Highly aggressive cancers, such as glioblastoma multiforme (GBM), use efficient cellular communication to develop resistance to therapy or to grow invasively. Tunneling nanotubes (TNTs), which are tiny membrane tunnels with diameters in the nanometer range, serve as effective communication channels for the exchange of signals and cargo between cells. TNTs enable the formation of extensive communication networks, contributing to rapid adaptability. However, the role of TNTs in cancer is still poorly understood. This study aimed to characterize TNTs in the GBM cell lines U87 MG and LN229 and to investigate their response to therapeutic interventions. To study these tiny structures and observe TNT network development in living cells after treatment, we developed and performed high-resolution confocal and stimulated emission depletion (STED) microscopy on living cells. The properties of the TNTs in both cell lines were characterized microscopically, including size, morphology, lifetime, formation, and cytoskeletal content. U87 MG cells had thinner (approximately 197 nm), longer (approximately 40 µm), and more stable (mean lifetime: approximately 88 min) TNTs than LN229 cells (mean diameter: approximately 338 nm; mean length: approximately 20 µm; mean lifetime: approximately 41 min). TNTs were formed exclusively by cell movement and were highly sensitive to standard fixation methods, highlighting the necessity of live cell imaging. Low-LET X-ray irradiation (1.79 Gy) led to a significant increase in the connectivity and complexity of TNT networks approximately ten hours after irradiation, indicating a stimulation of cellular communication along TNTs. In contrast, the standard chemotherapeutic agents temozolomide (TMZ) and cytarabine (AraC) showed no significant effect on TNT networks in U87 MG cells. However, the actin polymerization inhibitor cytochalasin B significantly inhibited TNT formation, confirming the importance of actin for TNTs. In a subsequent study, the migration behavior of the two cell lines was examined after irradiation with different types of radiation (X-rays, protons, and carbon ions) with and without spatial shielding. Depending on the irradiation conditions, cell migration was influenced differently in terms of speed and orientation. The results suggest that cell orientation may be more important than cell velocity for invasiveness. Additionally, partial irradiation induced altered migration in shielded cells compared to sham-irradiated cells, indicating bystander effects. In addition, the transfer of calcium and mitochondria by TNTs has been observed. Both could contribute to therapy resistance development. Furthermore, initial steps were carried out to perform co-culture experiments of non-irradiated and irradiated cells, to establish the study of non-targeted effects caused by cellular communication along TNTs. Here, the transfer of a cytoplasmic dye along TNTs was observed. In conclusion, this dissertation provides a detailed characterization of TNTs in GBM cells, revealing their dynamic nature and differential response to radiation and chemotherapy. The results suggest that communication via TNTs is enhanced by radiation, potentially contributing to radioresistance and influencing migration behavior. In contrast, chemotherapy does not affect TNT-mediated communication. This identifies cellular communication along TNTs as a critical component of GBM therapy and as a potential novel therapeutic target.     «

Highly aggressive cancers, such as glioblastoma multiforme (GBM), use efficient cellular communication to develop resistance to therapy or to grow invasively. Tunneling nanotubes (TNTs), which are tiny membrane tunnels with diameters in the nanometer range, serve as effective communication channels for the exchange of signals and cargo between cells. TNTs enable the formation of extensive communication networks, contributing to rapid adaptability. However, the role of TNTs in cancer is still poo...     »