TNFR, TRAF2, NF-κB mRNA Levels of Glioblastoma Multiforme Cells Treated by Conditioned Medium of Umbilical Cord-derived Mesenchymal Stem Cells

Novi Silvia Hardiany, Yohana Yohana, Septelia Inawati Wanandi


BACKGROUND: Glioblastoma multiforme (GBM) is a human malignant brain tumor which is arise from glial cells. Our previous study proved that GBM cells proliferation increased after treating by conditioned medium of umbilical cord-derived mesenchymal stem cells (CM-UCSCs). Cells proliferation is probably mediated by tumor necrosis factor (TNF)-α which could bind to membrane receptor and induce signaling pathway. Therefore, this research was intended to analyze the mRNA expression of TNF-α signaling pathway molecules on CM-treated GBM cells by measuring TNF receptor 1 and 2 (TNFR1 and TNFR2), TNFR associated factor 2 (TRAF2), nuclear factor kappa B (NF-κB) mRNA level, and TNFR2 protein level.

METHODS: UCSCs and human glioblastoma T98G cells were cultured and harvested after 80% confluence. CM was prepared by growing UCSCs in serum alpha Minimum Essential Media (α-MEM) for 24 hours. Fifty percent concentration of CM-UCSCs was used to treat T98G cells for 24 hours. TNF-α level in CM-UCSC was detected using enzyme linked-immunosorbent assay (ELISA), while the expression of TNFR1, TNFR2, TRAF2 and NF-κB were detected using quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR), and TNFR2 protein level was detected using sandwich ELISA.

RESULTS: TNF-α level was detected in CM-UCSCs 4.4 pg/mL. Moreover, the expression of TNFR1, TNFR2, TRAF2 and NF-κB were significantly 1.4-fold, 4.9-fold, 5.6-fold, 1.8-fold respectively higher in T98G treated cells than control. TNFR2 protein level in T98G treated cells was 11.57 pg/mg protein higher than control.

CONCLUSION: The expression of molecules involved in TNF-α signaling pathway were up regulated in T98G cells treated by CM-UCSCs.


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Hanif F, Muzaffar K, Perveen K, Malhi SM, Simjee SU. Glioblastoma multiforme : A review of its epidemiology and pathogenesis through clinical presentation and treatment. Asian Pac J Cancer P. 2017; 18: 3-9, PMID.

Iser IC, Ceschini SM, Onzi GR, Bertoni APS, Lens Guido, Wink MR. Conditioned medium from adipose-derived stem cells (ADSCs) promotes conditioned medium from adipose-derived stem cells (ADSCs) promotes epithelial-to-mesenchymal-like transition (EMT-Like ) in glioma cells in vitro. Mol Neurobiol. 2016; 53: 7184-99, CrossRef.

Koontongkaew S. The tumor microenvironment contribution to development, growth, invasion and metastasis of head and neck squamous cell carcinomas. J Cancer. 2013; 4: 66-83, PMID.

Akimoto K, Kimura K, Nagano M, Takano S, To’a Salazar G, Yamashita T, et al. Umbilical cord blood-derived mesenchymal stem cells inhibit, but adipose tissue-derived mesenchymal stem cells promote, glioblastoma multiforme proliferation. Stem Cells Dev. 2013; 22: 1370-86, CrossRef.

Vieira de Castro J, Gomes ED, Grania S, Anjo SI, Baltazar F, Manadas B, et al. Impact of mesenchymal stem cells’ secretome on glioblastoma pathophysiology. J Transl Med. 2017; 15: 200, CrossRef.

Cunningham CJ, Redondo-castro E, Allan SM. The therapeutic potential of the mesenchymal stem cell secretome in ischaemic stroke. J Cereb Blood Flow Metab. 2018; 38: 1276-92, CrossRef.

Ham B, Fernandez MC, Costa ZD, Brodt P. The diverse roles of the TNF axis in cancer progression and metastasis. Trends Cancer Res. 2016; 11: 1-27, PMID.

Martínez-Reza I, Díaz L, García-Becerra R. Preclinical and clinical aspects of TNF-α and its receptors TNFR1 and TNFR2 in breast cancer. J Biomed Sci. 2017; 24: 90, CrossRef.

Chopra M, Lang I, Salzmann S, Pachel C, Kraus S, Bauerlein CA, et al. Tumor necrosis factor induces tumor promoting and anti-tumoral effects on pancreatic cancer via TNFR1. Plos One. 2013; 8: e75737, CrossRef.

Cai X, Cao C, Li J, Chen F, Zhang S, Liu B, et al. Inflammatory factor TNF- α promotes the growth of breast cancer via the positive feedback loop of TNFR1/NF- κ B and/or p38. Oncotarget. 2017; 8: 58338-52, CrossRef.

Hardiany NS, Huang P, Dewi S, Paramita R, Wanandi SI. Analysis of pluripotency marker expression in human glioblastoma multiforme cells treated with conditioned medium of umbilical cord-derived mesenchymal stem cells. F1000Research. 2018; 7: 106, CrossRef.

Hardiany NS, Yohana, Wanandi SI. The impact of conditioned medium of umbilical cord-derived mesenchymal stem cells toward apoptosis and proliferation of glioblastoma multiforme cells. IOP Conf Ser Earth Environ Sci. 2019; 217: 012035, CrossRef.

Hardiany NS, Sadikin M, Siregar N, Wanandi SI. The suppression of manganese superoxide dismutase decreased the survival of human glioblastoma multiforme T98G cells. Med J Indones. 2017; 26: 19-25, CrossRef.

Purnamawati P, Pawitan JA, Rachman A, Wanandi SI. Effects of umbilical cord- and adipose-derived stem cell secretomes on ALDH1A3 expression and autocrine TGF-β1 signaling in human breast cancer stem cells. F1000Research. 2018; 7: 249, CrossRef.

Zhang K, Li QI, Kang X, Wang Y, Wang S. Identification and functional characterization of lncRNAs acting as ceRNA involved in the malignant progression of glioblastoma multiforme. Oncology Report. 2016; 36: 2911-25, CrossRef.

Wei B, Liang J, Hu J, Mi Y, Ruan J, Zhang J, et al. TRAF2 is a valuable prognostic biomarker in patients with prostate cancer. Med Sci Monit. 2017; 23: 4192-204, CrossRef.

Fashi M, Agha-alinejad H, Mahabadi HA, Rezaei B, Pakrad B, Rezaei S. The Effects of aerobic exercise on NF-κB and TNF-α in lung tissue of male rat. Novel Biomed. 2015; 3: 131-4, CrossRef.

Hardiany NS, Mulyawan W, Wanandi SI. Correlation between oxidative stress and tumor grade in glioma cells from patients in Jakarta. Med J Indones. 2012; 21: 122-9, CrossRef.

Mehling BM, Manvelyan M, Benesh G, Wu DC. Characterization of human umbilical cord mesenchymal stem cells-derived conditioned medium. J Stem Cell Res Ther. 2016; 1: 218-20, article.

Ten Hagen TL, Van Der Veen AH, Nooijen PT, Van Tiel ST, Seynhaeve AL, Egermont AM. Low dose tumor necrosis-alpha augments antitumor activity of stealth liposomal doxorubicin (DOXIL) in soft tissue sarcoma-bearing rats. Int J Cancer. 2000; 87: 829-37, CrossRef.

Bajetto A, Pattarozzi A, Corsaro A, Barbieri F, Daga A, Bosio A, et al. Different effects of human umbilical cord mesenchymal stem cells on glioblastoma stem cells by direct cell interaction or via released soluble factors. Front Cell Neurosci. 2017; 11: 312, CrossRef.

Dong L, Pu Y, Zhang L, Qi Q, Xu L, Li W, et al. Human umbilical cord mesenchymal stem cell-derived extracellular vesicles promote lung adenocarcinoma growth by transferring miR-410. Cell Death Dis. 2018; 9: 218, CrossRef.

Zhao P, Zhang Z. TNF-α promotes colon cancer cell migration and invasion by upregulating TROP-2. Oncol Lett. 2018; 15: 3820-7, CrossRef.

Chowdhury I, Bhat GK. Tumor Necrosis Factor (TNF) – From Bench to Bed Side. New York: Nova Science Publisher; 2009.

Puliyappadamba VT, Chakraborty S, Hatanpaa KJ, Habib AA. The role of NF-κB in the pathogenesis of glioma. Mol Cell Oncol. 2014; 1: e963478, CrossRef.

Rinkenbaugh AL, Cogswell PC, Calamini B, Dunn DE, Persson AI, Weiss WA, et al. IKK/NF-κB signaling contributes to glioblastoma stem cell maintenance. Oncotarget. 2016; 7: 69173-87, CrossRef.

Friedmann-morvinski D, Narasimamurthy R, Xia Y, Myskiw C, Soda Y, Verma IM. Targeting NFκB in glioblastoma : a therapeutic approach. Sci Adv. 2016; 2: e1501292, CrossRef.

Hu X, Li B, Li X, Zhao X, Wan L, Lin G, et al. Transmembrane TNFα promotes suppressive activities of myeloid-derived suppressor cells via TNFR2. J Imunol. 2018; 192: 1320-31, CrossRef.

Nubel T, Chmitt SS, Aina BK, Fritz G. Lovastatin stimulates p75 TNF receptor (TNFR2) expression in primary human endothelial cells. Int J Mol Med. 2005; 27: 1139-45, CrossRef.


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