Progranulin reduces insoluble TDP-43 levels, slows down axonal degeneration and prolongs survival in mutant TDP-43 mice

Abstract

Background: TAR DNA binding protein 43 (TDP-43) is the main disease protein in most patients with amyotrophic lateral sclerosis (ALS) and about 50% of patients with frontotemporal dementia (FTD). TDP-43 pathology is not restricted to patients with missense mutations in TARDBP, the gene encoding TDP-43, but also occurs in ALS/FTD patients without known genetic cause or in patients with various other ALS/FTD gene mutations. Mutations in progranulin (GRN), which result in a reduction of ~ 50% of progranulin protein (PGRN) levels, cause FTD with TDP-43 pathology. How loss of PGRN leads to TDP-43 pathology and whether or not PGRN expression protects against TDP-43-induced neurodegeneration is not yet clear.

Methods: We studied the effect of PGRN on the neurodegenerative phenotype in TDP-43(A315T) mice.

Results: PGRN reduced the levels of insoluble TDP-43 and histology of the spinal cord revealed a protective effect of PGRN on the loss of large axon fibers in the lateral horn, the most severely affected fiber pool in this mouse model. Overexpression of PGRN significantly slowed down disease progression, extending the median survival by approximately 130 days. A transcriptome analysis did not point towards a single pathway affected by PGRN, but rather towards a pleiotropic effect on different pathways.

Conclusion: Our findings reveal an important role of PGRN in attenuating mutant TDP-43-induced neurodegeneration.

Keywords: Amyotrophic lateral sclerosis; Frontotemporal dementia; Progranulin; TDP-43.

Fig. 1

PGRN overexpression has no effect on TDP-43 RNA levels. RNA expression of the human TDP-43(A315T) transgene (a), mouse Tardbp (b) and mouse Grn (c) in the spinal cord is unchanged by PGRN overexpression. ***p < 0.001, one-way ANOVA. d No significant changes were observed in Iba1 expression in the spinal cord. Data are shown as mean ± SEM

Fig. 2

Human PGRN overexpression has no effect on mouse PGRN or CTSD protein levels. a-b Protein expression of mouse PGRN in brain and spinal cord from NTG, TDP-43(A315T) and TDP-43(A315T)xGRN mice. c-d Protein expression of human PGRN and mouse CTSD in brain and spinal cord NTG, TDP-43(A315T) and TDP-43(A315T)xGRN mice. °Aspecific band

Fig. 3

PGRN overexpression reduces insoluble TDP-43 levels. a, e Western blot analysis of total, soluble and insoluble TDP-43 in brain (a) and spinal cord (e) from NTG, TDP-43(A315T) and TDP-43(A315T)xGRN mice. Quantification of blots (b-d and f-h) are shown as mean ± SEM (n = 3 per group, * p < 0.05, ** p < 0.001, *** p < 0.0001, Tukey-Kramer multiple comparison test)

Fig. 4

PGRN overexpression prevents degeneration of large axon fibers. a Schematic overview of the region in the lateral spinal cord used for the analysis of axon fibers as indicated by the black square. b No difference was found in the number of small axon fibers (1–4 μm²) in this region. c Representative images of the axon fibers in the lateral spinal cord (Scale bar = 10 μm). d The mean number of large axon fibers in NTG and TDP-43(A315T)xGRN mice was significantly higher across all size groups, compared to TDP-43(A315T) mice (n = 6–9 per group, *p < 0.05, Wilcoxon signed-rank test). Data are shown as mean ± SEM

Fig. 5

PGRN overexpression increases survival and disease duration in TDP-43(A315T) mice. a Disease onset is not affected by PGRN overexpression (n = 26 and 54 for TDP-43(A315T) and TDP-43(A315T)xGRN mice, respectively). b The survival is significantly increased by PGRN overexpression. Median survival: 364 d (TDP-43(A315T)) versus 491 d (TDP-43(A315T)xGRN). **p < 0.01, Log-rank test. c PGRN overexpression significantly increased the survival after symptom onset. Median disease duration: 91 d (TDP-43(A315T)) versus 150 d (TDP-43(A315T)xGRN). **p < 0.01, Log-rank test