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Dissertation SungWon Final revised.pdf (11.67 MB)

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ALS Astrocytes Adopt Natural Killer Properties to Induce Motor Neuron Death

Song, SungWon
http://rave.ohiolink.edu/etdc/view?acc_num=osu1416847886

Abstract Details

2014, Doctor of Philosophy, Ohio State University, Molecular, Cellular and Developmental Biology.
Selective degeneration and death of one or more subtypes of neurons is the defining feature of human neurodegenerative disorders. Although traditionally these diseases have been thought to mainly affect neurons, a vulnerable cell type in the central nervous system, the results of more recent studies indicate the involvement of non-neuronal cells, such as glial cells. Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease characterized by progressive, fatal paralysis due to degeneration and death of motor neurons. Like other neurodegenerative disorders, numerous studies indicate that the motor neuron damage found in ALS is accelerated by multiple glial cell types. These glia selectively become toxic for motor neurons during the disease. Among those, recent studies have highlighted the important role of ALS astrocytes, which lose their ability to support neurons and develop an aberrant phenotype, gaining toxicity to kill motor neurons. This is highlighted by in vitro models recapitulating the neurotoxic properties of ALS astrocytes. These model systems allow the identification of molecular mechanisms and targets in ALS astrocyte toxicity. They also enable the development of new therapeutic strategies for ALS. For the development of reliable and reproducible in vitro models of ALS, it is crucial to have easy access into motor neurons and astrocytes. Motor neurons are impossible to obtain directly from patients and their extraction from postmortem tissues is limited, due to the complexity of isolating living cells from an adult brain or spinal cord. Reprogramming of somatic cells to pluripotency was suggested as a valuable resource for the generation of patient-specific motor neurons and astrocytes. In the current study, we successfully generated four non-ALS control, three familial ALS, and nine sporadic ALS-induced pluripotent stem cell (iPSC) lines. This was performed by delivering four reprogramming transcription factors, Oct3/4, Klf4, Sox2, and c-Myc into donor- derived skin fibroblasts. Human embryonic stem cells (ESCs) or iPSCs can be differentiated into functional motor neurons. By utilizing three of motor neuron-inducing transcription factors—Ngn 2, Islet-1, and Lhx3—we could accelerate the production of functional human motor neurons by 50 days, with >60-70% improved efficiency. Using this technique for the efficient and rapid production of human motor neurons, we finally established the in vitro human co-culture model for ALS. We found astrocytes derived from both familial ALS (FALS) and sporadic ALS (SALS) patients were toxic to human motor neurons. This opens a new avenue of study, in which cross talk between multiple cellular components of ALS in humans can be examined. These findings that astrocytes derived from ALS patients are toxic to human motor neurons add to the growing evidence that non-cell-autonomous mechanisms contribute to motor neuron death in ALS, and point to a crucial role of glia in ALS pathogenesis. However, the mechanisms driving ALS astrocyte toxicity toward motor neurons in ALS still remain unknown. In the current study, we identified unexpected cellular properties of murine and human FALS and SALS astrocytes in eliminating motor neurons. We showed mouse and human ALS astrocytes express cytolytic proteins, perforin, and granzyme B. These ALS astrocytes actively release perforin to generate pores on motor neuron membranes and deliver apoptosis-inducing a factor, granzyme B, into motor neurons to induce target cell death. Suppression of perforin or granzyme B expression in mouse and human FALS and SALS astrocytes resulted in an almost complete protection of motor neuron survival during in vitro co-culture, suggesting functional relevance of this granule exocytosis pathway in explaining ALS astrocyte toxicity to motor neurons. Despite the fact that ALS astrocytes are toxic to motor neurons, little is known about the mechanisms behind the death of motor neurons by astrocytes. In the current study, we showed that a decrease in major histocompatibility complex class I (MHC class I) presentation by motor neurons is associated with their susceptibility to ALS astrocyte-induced toxicity and can act as a signal for the release of perforin and granzyme B by ALS astrocytes. Spinal motor neurons displayed marked reduction in MHC class I presentation at symptomatic stages of ALS pathology. Restored expression of MHC class I in motor neurons rescued them from ALS astrocyte-mediated death in vitro in both mouse and human, and significantly extended survival in ALS mice. We also found that ALS astrocytes express MHC class I receptors, which enable ALS astrocytes to sense the reduced level of MHC class I in motor neurons, recognizing motor neurons to be killed. These findings extend our knowledge of how ALS astrocytes selectively discriminate their target neurons—motor neurons—before engaging in killing. Overall, our results suggest that ALS astrocytes show drastic phenotypic changes during the disease and utilize mechanisms, typically ascribed to the cytotoxic process in an innate immune system, in recognition and death of motor neurons. Our discoveries have the great potential for therapeutic intervention to modify disease progression in ALS.
Brian Kaspar (Advisor)
Denis Guttridge (Committee Member)
Arthur Burghes (Committee Member)
Christopher Walker (Committee Member)
218 p.
Biomedical Research

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Citations

  • Song, S. (2014). ALS Astrocytes Adopt Natural Killer Properties to Induce Motor Neuron Death [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1416847886

    APA Style (7th edition)

  • Song, SungWon. ALS Astrocytes Adopt Natural Killer Properties to Induce Motor Neuron Death. 2014. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1416847886.

    MLA Style (8th edition)

  • Song, SungWon. "ALS Astrocytes Adopt Natural Killer Properties to Induce Motor Neuron Death." Doctoral dissertation, Ohio State University, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=osu1416847886

    Chicago Manual of Style (17th edition)

osu1416847886
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This open access ETD is published by The Ohio State University and OhioLINK.