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Title: Contributions of retinoic acid signalling to regenerating neurons
Author: Nǡnescu, Sonia Elena
ISNI:       0000 0004 2745 7625
Awarding Body: University of Aberdeen
Current Institution: University of Aberdeen
Date of Award: 2012
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A potent gene transcription regulator, retinoic acid (RA) is known to influence more than 500 genes, whose functions include regulation of neuronal differentiation and patterning of the developing nervous system. Furthermore, its ability to stimulate neurite outgrowth has been documented in primary neurons such as postnatal cerebellar granule neurons, adult cortical neurons, or embryonic and adult dorsal root ganglia neurons (DRGs) (Corcoran et al. 2000, Wong et al. 2006, Yip et al. 2006, Puttagunta et al. 2011). It has been previously proposed that RA synthesis takes place only in the cytoplasm mediated by three retinaldehyde dehydrogenases (Raldhs) namely Raldh1, Raldh2 and Raldh3. From this cytoplasmic location RA is then transported to the nucleus where RA receptors (RAR) α, β and γ initiate gene transcription. RA is then removed from the nucleus and degraded by microsomal cytochrome 450 (Cyp26) A1, B1 or C1 enzymes. New studies have pointed to additional roles for RARs in the cytoplasm to regulate kinase function. Thus, as well as being a highly potent gene regulator, RA might act on several different levels involving separate mechanisms all converging towards its pro-regenerative abilities. The starting hypothesis for this thesis was that neurons in culture that re-grow dendrites and axons following their removal from their previously interconnected state in the brain activate intrinsic RA signalling mechanisms. Neurons that regenerate in culture following dissection of CNS were termed throughout the thesis as “regenerating neurons” by extrapolation from a similar terminology used for neurons cultured from the dissected CNS of Lymnea stagnalis (Farrar et al. 2009). The aim of this thesis was to determine if the synthesis of RA occurs within these neurons and, if so, where such synthesis is localised within sub-regions of the regenerating neuron. Such localisation of RA signalling can be genomic (RAR control of nuclear transcription) or non-genomic (RAR control of cytoplasmic kinases). Three types of neurons from hippocampus, cortex and retina, as well as the tissue of origin, were analysed by means of immunochemistry for the expression of Raldh1 and Raldh2 as well as RAR α, β and γ. Since many genes can be regulated by RA, a possible candidate gene, and effects of its inhibition on neurite outgrowth with or without RA, was investigated. Considering the promising results that appear on the horizon concerning the potential therapeutic effects that RA may have on neuronal regeneration, various retinoids were tested for genomic and non-genomic activity. The genomic activity was determined using a reporter cell line that has lac-Z driven by a RA response element while the non-genomic activity was assessed though an assay performed with MCF-7 cells that measured the phosphorylation of Erk1/2 as one of the potential RA-regulated downstream kinases. The main finding of this thesis was that in-vivo, neurons in postnatal hippocampus, cortex and retina showed very little expression of the components for RA synthesis while expression of RARs seemed to decrease dramatically within days of postnatal life. Nevertheless neurons from all these regions, when regenerating in culture, possessed the ability to synthesize RA as well as responding to it due to the up regulation of the necessary receptors. This thesis also suggested that the Raldhs, the main enzymes responsible for RA synthesis, are much more dynamic than previously considered and their location as well as intensity can change over time, or as a response to typical neuronal plasticity manipulation. Since all regenerating neurons switch on RA signalling within a couple of days of culture, this suggests that RA could be therefore the decisive factor that tips the balance between the two possible outcomes for a neuron that has its neurites cut away: to live and regenerate or to die. In the attempt to identify potential retinoids with greater effect in stimulating the ability of neurons to regenerate, it was found that some retinoids appeared to have a substantial influence on neurite outgrowth while different retinoids had unique combinations of genomic and non-genomic potencies.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Tretinoin ; Nervous system