Although numerous transcription factors (TFs) are expressed by oligodendrocytes (OGs), the role(s) of most of these TFs in oligodendrogenesis remains to be elucidated. One such TF is Hoxa2, which was recently shown to be expressed by O4-positive (+) pro-OGs. Hence, the main objectives of this thesis were to determine the expression profile and function(s) of Hoxa2 during OG development. Immunocytochemical analysis of primary mixed glial cultures demonstrated that Hoxa2 is expressed throughout oligodendrogenesis, diminishing only with the acquisition of a myelinating phenotype. Subsequently, immunohistochemical analysis suggested that Hoxa2 is expressed by migratory oligodendroglial cells in the embryonic spinal cord. However, double immunofluorescent analysis of Hoxa2 transgenic knockout mice showed that OG specification and early maturation proceed normally in the absence of Hoxa2 in the spinal cord.
As Hoxa2 is one of 39 murine Hox genes, which exhibit extensive overlapping expression profiles in the spinal cord, we decided to examine the expression of an additional Hox TF, Hoxb4, during OG development. Immunocytochemical analysis of primary mixed glial cultures demonstrated that Hoxb4 is also expressed throughout OG development. Furthermore, comparison of the expression profiles of Hoxb4 and Olig2 suggested that Hoxb4 is expressed by oligodendroglial cells in the spinal cord. Hence, Hoxb4, as well as other Hox TFs could compensate for Hoxa2 in the spinal cord in its absence.
As the anterior boundary of most Hox genes has been found to be in the hindbrain or spinal cord, we decided to look at the telencephalon which would be less likely to have compensatory mechanisms. Our results showed that similar to the spinal cord, Hoxa2 is expressed by oligodendroglial cells in the telencephalon. Subsequently, it was found that over-expressing Hoxa2 in CG4 cells, an oligodendroglial cell line derived from the perinatal rat cerebral cortex, impairs their differentiation. In an attempt to determine the mechanism by which it accomplishes this, we examined the expression of polysialylated neural cell adhesion molecule (PSA-NCAM), which has been implicated in this process. Contrary to our expectations, however, it was found that over-expressing Hoxa2 in CG4 cells results in significantly fewer PSA-NCAM+ cells. Hence, the results suggest that Hoxa2s effect on OG differentiation is independent of its effect on PSA-NCAM expression.
The expression of Hox genes is enhanced by retinoic acid (RA), which, in turn, both inhibits, as well as promotes OG differentiation. Although the reason for these opposing roles is uncertain, examination of the experimental protocols utilized by different research groups reveals disparities in age, CNS region, as well as RA concentration. As a result, RAs effect on oligodendrogenesis could be stage- and/or concentration-dependent. In order to determine which of these factors could impact RAs effect on OG differentiation we treated CG4 cells with two different concentrations of RA at two distinct time points. The results showed that both factors (concentration and time/stage) can impact RAs effect on CG4 cell differentiation. In an attempt to determine the mechanism by which it accomplishes this, we examined the expression of PSA-NCAM. Contrary to our expectations, the results suggest that RAs effect on CG4 differentiation is independent of its effect on PSA-NCAM expression. The results of this thesis suggest that Hoxa2 and RA could play multiple roles in OG development. Although these roles appear to be similar, further research will be needed to determine whether Hoxa2 acts a downstream effector in the RA signaling pathway in oligodendroglial cells.
| Identifer | oai:union.ndltd.org:USASK/oai:usask.ca:etd-08222007-140825 |
| Date | 23 August 2007 |
| Creators | Nicolay, Danette Jacine |
| Contributors | Verge, Valerie M. K., Nazarali, Adil J., Foldvari, Marianna, Doucette, J. Ronald, Devon, Richard, Bandy, Brian, Alcorn, Jane, Yong, V. Wee |
| Publisher | University of Saskatchewan |
| Source Sets | University of Saskatchewan Library |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://library.usask.ca/theses/available/etd-08222007-140825/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Saskatchewan or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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