Heat Shock Factor 1 Depletion Sensitizes A172 Glioblastoma Cells to Temozolomide via Suppression of Cancer Stem Cell-Like Properties
Abstract
:1. Introduction
2. Results
2.1. BIS Depletion Decreased Protein Levels of HSF1 as Well as Its Nuclear Localization in A172 Glioblastoma Cells under SP-Forming Conditions
2.2. mRNA and Protein Levels of HSF1 Increased in SP-Forming Conditions and HSF1 Knockdown Inhibited SP Formation and Matrix Metalloprotease 2 (MMP2) Activity in A172 Cells
2.3. Transcription of HSF1 Target Genes Was Not Affected by Depletion of BIS or HSF1 in SP-Forming Conditions
2.4. SP A172 Cells Were Less Sensitive to TMZ Treatment Than Cells Cultured in the ML
2.5. HSF1 Depletion Increased TMZ Sensitivity and Was Accompanied by a Decrease in SP Formation and SOX2 Expression
3. Discussion
4. Materials and Methods
4.1. Cell Culture and Small Interfering RNA (siRNA) Transfection
4.2. Sphere-Formation Assay
4.3. Western Blot
4.4. Subcellular Fractionation
4.5. Confocal Microscopy
4.6. Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR)
4.7. Gelatin Zymography
4.8. Cell Viability Assay
4.9. Statistics
5. Conclusions
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
Abbreviations
BIS | Bcl-2 (B-cell lymphoma 2) interacting cell death suppressor |
CSC | Cancer stem cell |
DTT | Dithiothreitol |
HSEs | Heat shock elements |
HSF1 | Heat shock factor 1 |
HSPs | Heat shock proteins |
GBM | Glioblastoma multiforme |
GSCs | Glioblastoma stem cells |
si-CTL | control siRNA |
siRNA | Small interfering RNA |
SP | Sphere |
MMP2 | Matrix metalloprotease 2 |
PARP | Poly(ADP-ribose) polymerase |
SOX2 | Sex determining region Y (SRY)-box 2 |
TMZ | Temozolomide |
MAP1LC3B | Microtubule-associated protein 1 light chain 3 beta |
MCL-1 | Myeloid leukemia cell differentiation protein |
MGMT | O6-methylguanine DNA methyltransferase |
ML | Monolayer |
qRT-PCR | Quantitative real-time polymerase chain reaction |
ROS | Reactive oxygen species |
STAT3 | Signal transducer and activator of transcription 3 |
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Gene | Sequence (5'-3') |
---|---|
BIS | Forward: 5′-GGAATTCGCATGAGCGCCGCCACCCACTCG-3′ |
Reverse: 5′-CTCGAGCTACGGTGCTGCTGGGTTACCAGG-3′ | |
SOX2 | Forward: 5′-TACCTCTTCCTCCCACTCCA-3′ |
Reverse: 5′-ACTCTCCTCTTTTGCACCCC-3′ | |
HSF1 | Forward: 5′-GGCCATGAAGCATGAGAATG -3′ |
Reverse: 5′-GTTCAGCATCAGGGGGATCT-3′ | |
HSPA1A | Forward: 5′-AGCTGGAGCAGGTGTGTAAC-3′ |
Reverse: 5′-CAGCAATCTTGGAAAGGCCC-3′ | |
HSPA8 | Forward: 5′-TAACCCCATCATCAGCGGAC-3′ |
Reverse: 5′-TCCCAACAGTCCACCTCAAAG-3′ | |
HSP27 | Forward: 5′-TGACGGTCAAGACCAAGGAT-3′ |
Reverse: 5′-ATGGTGATCTCGTTGGACTG-3′ | |
MGMT | Forward: 5′-CCGTTTGCGACTTGGTACTT-3′ |
Reverse: 5′-CCTTGCCCAGGAGCTTTATTT-3′ | |
β-ACTIN | Forward: 5′-AGTACTCCGTGTGGATCGGC-3′ |
Reverse: 5′-GCTGATCCACATCTGCTGGA-3′ |
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Im, C.-N.; Yun, H.H.; Lee, J.-H. Heat Shock Factor 1 Depletion Sensitizes A172 Glioblastoma Cells to Temozolomide via Suppression of Cancer Stem Cell-Like Properties. Int. J. Mol. Sci. 2017, 18, 468. https://doi.org/10.3390/ijms18020468
Im C-N, Yun HH, Lee J-H. Heat Shock Factor 1 Depletion Sensitizes A172 Glioblastoma Cells to Temozolomide via Suppression of Cancer Stem Cell-Like Properties. International Journal of Molecular Sciences. 2017; 18(2):468. https://doi.org/10.3390/ijms18020468
Chicago/Turabian StyleIm, Chang-Nim, Hye Hyeon Yun, and Jeong-Hwa Lee. 2017. "Heat Shock Factor 1 Depletion Sensitizes A172 Glioblastoma Cells to Temozolomide via Suppression of Cancer Stem Cell-Like Properties" International Journal of Molecular Sciences 18, no. 2: 468. https://doi.org/10.3390/ijms18020468