Date of Award
Daniel A. Linseman
ALS, BH3-only Proteins, G93A mutant SOD1, Immunohistochemistry, Neurodegeneration, Real-Time PCR
Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease involving rapid degeneration of motor neurons in the spinal cord and retraction of their axonal projections to the neuromuscular junctions. Several known mutations linked to some familial cases of ALS have been linked to mutations in Cu/Zn superoxide dismutase (SOD1), resulting in mitochondrial oxidative stress and intrinsic apoptosis. Transgenic mice expressing a G93A mutant of SOD1 provide an in vivo model to investigate motor neuron death during disease progression. The principal regulators of intrinsic apoptosis are the Bcl-2 family proteins. While some members of this family are pro-survival, the Bcl-2 homology-3 domain (BH3)-only proteins are pro-apoptotic. Since cooperation of various BH3-only proteins is often necessary to induce apoptosis, we hypothesized that multiple BH3-only proteins are induced in spinal motor neurons during the progression of ALS. Furthermore, we postulated that these pro-apoptotic proteins act in a coordinated manner to cause motor neuron death. We utilized laser capture microdissection (LCM) to collect highly enriched populations of spinal motor neurons from wild type vs. SOD1 mutant mice. RNA was then isolated from the captured motor neurons and used for quantitative real time PCR analysis of BH3-only transcript expression. We did not detect any significant differences in the expression of BH3-only transcripts between end stage SOD1 mutant mice and age-matched wild type animals. In contrast, immunohistochemical staining for the BH3-only proteins, Bik, Bad, BNip3, Bid, Noxa, Puma, and Hrk/Dp5, demonstrated selective staining of Hrk/Dp5, Bnip3, and Bid in astrocytes of lumbar spinal cord from end stage mutant SOD1 mice. Hrk/Dp5, Bnip3, and Bid were not observed in astrocytes of wild type mouse spinal cords. These novel findings indicate a potentially important role for astrocytes expressing Hrk/Dp5, Bnip3, and Bid in ALS disease progression.
Andrianakos, Anna George, "Role of Pro-Apoptotic BCL2-homology-3 Domain (BH3)-only Proteins in the Mutant SOD1 Mouse Model of ALS" (2009). Electronic Theses and Dissertations. 750.
Recieved from ProQuest
Anna George Andrianakos