Investigating the MeCP2 (E1/E2) regulatory network in the human control and Rett syndrome brain
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Date
2018-12-12
Authors
Pejhan, Shervin
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Abstract
Rett Syndrome (RTT) is a neurodevelopmental disorder characterized by neurological regression and autism spectrum features, usually in females with mutation in MECP2. Animal and cell models have contributed to our knowledge about RTT and MeCP2 function. However, many findings remain unverified in the human brain.
Based on a study on the rat brain, miR132, a neuronal microRNA that is highly conserved among vertebrates, is suggested to inhibit MeCP2. This microRNA is induced itself by Brain Derived Neurotrophic Factor (BDNF), a neurotrophin, which is controlled by MeCP2.
In an effort to study the possible translation of this suggested regulatory network from rat to human brain, and to evaluate the hypothesized changes of this network in the brain of RTT patients, I examined post-mortem brain samples of RTT patients and compared them with their age-, and sex-matched controls.
For the first aim of my project, I evaluated the transcript and protein levels of MECP2/MeCP2 isoforms, BDNF/BDNF, and the level of miR132 by real time RT-PCR, Western blot, and ELISA in four brain regions of three human RTT brains and their age-, post-mortem delay-, and sex-matched controls.
The transcript level of the studied elements was significantly different in RTT patients; even though the change was not identical in all studied parts of the brain. The protein level of the studied elements did not follow the same pattern as the transcripts.
For the second aim of my project, I performed immunohistochemistry studies on three brain regions of four RTT patients and their age-, and sex-matched controls. My results showed significantly reduced immunoreactivity for MeCP2 protein, exclusively in the glial cells of the white matter, but surprisingly, neurons did not show a noticeable decrease in MeCP2 staining. Immunolabeling of brain samples for BDNF revealed a dominant astroglial/endothelial pattern without noticeable difference between RTT and control tissues.
Furthermore, for the first time, the effect of post-mortem delay in fixation on MeCP2 and BDNF immunolabeling of human brain samples was studied in this project. MeCP2 protein appeared to be more sensitive to post-mortem delay than BDNF.
Despite challenges in evaluating autopsy samples, this project provides insight on RTT pathobiology, while highlighting the importance of studying human RTT brain in parallel with rapidly growing research on animal and cell models of the disease.
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Keywords
MECP2 mutations, Rett syndrome, MeCP2 isoforms, BDNF, miR132, Epigenetics