Abstract
Motor neuron degeneration in amyotrophic lateral sclerosis (ALS) caused by mutations in superoxide dismutase 1 (SOD1) is partly non-cell autonomous, involving cellular dysfunction of astrocytes. Whether non-cell autonomous effects occur in other forms of ALS, such as TAR DNA binding protein 43 (TDP-43)-related disease, remains unclear. Here, we characterised the impact of mutant TDP-43 expression on primary astrocytes derived from transgenic TDP-43A315T mice. Mutant TDP-43 astrocytes revealed evidence for TDP-43 pathology, shown by cytoplasmic TDP-43 inclusions and accumulation in insoluble cell fractions which was exacerbated by proteasomal inhibition. l-glutamate uptake, measured using an [3H]D-aspartate assay, was impaired in mutant TDP-43 astrocytes, while ATP accumulation was abnormal, suggesting mutant TDP-43 induced astrocytic dysfunction. Astrocyte activation coupled with spinal and cortical motor neuron loss in transgenic TDP-43A315T mice could imply non-cell autonomous effects of astrocytes in vivo. These data demonstrate mutant TDP-43-mediated cell autonomous effects on astrocytes that may contribute to motor neuron pathology in ALS.
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Acknowledgements
PMB is pleased to contribute a paper to this Special Issue honouring Michael Robinson who has been a colleague furthering the neurochemical cause via Neurochemistry International and internationally (ISN) for some 20 years. Funding for this project was provided by the Australian NHMRC (Project Grant 1023780 P.M.B.; Fellowship 1020401 P.M.B.; Fellowship 1137024 B.J.T; and joint NHMRC-ARC Fellowship 1110040 S.K.B) and the Stafford Fox Medical Research Foundation. The Florey Institute of Neuroscience & Mental Health acknowledge Victorian Government Operational Infrastructure Support.
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BJT, PMB and HM conceived the study design. CLL, MDFC and DT performed the experimental procedures. SKB and BJT collated the data and wrote the manuscript and all authors approved the final version.
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Special Issue In honor of Professor Michael Robinson
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11064_2020_3048_MOESM1_ESM.tiff
Supplementary file1 (TIFF 2703 kb) Suppl Figure 1: Inhibiting the UPS alters astrocyte morphology. Untreated 12 DIV TDP-43A315T astrocytes had increased GFAP expression compared to WT astrocytes. Treating astrocytes with increasing doses of MG-132 altered astrocyte morphology and GFAP expression but there was no longer a noticeable difference between WT and TDP-43A315T astrocytes. Scale bar 50 µm.
11064_2020_3048_MOESM2_ESM.tiff
Supplementary file2 (TIFF 2703 kb) Supplemental Figure 2: Inhibiting the Ubiquitin Proteasome System (UPS) causes decreased TDP-43 in the soluble nuclear fraction in 12 DIV TDP-43A315T astrocytes. The synthetic peptide MG-132 was used at increasing doses to inhibit the UPS. MG-132 had no effect on TDP-43 levels in the nucleus in WT astrocytes and whilst TDP-43A315T astrocytes of matching doses had significantly increased TDP-43 levels compared to their WT counterparts, the 3.0 μM dose of MG-132 caused a significant decrease in the amount of TDP-43 in the nuclear protein fraction compared to untreated TDP-43A315T astrocytes (A). There was no difference in the proportion of cytoplasmic TDP-43 to total soluble TDP-43 in any groups, regardless of genotype (B) indicating re-distribution of TDP-43 to the insoluble protein fraction in TDP-43A315T astrocytes. #p<0.05 compared to untreated TDP43A315T, *p<0.05 compared to the WT treated with the same concentration. n = 3 mice per genotype for all groups.
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Barton, S.K., Lau, C.L., Chiam, M.D.F. et al. Mutant TDP-43 Expression Triggers TDP-43 Pathology and Cell Autonomous Effects on Primary Astrocytes: Implications for Non-cell Autonomous Pathology in ALS. Neurochem Res 45, 1451–1459 (2020). https://doi.org/10.1007/s11064-020-03048-5
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DOI: https://doi.org/10.1007/s11064-020-03048-5