MiR-21-5p regulates the dynamic of mitochondria network and rejuvenates the senile phenotype of bone marrow stromal cells (BMSCs) isolated from osteoporotic SAM/P6 mice

Mateusz Sikora, Agnieszka Śmieszek, Ariadna Pielok, Krzysztof Marycz

Progression of senile osteoporosis is associated with deteriorated regenerative potential of bone marrow-derived mesenchymal stem/stromal cells (BMSCs). According to the recent results, the senescent phenotype of osteoporotic cells strongly correlates with impaired regulation of mitochondria dynamics. Moreover, due to the ageing of population and growing osteoporosis incidence, more efficient methods concerning BMSCs rejuvenation are intensely investigated. Recently, miR-21-5p was reported to play a vital role in bone turnover, but its therapeutic mechanisms in progenitor cells delivered from senile osteoporotic patients remain unclear. Therefore, the goal of this paper was to investigate for the first time the regenerative potential of miR-21-5p in the process of mitochondrial network regulation and stemness restoration using the unique model of BMSCs isolated from senile osteoporotic SAM/P6 mice model. BMSCs were isolated from healthy BALB/c and osteoporotic SAM/P6 mice. We analysed the impact of miR-21-5p on the expression of crucial markers related to cells' viability, mitochondria reconstruction and autophagy progression. Further, we established the expression of markers vital for bone homeostasis, as well as defined the composition of extracellular matrix in osteogenic cultures. The regenerative potential of miR-21 in vivo was also investigated using a critical-size cranial defect model by computed microtomography and SEM-EDX imaging. MiR-21 upregulation improved cells' viability and drove mitochondria dynamics in osteoporotic BMSCs evidenced by the intensification of fission processes. Simultaneously, miR-21 enhanced the osteogenic differentiation of BMSCs evidenced by increased expression of Runx-2 but downregulated Trap, as well as improved calcification of extracellular matrix. Importantly, the analyses using the critical-size cranial defect model indicated on a greater ratio of newly formed tissue after miR-21 application, as well as upregulated content of calcium and phosphorus within the defect site. Our results demonstrate that miR-21-5p regulates the fission and fusion processes of mitochondria and facilitates the stemness restoration of senile osteoporotic BMSCs. At the same time, it enhances the expression of RUNX-2, while reduces TRAP accumulation in the cells with deteriorated phenotype. Therefore, miR-21-5p may bring a novel molecular strategy for senile osteoporosis diagnostics and treatment.