Senescence plays a role in myotonic dystrophy type 1
Mikel García-Puga, Ander Saenz-Antoñanzas, Gorka Gerenu, Alex Arrieta-Legorburu, Roberto Fernández-Torrón, Miren Zulaica, Amets Saenz, Joseba Elizazu, Gisela Nogales-Gadea, Shahinaz M. Gadalla, Marcos J. Araúzo-Bravo, Adolfo López de Munain, Ander Matheu
Mikel García-Puga, Ander Saenz-Antoñanzas, Gorka Gerenu, Alex Arrieta-Legorburu, Roberto Fernández-Torrón, Miren Zulaica, Amets Saenz, Joseba Elizazu, Gisela Nogales-Gadea, Shahinaz M. Gadalla, Marcos J. Araúzo-Bravo, Adolfo López de Munain, Ander Matheu
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Research Article Aging Cell biology

Senescence plays a role in myotonic dystrophy type 1

Abstract

Myotonic dystrophy type 1 (DM1; MIM #160900) is an autosomal dominant disorder, clinically characterized by progressive muscular weakness and multisystem degeneration. The broad phenotypes observed in patients with DM1 resemble the appearance of an accelerated aging process. However, the molecular mechanisms underlying these phenotypes remain largely unknown. Transcriptomic analysis of fibroblasts derived from patients with DM1 and healthy individuals revealed a decrease in cell cycle activity, cell division, and DNA damage response in DM1, all of which related to the accumulation of cellular senescence. The data from transcriptome analyses were corroborated in human myoblasts and blood samples, as well as in mouse and Drosophila models of the disease. Serial passage studies in vitro confirmed the accelerated increase in senescence and the acquisition of a senescence-associated secretory phenotype in DM1 fibroblasts, whereas the DM1 Drosophila model showed reduced longevity and impaired locomotor activity. Moreover, functional studies highlighted the impact of BMI1 and downstream p16INK4A/RB and ARF/p53/p21CIP pathways in DM1-associated cellular phenotypes. Importantly, treatment with the senolytic compounds Quercetin, Dasatinib, or Navitoclax reversed the accelerated aging phenotypes in both DM1 fibroblasts in vitro and in Drosophila in vivo. Our results identify the accumulation of senescence as part of DM1 pathophysiology and, therefore, demonstrate the efficacy of senolytic compounds in the preclinical setting.

Authors

Mikel García-Puga, Ander Saenz-Antoñanzas, Gorka Gerenu, Alex Arrieta-Legorburu, Roberto Fernández-Torrón, Miren Zulaica, Amets Saenz, Joseba Elizazu, Gisela Nogales-Gadea, Shahinaz M. Gadalla, Marcos J. Araúzo-Bravo, Adolfo López de Munain, Ander Matheu

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Figure 5

DM1 fibroblasts undergo premature senescence.

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DM1 fibroblasts undergo premature senescence. (A) Results of 3T3 protoco...
(A) Results of 3T3 protocol where replicative senescence was induced in fibroblasts by seeding at a density of 30 × 103 cells per 10 cm2 every 3 days. Population doubling (PDL) of DM1 cells (n ≥ 5) and controls (n = 4) was calculated at each passage. (B and C) Representative images and quantification of SA–β-galactosidase+ cells at early (between 3 and 10) and late passage (between 35 and 40) in control and DM1 fibroblasts (n ≥ 5). Scale bar: 100 μm. (D and E) Representative immunoblot of p16INK4A, p21CIP1, and Lamin B1 proteins at early (between 5 and 10) and late (between 35 and 40) passages in DM1 relative to control fibroblasts (n ≥ 4). (F–H) mRNA levels of p16INK4A, p14ARF, p21CIP1, p27KIP1, and BMI1 in DM1 and controls (n ≥ 4), at early and late passages. (I) Antibody array of soluble factors secreted by control and DM1 fibroblasts. For each cell culture, control signals were averaged and used as the baseline. Signals above baseline are shown in yellow; signals below baseline are shown in blue. The heatmap indicates fold changes from baseline between DM1 (n = 3) and control fibroblasts (n = 3). (J) IL6, TNFα, and CCL5 mRNA levels at early and late passages (n ≥ 4). (K) Representative immunoblot of Lamin B1 proteins in PBMCs from DM1 and controls (n ≥ 5). (L) Measurement of IL6 mRNA in PBMCs derived from patients with DM1 (n ≥ 56) and controls (n ≥ 22). (M) Pearson’s correlation analysis between expression of senescence markers and leukocyte telomere length in patients with DM1. Telomere length was measured in DNA samples extracted with Qiagen in a previous study (18). P values were calculated using the Student’s t test with P value corrected for FDR. *P < 0.05, **P < 0.01, ***P < 0.001.