Reversal of DNA Damage-Induced Senescence in Endothelial Cells Using mRNA Telomerase

Introduction:: Radiation-induced vascular senescence is closely linked to cardiovascular disease. Such senescence is stimulated on by DNA damage resulting from radiation exposure, leading to genomic instability, telomere dysfunction, epigenetic shifts, and irregular gene expression. These changes often trigger an increase in inflammatory cytokine production, as the damaged cells turn into a state known as the secretory-associated senescent phenotype (SASP). Our previous research showed that telomerase mRNA (hTERT) therapy could rescue senescent iPSC-derived endothelial cells (ECs) in patients with progeria, a premature aging syndrome. This therapy not only normalized telomere length, but also restored the replicative capacity of the senescent ECs. Alongside these benefits, we observed a global normalization of transcriptional profiles, improved endothelial functions, and decreased DNA damage signals and inflammatory cytokine levels. Consequently, we propose that telomerase mRNA therapy may reduce radiation-induced DNA damage and senescence, restoring normal cellular functions.

Materials and Methods:: In our study, we used human aortic ECs (HAEC) that were transiently transfected with either hTERT-mRNA or GFP, or left untreated. The cells were exposed to 5 grays of radiation, and we measured levels of γH2A.X and cellular viability at different timepoints before and after radiation through immunoblotting, immunofluorescent staining, and FACS analysis. Now, in an attempt to prolong the positive effects of hTERT therapy, we're employing circular mRNA, which sustains hTERT expression and potentially enhances its cellular benefits. Upcoming analyses will measure telomerase expression, γH2A.X levels, cell viability, and telomere length, comparing these results to those achieved with linear hTERT mRNA.


Results, Conclusions, and Discussions:: As anticipated, radiation-exposed HAEC showed an increased level of DNA damage, as indicated by γH2A.X. However, HAEC pre-treated with hTERT-mRNA exhibited a marked reduction in γH2A.X, correlating with a significant decrease in cell death. While telomere length remained unchanged between the two cell types 72 hours post-radiation, hTERT-treated cells displayed a considerable reduction in another DNA damage marker (53BP1), inflammatory cytokines (IL6, IL8), and cells displaying the senescence marker β-gal.



Conclusion: Telomerase mRNA therapy appears to prevent radiation-induced DNA damage and cellular senescence through a non-canonical function of telomerase.




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