Merlin-Deficient Schwann Cells Are More Susceptible to Radiation Injury than Normal Schwann Cells In Vitro

Abstract

Objectives Vestibular schwannomas (VS) are intracranial tumors, which are caused by NF2 gene mutations that lead to loss of merlin protein. A treatment for VS is stereotactic radiosurgery, a form of radiation. To better understand the radiobiology of VS and radiation toxicity to adjacent structures, our main objectives were (1) investigate effects of single fraction (SF) radiation on viability, cytotoxicity, and apoptosis in normal Schwann cells (SCs) and merlin-deficient Schwann cells (MD-SCs) in vitro, and (2) analyze expression of double strand DNA breaks (γ-H2AX) and DNA repair protein Rad51 following irradiation. Study Design This is a basic science study. Setting This study is conducted in a research laboratory. Participants Patients did not participate in this study. Main Outcome Measures In irradiated normal SCs and MD-SCs (0–18 Gy), we measured (1) viability, cytotoxicity, and apoptosis using cell-based assays, and (2) percentage of cells with γ-H2AX and Rad51 on immunofluorescence. Results A high percentage of irradiated MD-SCs expressed γ-H2AX, which may explain the dose-dependent losses in viability in rodent and human cell lines. In comparison, the viabilities of normal SCs were only compromised at higher doses of radiation (>12 Gy, human SCs), which may be related to less Rad51 repair. There were no further reductions in viability in human MD-SCs beyond 9 Gy, suggesting that <9 Gy may be insufficient to initiate maximal tumor control. Conclusion The MD-SCs are more susceptible to radiation than normal SCs, in part through differential expression of γ-H2AX and Rad51. Understanding the radiobiology of MD-SCs and normal SCs is important for optimizing radiation protocols to maximize tumor control while limiting radiation toxicity in VS patients.

Primary Vestibular Schwannoma Cells Activate p21 and RAD51-Associated DNA Repair Following Radiation-Induced DNA Damage

“…RAD51-associated DNA repair in response to radiation has been studied in normal human Schwann cells and merlin-deficient Schwann cells (MDSC). In a recent in vitro investigation by Cohen et al, human MDSCs exhibited high levels of g-H2AX 6 h after exposure to 6, 12, and 18 Gy of single fraction radiation (18). Although the levels of g-H2AX were similar across the three radiation groups, MDSCs demonstrated a dose-dependent loss in cell viability at 96 h. The authors show that a lower dose of radiation (i.e., 6 Gy), MDSCs can initiate a robust RAD51 DNA repair response to evade cell loss.…”

“…A recent in vitro study showed that merlin-deficient Schwann cells (MD-SCs) displayed upregulation of the DNA repair protein RAD51 in response to IR-induced DNA damage when compared to normal human Schwann cells (18). However, little is known about the activity of RAD51 in response to radiation-induced DSBs in VS.…”

Understanding the Radiobiology of Vestibular Schwannomas to Overcome Radiation Resistance

“…Other tumors can evade radiation injury by upregulating DNA repair proteins, such as RAD51, DNA-PK [ 160 , 165 ], RPA1 [ 166 ], LIG4 [ 167 ], HIF-1 [ 168 ], HDAC [ 169 ], Wee1 [ 170 ], CDK1 [ 171 ], and Chk1 [ 160 , 165 , 172 , 173 , 174 ]. In an in vitro study using normal human Schwann cells and merlin-deficient Schwann cells (MD-SCs), MD-SCs produced a robust RAD51 response when exposed to 6 Gy of radiation when compared to normal Schwann cells [ 175 ]. These findings suggest a possible mechanism of radiation resistance in schwannomas that warrants further investigation.…”

“…Although the molecular mechanisms for why faster-growing tumors would be less responsive to radiation are unknown, it was proposed that fast-growing tumors may be more radioresistant because they have efficient DNA repair mechanisms [ 30 ]. In an in vitro study, Cohen et al found that MD-SCs had a more robust upregulation of DNA repair protein RAD51 after exposure to 6 Gy of radiation than normal Schwann cells, suggesting that VS may upregulate DNA repair mechanisms in order to resist radiation injury [ 175 ]. However, at higher doses of radiation (12 and 18 Gy single fraction), merlin-deficient Schwann cells did not activate RAD51 more than baseline, suggesting that higher doses of radiation may be required to prevent activation of DNA repair proteins.…”