Mesenchymal stromal cells having inactivated RB1 survive following low irradiation and accumulate damaged DNA: Hints for side effects following radiotherapy

ABSTRACT

Following radiotherapy, bone sarcomas account for a significant percentage of recurring tumors. This risk is further increased in patients with hereditary retinoblastoma that undergo radiotherapy. We analyzed the effect of low and medium dose radiation on mesenchymal stromal cells (MSCs) with inactivated RB1 gene to gain insights on the molecular mechanisms that can induce second malignant neoplasm in cancer survivors.

MSC cultures contain subpopulations of mesenchymal stem cells and committed progenitors that can differentiate into mesodermal derivatives: adipocytes, chondrocytes, and osteocytes. These stem cells and committed osteoblast precursors are the cell of origin in osteosarcoma, and RB1 gene mutations have a strong role in its pathogenesis. Following 40 and 2000 mGy X-ray exposure, MSCs with inactivated RB1 do not proliferate and accumulate high levels of unrepaired DNA as detected by persistence of gamma-H2AX foci. In samples with inactivated RB1 the radiation treatment did not increase apoptosis, necrosis or senescence versus untreated cells. Following radiation, CFU analysis showed a discrete number of cells with clonogenic capacity in cultures with silenced RB1.

We extended our analysis to the other members of retinoblastoma gene family: RB2/P130 and P107. Also in the MSCs with silenced RB2/P130 and P107 we detected the presence of cells with unrepaired DNA following X-ray irradiation. Cells with unrepaired DNA may represent a reservoir of cells that may undergo neoplastic transformation. Our study suggests that, following radiotherapy, cancer patients with mutations of retinoblastoma genes may be under strict controls to evaluate onset of secondary neoplasms following radiotherapy.

KEYWORDS:

• Apoptosis
• DNA damage
• mesenchymal stem cells
• senescence

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.

Acknowledgments

We thank Drs. Vincenzo Condè, Daniela Di Pinto, Martina Di Martino (Servizio di Oncologia Pediatrica, AOU, Seconda Università di Napoli) for bone marrow harvests and technical assistance. We thank Dr. Giovanni Fulgione (Servizio di Radiologia, AOU, Seconda Università di Napoli) for irradiation technical assistance. We thank Mrs. Maria Rosaria Cipollaro (Dept. of Experimental Medicine, Second University of Naples) for technical assistance.

Authors' contributions

Stefania Capasso, Nicola Alessio, Anna Calarco and Giovanni Di Bernardo carried out collection and assembly of data and contributed to data analysis and interpretation.

Marilena Cipollaro, Fiorina Casale and Salvatore Cappabianca carried out data analysis and interpretation.

Umberto Galderisi and Gianfranco Peluso carried out conception and design, contributed to data analysis and wrote the manuscript.

Additional information

Funding

The research leading to these results has received funding from the European Union Euratom Seventh Framework Program RISK-IR project under grant agreement n°323267 to UG, and from Progetto PON – ‘Ricerca e Competitivita´ 2007–2013’ – PON01_00802 entitled ‘Sviluppo di molecole capaci di modulare vie metaboliche intracellulari redoxsensibili per la prevenzione e la cura di patologie infettive, tumorali, neurodegenerative e loro delivery mediante piattaforme nano tecnologiche’ to GP.