ABSTRACT
BACKGROUND: Radiotherapy is essential in the treatment of prostate cancer. An alternative to conventional photon radiotherapy is the application of carbon ions, which provide a superior intratumoral dose distribution and less induced damage to adjacent healthy tissue. A common characteristic of prostate cancer cells is their dependence on androgens which is exploited therapeutically by androgen deprivation therapy in the advanced prostate cancer stage. Here, we aimed to analyze the transcriptomic response of prostate cancer cells to irradiation by photons in comparison to carbon ions, focusing on DNA damage, DNA repair and androgen receptor signaling. METHODS: Prostate cancer cell lines LNCaP (functional TP53 and androgen receptor signaling) and DU145 (dysfunctional TP53 and androgen receptor signaling) were irradiated by photons or carbon ions and the subsequent DNA damage was assessed by immuno-cytofluorescence. Furthermore, the cells were treated with an androgen-receptor agonist. The effects of irradiation and androgen treatment on the gene regulation and the transcriptome were investigated by RT-qPCR and RNA sequencing, followed by bioinformatic analysis. RESULTS: Following photon or carbon ion irradiation, both LNCaP and DU145 cells showed a dose-dependent amount of visible DNA damage that decreased over time, indicating occurring DNA repair. In terms of gene regulation, mRNAs involved in the TP53-dependent DNA damage response were significantly upregulated by photons and carbon ions in LNCaP but not in DU145 cells, which generally showed low levels of gene regulation after irradiation. Both LNCaP and DU145 cells responded to photons and carbon ions by downregulation of genes involved in DNA repair and cell cycle, partially resembling the transcriptome response to the applied androgen receptor agonist. Neither photons nor carbon ions significantly affected canonical androgen receptor-dependent gene regulation. Furthermore, certain genes that were specifically regulated by either photon or carbon ion irradiation were identified. CONCLUSION: Photon and carbon ion irradiation showed a significant congruence in terms of induced signaling pathways and transcriptomic responses. These responses were strongly impacted by the TP53 status. Nevertheless, irradiation mode-dependent distinct gene regulations with undefined implication for radiotherapy outcome were revealed. Androgen receptor signaling and irradiations shared regulation of certain genes with respect to DNA-repair and cell-cycle.
Subject(s)
Photons , Prostatic Neoplasms , Receptors, Androgen , Signal Transduction , Transcriptome , Tumor Suppressor Protein p53 , Humans , Male , Prostatic Neoplasms/radiotherapy , Prostatic Neoplasms/metabolism , Prostatic Neoplasms/pathology , Receptors, Androgen/metabolism , Receptors, Androgen/genetics , Tumor Suppressor Protein p53/metabolism , Transcriptome/radiation effects , Signal Transduction/radiation effects , DNA Damage/radiation effects , Heavy Ion Radiotherapy , DNA Repair , Gene Expression Regulation, Neoplastic/radiation effects , Gene Expression Regulation, Neoplastic/drug effects , Cell Line, Tumor , Carbon/pharmacologyABSTRACT
In skeletal muscle tissue engineering, innervation and vascularization play an essential role in the establishment of functional skeletal muscle. For adequate three-dimensional assembly, biocompatible aligned nanofibers are beneficial as matrices for cell seeding. The aim of this study was to analyze the impact of Schwann cells (SC) on myoblast (Mb) and adipogenic mesenchymal stromal cell (ADSC) cocultures on poly-É-caprolactone (PCL)-collagen I-nanofibers in vivo. Human Mb/ADSC cocultures, as well as Mb/ADSC/SC cocultures, were seeded onto PCL-collagen I-nanofiber scaffolds and implanted into the innervated arteriovenous loop model (EPI loop model) of immunodeficient rats for 4 weeks. Histological staining and gene expression were used to compare their capacity for vascularization, immunological response, myogenic differentiation, and innervation. After 4 weeks, both Mb/ADSC and Mb/ADSC/SC coculture systems showed similar amounts and distribution of vascularization, as well as immunological activity. Myogenic differentiation could be observed in both groups through histological staining (desmin, myosin heavy chain) and gene expression (MYOD, MYH3, ACTA1) without significant difference between groups. Expression of CHRNB and LAMB2 also implied neuromuscular junction formation. Our study suggests that the addition of SC did not significantly impact myogenesis and innervation in this model. The implanted motor nerve branch may have played a more significant role than the presence of SC.
