gms | German Medical Science

Objectives: Senescent cells have detrimental effects across tissues with aging, in particular in the liver and brain. However, they also may have beneficial effects on tissue repair, specifically on skin wound healing. Whether senescence does play a role in the dynamic bone healing process is currently unknown. As well, the potential role of senescent cells in fracture healing has not been defined.

Methods: We first performed an in-silico analysis of public mRNAseq data evaluating six different time points of long bone healing (One-way ANOVA). We then analyzed senescent biomarkers in tibial fracture healing in vivo (C57BL/6) with qPCR and Fluorescence in situ hybridization (FISH, One-way ANOVA, Student's t-test). After that, we utilized a genetic mouse model (Alb.B6.Cdkn2a-Luciferase) to verify our findings via qPCR, X-ray, micro-CT and luminescence detection (One-way ANOVA, Student's t-test). Subsequently, we eliminated senescent cells via application of modern senolytics (Dasatinib plus Quercetin) to assess the impact on fracture healing via qPCR, X-ray, micro-CT and biomechanical testing (One-way ANOVA, Student's t-test).

Results and conclusion: We found that senescence and senescence-associated secretory phenotype (SASP) markers increased during fracture healing. We established a time course of senescence biomarkers during murine fracture healing and directly identified cells in the fracture callus that displayed hallmarks of senescence, including distension of satellite heterochromatin and telomeric DNA damage. In the reporter mouse model, we demonstrated transient in vivo senescent cell accumulation mirroring our in silico and in vivo transcriptional prediction during callus formation.

Finally, and with intermittent treatment of young adult mice following fracture with drugs that selectively eliminate senescent cells ("Senolytics", Dasatinib plus Quercetin), we showed that this regimen both decreased senescence and SASP markers in the fracture callus and significantly accelerated the time course of fracture healing in both x-ray and biomechanical callus healing assessment.

Our findings demonstrate that senescent cells accumulate transiently in the murine fracture callus and, in contrast to the skin, their clearance does not impair but rather improves fracture healing. Senolytic therapy was supportive for the fracture healing process.

Figure 1 [Fig. 1]