DNA-damage, survival, differentiation, and matrix mineralization in vitro of a murine multipotent mesenchymal precursor cell line

Abstract

Radiation therapy is one of the most effective and indispensable treatment modalities for cancer patients. Known tissue complications caused by radiation-induced stem cell depletion, may result in structural and functional alterations of the surrounding matrix. Although, studies have demonstrated that ionizing radiation can induce apoptosis and senescence, little is known about the effects of therapeutic irradiation concerning the commitment of mesenchymal stem cells to the osteoblastic lineage. C3H10T1/2 clone eight cells were used reflecting an early stage of differentiation. Notably, radiation doses of 2 Gy reduced proliferation, but had no significant effect on cell viability. Cell cycle analysis revealed that the yield of cells captured in the G₂/M phase of the cell cycle was markedly and dose-dependently increased. Instead of apoptosis we detected increased activity of stress-induced premature cellular senescence. Histochemical staining and quantification of the hydroxyapatite content of the extracellular bone matrix revealed positive staining for alizarin red S. Expression of TP53 encoding for tumour suppressor protein p53 and its downstream target cyclin-dependent kinase inhibitor 1A (p21(Cip1/Waf1)) were significantly increased. Gene expression analysis of two osteoblast specific genes, Runx2 and osteocalcin were assessed. Here, we confirmed that exposure to X-rays was dose dependently effective in decreasing cellular survival. Our results indicate that the direct impairment of proliferation and osteogenic differentiation potential of MSCs by irradiation may contribute partly to post-irradiation osteoporosis.