent model for studying the role of cell-autonomous mechanisms upon injury. To determine which elements of the regeneration program are necessary and sufficient to activate and support regeneration, this model provides a straightforward means of consistently prompting the regenerative course. Sensory neurons, located in the periphery, possess two branches: the branch that innervates a peripheral sensory organ and the central branch, which projects into the spinal cord. Following a peripheral nerve crush lesion, DRG neurons initiate a genetic program that drives axonal regeneration to reinnervate appropriate targets. Similar damage to the central branch of the same cell, however, fails to trigger any such cellular growth and repair . However, if a peripheral conditioning lesion (CL) precedes an injury to the central branch, some growth is observed within the inhibitory environment of the CNS .
All evidence indicate that the CL effect is dependent upon transcription, and further data show that over 1,000 genes change in expression level after a peripheral branch injury, compared to very few after a central branch lesion [17-20]. In addition to regeneration-associated genes, cytoskeletal proteins, neurotrophic factors, receptors, and cell adhesion molecules , many transcription factors are also rapidly upregulated, as recent microarray analyses suggest [19, 20].
In previous studies, we have conducted microarray analyses and compared the results of peripheral sensory ganglia after CNS and peripheral lesions and corticospinal and raphaespinal neurons after central lesions in the rat, reporting numerous changes in the transcriptional machinery. Among the genes with altered gene expression are a number of previously investigated transcription factors with potential importance for axonal regeneration. Transcription factors exhibiting upregulation of more than 75% of the levels in the intact condition were prioritized, as well as those demonstrating prolonged changes over fourteen days following injury. Using an in vitro neurite outgrowth assay, I will assess each factor’s influence on promoting growth, continue to establish the factor’s localization and activation in vivo, and further develop the gene delivery system for adult DRG neurons to propogate the transcriptional changes, allowing for initiation of an intrinsic growth program.
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