Bone Marrow-Derived Stem Cells Reduce Radiation-Induced Damage to Salivary Glands.

Abstract

The salivary glands are often included in the field of irradiation during radiotherapy of head and neck cancer. This can result in severe side-effects that reduces the quality of life of the patient and may even limit the treatment dose. Late damage to the salivary glands is mainly caused by exhaustion of the tissue specific stem cells. Post-irradiation replacement of salivary gland stem cells with donor stem cells may ameliorate radiation-induced complications. Bone marrow-derived stem cells (BMSC) have been shown to be multipotent and thereby able to engraft in many tissues after injury. In this study, we assessed the potential of BMSC to reduce irradiation-induced salivary gland damage. C57BL/6 mice were transplanted with bone marrow from eGFP transgenic animals. After two months the salivary glands of these chimeric mice were locally irradiated with 15 Gy. BMSC were mobilized 10, 30 and/or 60 days after irradiation by s.c. injection of rHu-PEG-G-CSF. Saliva secretion (μl/15 minutes) was measured up to 90 days after irradiation by pilocarpine induction. Hereafter, the glands were extirpated and examined for eGFP-expression. From every individual animal one parotid and one submandibular gland was used to prepare single cell suspensions in order to detect eGFP-positve cells by flow cytometry. The other parotid and submandibular glands were analyzed using confocal laser fluorescence scanning microscopy and light microscopy. G-CSF treatment yielded in an increase of saliva flow for all time points. The optimal time-point for mobilization, however, was 30 days after irradiation as is demonstrated by an improvement of salivary flow from 5 to 30% when compared to radiation alone. FACS analysis showed that up to 10% of the isolated cells were eGFP-positive. Microscopic analysis revealed a similar amount of positive cells and an improved morphology. Immuno-histochemistry using anti-SM-actin antibodies showed the close vicinity of actin and eGFP within the cells, demonstrating the occurence of BMSC derived myoepithelial cells in irradiated salivary glands. Furthermore, using cell-type specific antibodies, the meyoepethilial nature of the eGFP positive was revealed. In conclusion, the results show that BMSC home to severely damaged salivary glands after mobilization. Hence, BMSC mobilization could become a promising modality to ameliorate radiation-induced complications in salivary glands after radiotherapy.