The Cytotoxic Effects of PTEN-mRNA-engineered Mesenchymal Stem Cell on U251 Glioma Cells

doi:10.13560/j.cnki.biotech.bull.1985.2016.04.032

Abstract

Abstract: Mesenchymal stem cells（MSCs）have been considered of great potential as ideal carriers for the delivery of anticancer agents since the discovery of their capacity of tumor-oriented migration and integration. Differing from DNA-based vectors，synthesized mRNA in vitro owns the properties of its easy transfection and mutagenesis-free. This study aims to investigate the effects of phosphatase and tensin homolog（PTEN）mRNA-engineered MSCs on human glioma U251 cells under indirect co-culture condition. PTEN mRNA by in vitro transcription was transfected into MSCs，and then the expression of protein PTEN in the transfected MSCs was detected by Western blot，and the migration and integration effects of PTEN mRNA transfection on MSC tumor cells were verified using transwell co-cultures. The cytotoxic effects of PTEN mRNA-engineered MSCs on the U251-Luc glioma cells were detected with luminescence and fluorescence microscopy under indirect co-culture condition. The in vitro synthesized PTEN mRNA was transfected to U251-Luc cells，protein PTEN expressed correctly and the expression was the highest at 24 hours after transfection. An enhanced migration rate was observed with MSCs transfected with PTEN mRNA compared to non-transfected MSCs（P<0.05）. A significant inhibition of U251 cells was observed when they were cultured with conditioned medium from PTEN mRNA-engineered MSCs（P<0.05）. The results suggest that anticancer gene-bearing mRNA synthesized in vitro provides new insights into the MSC-mediated targeted gene therapy of glioblastoma.

Key words: mesenchymal stem cells, synthesized mRNA, PTEN, gene therapy, U251 glioma cell

GUO Xing-rong, WANG Xiao-li, YUAN Ya-hong ,TU Han-jun. The Cytotoxic Effects of PTEN-mRNA-engineered Mesenchymal Stem Cell on U251 Glioma Cells[J]. Biotechnology Bulletin, 2016, 32(4): 234-241.

References

［1］Van Meir EG, Hadjipanayis CG, Norden AD, et al. Exciting new advances in neuro-Oncology the avenue to a cure for malignant glioma［J］. CA Cancer J Clin, 2010, 60（3）：166-193.
［2］Westphal M, Lamszus K. The neurobiology of gliomas：from cell biology to the development of therapeutic approaches［J］. Nat Rev Neurosci, 2011, 12（9）：495-508.
［3］Ferguson SD. Malignant gliomas：diagnosis and treatment［J］. Neuro Surgery, 2011, 57（11）：558-569.
［4］Loebinger MR, Janes SM. Stem cells as vectors for antitumour therapy［J］. Thorax, 2010, 65（4）：362-369.
［5］Dai LJ. Potential implications of mesenchymal stem cells in cancer therapy［J］. Cancer Lett, 2011, 305（5）：8-20.
［6］Buono R, Abrate A, Esposito A, et al. Mesenchymal stem cell mediated cancer therapy inhibits tumor growth in the transgenic adenocarcinoma of the mouse prostate（Tramp）model［J］. J Urology, 2013, 189（5）：203-207.
［7］Zhang X, Zhang L, Xu W, et al. Experimental therapy for lung cancer：umbilical cord-Derived mesenchymal stem cell-mediated interleukin-24 delivery［J］. Curr Cancer Drug Tar, 2013, 13（1）：92-102.
［8］Leonhardt C, Schwake G, St?gbauer TR, et al. Single-cell mRNA transfection studies：Delivery, kinetics and statistics by numbers［J］. Nanomed-Nanotechnol, 2014, 10（4）：679-688.
［9］Wang XL, Hu p, Guo XR, et al. Reprogramming human umbilical cord mesenchymal stromal cells to islet-like cells with the use of in vitro-synthesized pancreatic-duodenal homebox 1 messenger RNA［J］. Cytotherapy, 2014, 16（11）：1519-1527.
［10］Guo XR, Wang XL, Li MC, et al. PDX-1 mRNA-induced reprogramming of mouse pancreas-derived mesenchymal stem cells into insulin-producing cells in vitro［J］. Clin Exp Med, 2014, 10（1）：152-160.
［11］ Warren L, Manos PD, Ahfeldt T, et al. Highly efficient reprogram-ming to pluripotency and directed differentiation of human cells with synthetic modified mRNA［J］. Cell Stem Cell, 2010, 7（5）：618-630.
［12］ Zangi L, Lui KO, von Gise A, et al. Modified mRNA directs the fate of heart progenitor cells and induces vascular regeneration after myocardial infarction［J］. Nat Biotechnol, 2013, 31（10）：898-907.
［13］Liu W, Zhou Y, et al. PTEN mutation：many birds with one stone in tumorigenesis［J］. Anticancer Res, 2008, 28（6A）：3613-3619.
［14］Ciuffreda L, Falcone I, Incani UC, et al. PTEN expression and function in adult cancer stem cells and prospects for therapeutic targeting［J］. Adv Biol Relat, 2014, 56：66-80.
［15］Muniyan S, Ingersoll MA, Batra SK, et al. Cellular prostatic acid phosphatase, a PTEN-functional homologue in prostate epithelia, functions as a prostate-specific tumor suppressor［J］. BBA, 2014, 1846（1）：88-98.
［16］ Chalhoub N, Baker SJ. PTEN and the PI3-kinase pathway in cancer［J］. Annu Rev Pathol, 2009, 4（11）：127-150.
［17］Moniri MR, Sun XY, Rayat J, et al. TRAIL-engineered pancreas-derived mesenchymal stem cells：characterization and cytotoxic effects on pancreatic cancer cells［J］. Cancer Gene Ther, 2012, 19（9）：652-658.
［18］Yang ZS, Tang XJ, Guo XR, et al. Cancer cell-oriented migration of mesenchymal stem cells engineered with an anticancer gene（PTEN）：an imaging demonstration［J］. Oncotargets Ther, 2014, 7：441-446.
［19］ Nakamizo A, Marini F, Amano T, et al. Human bone marrow-derived mesenchymal stem cells in the treatment of gliomas［J］. Cancer Res, 2005, 65（12）：3307-3318.
［20］ Menon LG, Kelly K, Yang HW, et al. Human bone marrow-derived mesenchymal stromal cells expressing S-TRAIL as a cellular delivery vehicle for human glioma therapy［J］. Stem Cells, 2009, 27（28）：2320-2330.
［21］ Dwyer RM, Khan S, Barry FP, et al. Advances in mesenchymal stem cell-mediated gene therapy for cancer［J］. Stem Cell Res Ther, 2010, 22（6）：2012-2018.
［22］ Knoop K, Kolokythas M, Klutz K, et al. Image-guided, tumor stroma-targeted I-131 therapy of hepatocellular cancer after systemic mesenchymal stem cell-mediated NIS gene delivery［J］. Mol Ther, 2011, 19：1704-1713.
［23］ Li M, Sancho-Martinez I, Belmonte J C, et al. Cell fate conversion by mRNA［J］. Stem Cell Res Ther, 2011, 15（11）：2-5.
［24］ Kormann MS, Hasenpusch G, Aneja MK, et al. Expression of therapeutic proteins after delivery of chemically modified mRNA in mice［J］. Nat Biotechnol, 2011, 29（2）：154-157.
［25］ Neyns B, Heirman C, Thielemans K, et al. Immunotherapy of cancer with dendritic cells loaded with tumor antigens and activated through mRNA electroporation［J］. Methods Mol Biol, 2010, 629（3）：405-452.
［26］Leslie NR, Downes CP. PTEN function：how normal cells control it and tumour cells lose it［J］. Biochem J, 2004, 382（Pt 1）：1-11.
［27］Benjamin DH, Fine B, Steinbach N, et al. A secreted PTEN phosphatase that enters cells to alter signaling and survival［J］. Science, 2013, 341（6411）：399-402.
［28］ Roshan MM, Young A, Reinheimer K, et al. Dynamic assessment of cell viability, proliferation and migration using real time cell analyzer system（RTCA）［J］. Cytotechnology, 2014, 67（2）：379-386.

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