The Impact of environmental and genetic insults on adult and developing brain oligodendrogenesis
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The focus of this dissertation was to understand how brain oligodendrocytes are affected by oxidative stress throughout development and in adulthood. The stressors ranged from the blast induced brain ischemia to neurodevelopmental disorders, all of which had been shown to affect the mitochondrial oxidative phosphorylation. Impaired mitochondrial function is accompanied by the loss of myelin indicative of impaired oligodendrocyte function. My overall hypothesis was that oligodendrocytes are damaged by oxidative stress but their precursor cells have significant regenerative capacity which may be augmented through stimulation of mitochondrial function. I have tested three postulates : 1) changes in myelin expression are associated with the loss of the central nervous system myelinating cells – oligodendrocytes; 2) the stem/progenitor cells in the brain neurogenic niches and/or local cortical oligodendrocyte precursor cells can increase the production of oligodendrocytes and thus compensate for the loss; and 3) the regeneration of oligodendrocytes can be augmented by pharmacological treatment with Phenylbutyrate, which targets oxidative phosphorylation and epigenetic control of the cell development. I have utilized three different oxidative stress, mouse and cerebral organoid, models. The first model, chapter 2, was the blast-induced brain injury in an adult mouse, in which the vascular-oxidative stress was generated by an external acoustic stimulus. Using this model, I found that blast-induced brain cortical injury and transient cortical demyelination were followed by a regrowth of myelinated fibers. Blast induced an initial loss of the oligodendrocyte population expressing Oligodendrocyte marker 4 which recovered within 21 days after blast. The recovery of Olig4 oligodendrocytes parallels and may have contributed to the regeneration myelinated cortical fibers. Blast had no effect on the proliferation of neural stem/progenitor cells in the telencephalic subventricular zone or in the hippocampal subgranular zone. In contrast, in the cortex, blast transiently increased cell proliferation and induced-oligodendrocytic cells expressing Olig2, an early determinant of oligodendrocyte fate specification. These results indicated that the regeneration of cortical oligodendrocyte population was due to a local activation of the oligodendrogenesis. The population of proliferating Oligodendrocytes Progenitor Cells (Ki67+/Olig4+) was small in adult mouse brain cortex, SVZ and SGZ, therefore it could not be evaluated. The second model, chapter 3, employed was Pyruvate Dehydrogenase Complex (PDC)-deficient mouse in which oxidative stress was induced prenatally by a hemizygous mutation of the Pyruvate Dehydrogenase gene. The PDC-deficient mice were analyzed at postnatal day 35, at which point the mice showed an overall reduction in brain cell proliferation and in the populations of both oligodendrocytes and their progenitor cells. Unlike the transient blast insult, in the PDC-deficient mice there was a no compensatory increase in oligodendrogenesis. However, treatment of the PDC-deficient mice with Phenylbutyrate from postnatal day 2 to day 35 - effectively upregulated the oligodendrocyte population and restored oligodendrocyte progenitor cells to the levels found in control PDC-normal mice The final model, chapter 4, a human pluripotent stem cell-based model, I employed cerebral organoids in which transient inhibition of the oxidative phosphorylation and the oxidative stress were induced by Rotenone. For the first time oligodendrogenesis was shown and quantified during development of human cerebral organoids (day 17 to day 27). A short-term, 3-day, rotenone treatment depleted proliferating organoid neural progenitor cell while increasing migration of the residual NPC. Rotenone treatment also depleted the population of Olig4+ oligodendrocytes and a similar trend was observed with oligodendrocyte progenitor cells. When allowed an additional post-Rotenone recovery time, the organoids showed spontaneous cellular regeneration that was enhanced by treatment with the Phenylbutyrate. Specifically, Phenylbutyrate restored the oligodendrocyte progenitor cell numbers and increased the oligodendrocyte population depleted by 3 days of Rotenone treatment. In contrast, after an extended, 6 days of rotenone neither spontaneous nor Phenylbutyrate-induced recoveries were noted. In summary, the cerebral organoids provide an effective model to study human brain developmental oligodendrogenesis, its disruption in oxidative stress and development of new corrective therapies. My overall findings showed that the population of brain cortical oligodendrocytes are depleted during demyelination in blast induced brain ischemia, PDC deficient mitochondrial dysfunction and in directly suppressed oxidative phosphorylation by Rotenone. Following transient insults, the oligodendrocyte population is capable of significant spontaneous recovery in both the mature and developing brain models. Both the spontaneous recovery following the direct blockade of mitochondrial function and of the permanently depleted oligodendrocytic population in genetic disorder may be augmented by Phenylbutyrate. Thus, Phenylbutyrate may have therapeutic utility for the regeneration of oligodendrocyte population after oxidative stress-causing insults.