mRNA from Ionizing radiation treatment of chondrocyte-like cells derived from human induced pluripotent stem cells: gene and microRNA expression profile and high-throughput screening of secreted cytokines, chemokines, and growth factors

Human induced pluripotent stem cells (hiPSCs) possess the ability to differentiate into chondrocytes and produce cartilaginous matrix without the need to biopsy healthy cartilage. However, this strategy is not without risk given that those transplanted cells can be subjected to IR during Head and Neck (H&N) (re-)treatment. The main aim of this study was to investigate the gene expression profile. The differentiated hiPSC into chondrocyte were irradiated in accordance with conventional therapy, hypofractionation and hyperfractionation protocols (1, 2, 3 Gy) intended for larynx region. Then, to fulfill objective we performed following high-throughput analysis: gene expression profile based on microarrays. Our findings allowed for the selection of crucial biological processes activated in hiPSC-derived chondrocytes after IR such as: “apoptotic process”, “cellular response to DNA damage stimulus” and “regulation of programmed cell death”. Moreover, we observed decrease of expression of genes involved in e.g. cell cycle and cell division. Microarray data were confirmed by RT-qPCR technique. We have proved that IR have a great impact on DDR mechanisms of hiPSCs differentiated towards chondrogenic lineage. To irradiate the chondrocyte-like cells the specially modified and redesigned the Alderson Radiation Therapy phantom (ART) was used to obtain similar condition to those in patient’s larynx region treatment. Eppendorf tube with the one million of chondrocyte-like cells was used to irradiate the cells and was delineated as the quasi-Gross tumor volume (GTV), defining 1 cm geometrical margins around GTV to create the quasi-Clinical Target Volume (CTV) was done. Total RNA from hiPSCs after chondrogenic differentiation were purified with miRNeasy Kit and RNeasy MinElute according to the manufacturer’s instructions (217004, Quiagen, Hilden, Germany).