Long-term cryopreservation model of rat salivary gland stem cells for future therapy in irradiated head and neck cancer patients

Irradiated head and neck cancer patients suffer from irreversible loss of salivary gland (SG) function, along with significant morbidity and compromised quality of life. To date there is no biologically-based treatment for this distress. Adult salivary gland stem cells are promising candidates for autologous transplantation therapy in the context of tissue-engineered artificial SGs or direct cell therapy. The major restrictions in handling such cells are their limited lifespan during in vitro cultivation, resulting in a narrow time-window for implantation and a risk of tumorigenic changes during culture. To overcome these difficulties, we tested in a rat model the possibility of establishing a personal/autologous SG stem cell bank. SG's integrin-α6β1-expressing cells were shown to hold a subpopulation of SG-specific progenitor-cells. Explanted and cultured single cell-originated clones were cryopreserved for up to 3 years and shown to exhibit genetic and functional stability similar to noncryopreserved cells, as was emphasized by soft agar assay, division potential assessment, flow cytometric analysis, real-time reverse transcriptase-polymerase chain reaction, in vitro three-dimensional differentiation assay, and immunofluorescence confocal microscopy. Future integration of the novel strategies presented herein to a clinical therapeutic model will allow safe preservation until transplantation and repeated transplantation if needed. These tools open a new venue for adult autologous stem-cell transplantation-based SG regeneration.