Characterization of Temozolomide Resistance Using a Novel Acquired Resistance Model in Glioblastoma Cell Lines
Abstract
:Simple Summary
Abstract
1. Introduction
2. Materials and Methods
2.1. Cell Culture, Generation of TMZ Resistant GBM Variants and Drug Treatment
2.2. Generation of GBM-RG Cells Expressing a 53BP1-Foci Reporter and Analysis of TMZ Induced Double-Strand Breaks (DSBs)
2.3. MTT Assay
2.4. Flow Cytometry Detection of Cell Cycle and EdU Incorporation
2.5. Apoptosis Assay by Nuclear Staining and Annexin V/PI Detection
2.6. Colony Formation Assay
2.7. Western Blot
2.8. RT2-PCR
2.9. Neurosphere Assay
2.10. Statistics
3. Results
3.1. RG2 Cells Exhibit Resistance to Chemotherapy and Higher Regrowth Capacity in Post-Treatment Phase
3.2. RG2 Cells Exhibit Enhanced Clonogenic Growth Capacity Accompanied by Higher Proliferation Activity in the Post-Treatment Phase
3.3. RG2 Cells Show Less DNA Damage in Individual Cells after TMZ Treatment in Association with the Downregulation of MMR Gene Expression
3.4. RG2 Cells Show a Reduction in Cell Cycle Arrest at G2/M Accompanied by an Increase in DNA Replication after TMZ Treatment
3.5. Apoptosis Is Involved in the Chemoresistance of U373-RG2 and LN229-RG2 Cells via Different Pathways
3.6. U373 RG2 Cells Exhibit Enhanced Self-Renewal Capacity in Association with Upregulation of Stem Cell Markers
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Primer Name | Sequence | Annealing Temperature (°C) |
---|---|---|
MSH2 | 60 | |
for. | TTTACCCGGAGGAGAGACTGC | |
rev. | TGCTCTCCCTTTTTGCCTTTC | |
MSH6 | 60 | |
for. | AGAGCAATGCAACGTGCAGA | |
rev. | TTTGGCGGCTACTTCGCCTA | |
CD133 | 60 | |
for. | CAGAAGGCATATGAATCC | |
rev. | CACCACATTTGTTACAGC | |
CD44 | 60 | |
for. | AGCACAATCCAGGCAACTCC | |
rev. | CTGGTATGAGCTGAGGCTGC | |
SOX2 | 60 | |
for. | ACCGGCGGCAACCAGAAGAACAG | |
rev. | GCGCCGCGGCCGGTATTTAT | |
GAPDH | 60 | |
for. | TCACCACCATGGAGAAGGC | |
rev. | GCTAAGCAGTTGGTGGTGCA | |
RPS13 | 60 | |
for. | CGAAAGCATCTTGAGAGGAACA | |
rev. | TCGAGCCAAACGGTGAATC |
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Zhu, Y.; Chen, Z.; Kim, S.N.; Gan, C.; Ryl, T.; Lesjak, M.S.; Rodemerk, J.; Zhong, R.D.; Wrede, K.; Dammann, P.; et al. Characterization of Temozolomide Resistance Using a Novel Acquired Resistance Model in Glioblastoma Cell Lines. Cancers 2022, 14, 2211. https://doi.org/10.3390/cancers14092211
Zhu Y, Chen Z, Kim SN, Gan C, Ryl T, Lesjak MS, Rodemerk J, Zhong RD, Wrede K, Dammann P, et al. Characterization of Temozolomide Resistance Using a Novel Acquired Resistance Model in Glioblastoma Cell Lines. Cancers. 2022; 14(9):2211. https://doi.org/10.3390/cancers14092211
Chicago/Turabian StyleZhu, Yuan, Zhen Chen, Su Na Kim, Chao Gan, Tatsiana Ryl, Michaela Silvia Lesjak, Jan Rodemerk, Rong De Zhong, Karsten Wrede, Philipp Dammann, and et al. 2022. "Characterization of Temozolomide Resistance Using a Novel Acquired Resistance Model in Glioblastoma Cell Lines" Cancers 14, no. 9: 2211. https://doi.org/10.3390/cancers14092211