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P53/PUMA are potential targets that mediate the protection of brain-derived neurotrophic factor (BDNF)/TrkB from etoposide-induced cell death in neuroblastoma (NB)

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Abstract

The over-expressions of brain-derived neurotrophic factor (BDNF) and its tyrosine kinase receptor TrkB have been reported to induce chemo-resistance in neuroblastoma (NB) cells. In this study, we investigated the roles of P53 and BCL2 family members in the protection of BDNF/TrkB from etoposide-induced NB cell death. TB3 and TB8, two tetracycline (TET)-regulated TrkB-expressing NB cell lines, were utilized. The expressions of P53 and BCL2 family members were detected by Western blot or RT-PCR. Transfection of siRNAs was used to knockdown P53 or PUMA. Activated lentiviral was used to over-express PUMA. Cell survival was performed by MTS assay, and the percentage of cell confluence was measured by IncuCyte ZOOM. Our results showed that etoposide treatment induced significant and time-dependent increase of P53, which could be blocked by pre-treatment with BDNF, and knockdown P53 by transfecting siRNA attenuated etoposide-induced TrkB-expressing NB cell death. PUMA was the most significantly changed BCL2 family member after treatment with etoposide, and pre-treatment with BDNF blocked the increased expression of PUMA. Transfection with siRNA inhibited etoposide-induced increased expression of PUMA, and attenuated etoposide-induced NB cell death. We also found that over-expression of PUMA by infection of activated lentiviral induced TrkB-expressing NB cell death in the absence of etoposide, and treatment of BDNF protected NB cells from PUMA-induced cell death. Our results suggested that P53 and PUMA may be potential targets that mediated the protection of BDNF/TrkB from etoposide-induced NB cell death.

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Abbreviations

BDNF:

Brain-derived neurotrophic factor

TrkB:

Tropomyosin-related kinase B

NB:

Neuroblastoma

MYCN:

Neuroblastoma MYC oncogene

PI3K:

Phosphoinositied-3-kinase

MAPK:

Mitogen-activated protein kinase

HNSCC:

Head and neck squamous cell carcinoma

TET:

Tetracycline

FBS:

Fetal bovine serum

DSBs:

DNA strand breaks

Wip1:

Wild-type p53-inducible phosphatase 1

DLBCL:

Diffuse large B-cell lymphoma

TNBC:

Triple-negative breast cancer

AML:

Acute myeloid leukemia

NSCLC:

Non-small cell lung cancer

FHIT:

Fragile histidine triad

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Acknowledgements

We thank the staff from Medical Research Center of Shengjing Hospital who gave us support throughout the experiments.

Funding

This work was supported by National Natural Science Foundation of China (Nos. 81472359, 81272538), Natural Science Foundation of Liaoning Province (No. 201602855), and 2013 Liaoning Climbing Scholar Foundation.

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Authors and Affiliations

  1. Medical Research Center, Shengjing Hospital of China Medical University, #36 Sanhao Street, Heping District, Shenyang, 110004, China

    Zhongyan Hua, Yue Zhan, Simeng Zhang, Yudi Dong, Min Jiang & Zhijie Li

  2. Department of Neurology, Shengjing Hospital of China Medical University, Shenyang, China

    Fei Tan

  3. Cellular & Molecular Biology Section, Pediatric Oncology Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA

    Zhihui Liu & Carol J. Thiele

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  1. Zhongyan Hua
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  2. Yue Zhan
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Contributions

ZH contributed to the study design, collection and interpretation of data, writing of the manuscript; YZ and SZ contributed to collection of data; YD, and MJ contributed to statistical analyses; FT, ZL, and CJT contributed to the interpretation of the data, and the writing of the manuscript; ZL contributed to the study design, interpretation of the data, the writing of the manuscript, and the submission of the manuscript for publication.

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Correspondence to Zhijie Li.

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Hua, Z., Zhan, Y., Zhang, S. et al. P53/PUMA are potential targets that mediate the protection of brain-derived neurotrophic factor (BDNF)/TrkB from etoposide-induced cell death in neuroblastoma (NB). Apoptosis 23, 408–419 (2018). https://doi.org/10.1007/s10495-018-1467-6

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