Molecular Biology Reports

, Volume 47, Issue 2, pp 1187–1197 | Cite as

Effects of temozolomide on U87MG glioblastoma cell expression of CXCR4, MMP2, MMP9, VEGF, anti-proliferatory cytotoxic and apoptotic properties

  • Seyedsaber Mirabdaly
  • Daniel Elieh Ali Komi
  • Yadollah Shakiba
  • Ali Moini
  • Amir KianiEmail author
Original Article


Temozolomide is an alkylating agent which is used in glioblastoma treatment. We aimed to investigate the effects of different concentrations of temozolomide and exposure time on U87MG glioblastoma cell expression of CXCR4, MMP2, MMP9 and VEGF. U87MG cells were cultured in different temozolomide concentrations and incubation time and the effects of temozolomide on inducing apoptosis was investigated. The levels of VEGF and CXCR4 expression were measured by RT-PCR and flowcytometry. Moreover, MMP2 and MMP9 activity and expression were assessed by ELISA and zymography. CXCR4 and VEGF expression levels decreased upon applying higher concentration of temozolomide. MMP2 and MMP-9 had lower activity in cells with longer exposure time or higher doses of temozolomide. Temozolomide induces the apoptosis in U87MG glioblastoma cells at therapeutic or higher dose. It is capable of decreasing their expression levels of VEGF and CXCR4.


CXCR4 MMP2 MMP9 Temozolomide U87MG VEGF 



Primary brain tumor


Central nervous system




Blood brain barrier



This work was performed in partial fulfillment of the requirements for, Pharm. Dr. of Seyedsaber Mirabdaly in the faculty of Pharmacy, Kermanshah University of Medical Sciences, Kermanshah, Iran. The authors gratefully acknowledge the Research Council of Kermanshah University of Medical Sciences (Grant Number: 90076) for the financial support.

Author contributions

All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Seyedsaber Mirabdaly, Daniel Elieh Ali Komi and Amir Kiani. The first draft of the manuscript was written by Amir Kiani and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.


This work was supported by Research Council of Kermanshah University of Medical Sciences.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.


  1. 1.
    Yuan D, Tao Y, Chen G, Shi T (2019) Systematic expression analysis of ligand-receptor pairs reveals important cell-to-cell interactions inside glioma. Cell Commun Signal 17(1):48. CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Chen R, Smith-Cohn M, Cohen AL, Colman H (2017) Glioma subclassifications and their clinical significance. Neurotherapeutics 14(2):284–297. CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Wirsching HG, Weller M (2016) The role of molecular diagnostics in the management of patients with gliomas. Curr Treat Opt Oncol 17(10):51. CrossRefGoogle Scholar
  4. 4.
    Berger G, Grauwet K, Zhang H, Hussey AM, Nowicki MO, Wang DI, Chiocca EA, Lawler SE, Lippard SJ (2018) Anticancer activity of osmium(VI) nitrido complexes in patient-derived glioblastoma initiating cells and in vivo mouse models. Cancer Lett 416:138–148. CrossRefPubMedGoogle Scholar
  5. 5.
    Wang J, Hu G, Quan X (2019) Analysis of the factors affecting the prognosis of glioma patients. Open Med (Warsaw, Poland) 14:331–335. CrossRefGoogle Scholar
  6. 6.
    Grauwet K, Chiocca EA (2016) Glioma and microglia, a double entendre. Nat Immunol 17(11):1240–1242. CrossRefPubMedGoogle Scholar
  7. 7.
    Alentorn A, Duran-Pena A, Pingle SC, Piccioni DE, Idbaih A, Kesari S (2015) Molecular profiling of gliomas: potential therapeutic implications. Expert Rev Anticancer Ther 15(8):955–962. CrossRefPubMedGoogle Scholar
  8. 8.
    Rao JU, Coman D, Walsh JJ, Ali MM, Huang Y, Hyder F (2017) Temozolomide arrests glioma growth and normalizes intratumoral extracellular pH. Sci Rep 7(1):7865. CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Li W, Zhang R, Yang J, Wang R (2017) Efficacy and prognosis of surgery combined with (125)I seed implantation in treatment of recurrent glioma. Oncol Lett 14(6):7201–7206. CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Walsh KM, Claus EB (2019) Diet and risk of glioma: targets for prevention remain elusive. Neuro-oncology. CrossRefPubMedGoogle Scholar
  11. 11.
    Richardson PJ (2016) CXCR4 and glioblastoma. Anticancer Agents Med Chem 16(1):59–74CrossRefGoogle Scholar
  12. 12.
    Yadav VN, Zamler D, Baker GJ, Kadiyala P, Erdreich-Epstein A, DeCarvalho AC, Mikkelsen T, Castro MG, Lowenstein PR (2016) CXCR4 increases in vivo glioma perivascular invasion, and reduces radiation induced apoptosis: a genetic knockdown study. Oncotarget 7(50):83701–83719. CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Li G, Xie Q, Yang Z, Wang L, Zhang X, Zuo B, Zhang S, Yang A, Jia L (2019) Sp1-mediated epigenetic dysregulation dictates HDAC inhibitor susceptibility of HER2-overexpressing breast cancer. Int J Cancer. CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Zheng F, Zhang Z, Flamini V, Jiang WG, Cui Y (2017) The axis of CXCR4/SDF-1 plays a role in colon cancer cell adhesion through regulation of the AKT and IGF1R signalling pathways. Anticancer Res 37(8):4361–4369. CrossRefPubMedGoogle Scholar
  15. 15.
    Gagner JP, Sarfraz Y, Ortenzi V, Alotaibi FM, Chiriboga LA, Tayyib AT, Douglas GJ, Chevalier E, Romagnoli B, Tuffin G, Schmitt M, Lemercier G, Dembowsky K, Zagzag D (2017) Multifaceted C-X-C chemokine receptor 4 (CXCR4) inhibition interferes with anti-vascular endothelial growth factor therapy-induced glioma dissemination. Am J Pathol 187(9):2080–2094. CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Liu H, Zeng Z, Wang S, Li T, Mastriani E, Li QH, Bao HX, Zhou YJ, Wang X, Liu Y, Liu W, Hu S, Gao S, Yu M, Qi Y, Shen Z, Wang H, Gao T, Dong L, Johnston RN, Liu SL (2017) Main components of pomegranate, ellagic acid and luteolin, inhibit metastasis of ovarian cancer by down-regulating MMP2 and MMP9. Cancer Biol Ther 18(12):990–999. CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Khadijeh N, Daniel Elieh Ali K, Habibolah K, Ali M, Asad V-R, Hamid Reza A, Mohammad Rasoul G, Amir K (2018) Investigation of serum levels and activity of matrix metalloproteinases 2 and 9 (MMP2, 9) in opioid and methamphetamine-dependent patients. Acta Med Iran 56:559–562Google Scholar
  18. 18.
    Komi DEA, Khomtchouk K, Santa Maria PL (2019) A review of the contribution of mast cells in wound healing: involved molecular and cellular mechanisms. Clin Rev Allergy Immunol. CrossRefPubMedGoogle Scholar
  19. 19.
    Farina P, Tabouret E, Lehmann P, Barrie M, Petrirena G, Campello C, Boucard C, Graillon T, Girard N, Chinot O (2017) Relationship between magnetic resonance imaging characteristics and plasmatic levels of MMP2 and MMP9 in patients with recurrent high-grade gliomas treated by Bevacizumab and Irinotecan. J Neurooncol 132(3):433–437. CrossRefPubMedGoogle Scholar
  20. 20.
    Kim SY, Lee EJ, Woo MS, Jung JS, Hyun JW, Min SW, Kim DH, Kim HS (2008) Inhibition of matrix metalloproteinase-9 gene expression by an isoflavone metabolite, irisolidone in U87MG human astroglioma cells. Biochem Biophys Res Commun 366(2):493–499. CrossRefPubMedGoogle Scholar
  21. 21.
    Zhang JF, Wang P, Yan YJ, Li Y, Guan MW, Yu JJ, Wang XD (2017) IL33 enhances glioma cell migration and invasion by upregulation of MMP2 and MMP9 via the ST2-NF-kappaB pathway. Oncol Rep 38(4):2033–2042. CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Elieh Ali Komi D, Bjermer L (2019) Mast cell-mediated orchestration of the immune responses in human allergic asthma: current insights. Clin Rev Allergy Immunol 56(2):234–247. CrossRefPubMedGoogle Scholar
  23. 23.
    Buijs N, Oosterink JE, Jessup M, Schierbeek H, Stolz DB, Houdijk AP, Geller DA, van Leeuwen PA (2017) A new key player in VEGF-dependent angiogenesis in human hepatocellular carcinoma: dimethylarginine dimethylaminohydrolase 1. Angiogenesis 20(4):557–565. CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Turkowski K, Brandenburg S (2018) VEGF as a modulator of the innate immune response in glioblastoma. Glia 66(1):161–174. CrossRefPubMedGoogle Scholar
  25. 25.
    Azambuja JH, da Silveira EF, de Carvalho TR, Oliveira PS, Pacheco S, do Couto CT, Beira FT, Stefanello FM, Spanevello RM, Braganhol E (2017) Glioma sensitive or chemoresistant to temozolomide differentially modulate macrophage protumor activities. Biochim Biophys Acta 1861:2652–2662. CrossRefGoogle Scholar
  26. 26.
    Hassler MR, Sax C, Flechl B, Ackerl M, Preusser M, Hainfellner JA, Woehrer A, Dieckmann KU, Rossler K, Prayer D, Marosi C (2015) Thalidomide as palliative treatment in patients with advanced secondary glioblastoma. Oncology 88(3):173–179. CrossRefPubMedGoogle Scholar
  27. 27.
    Hafazalla K, Sahgal A, Jaja B, Perry JR, Das S (2018) Procarbazine, CCNU and vincristine (PCV) versus temozolomide chemotherapy for patients with low-grade glioma: a systematic review. Oncotarget 9(72):33623–33633. CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Thomas A, Tanaka M, Trepel J, Reinhold WC, Rajapakse VN, Pommier Y (2017) Temozolomide in the era of precision medicine. Cancer Res 77(4):823–826. CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Stupp R, Gander M, Leyvraz S, Newlands E (2001) Current and future developments in the use of temozolomide for the treatment of brain tumours. Lancet Oncol 2(9):552–560. CrossRefPubMedGoogle Scholar
  30. 30.
    Lee SY (2016) Temozolomide resistance in glioblastoma multiforme. Genes Dis 3(3):198–210. CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Yi GZ, Liu YW, Xiang W, Wang H, Chen ZY, Xie SD, Qi ST (2016) Akt and beta-catenin contribute to TMZ resistance and EMT of MGMT negative malignant glioma cell line. J Neurol Sci 367:101–106. CrossRefPubMedGoogle Scholar
  32. 32.
    Agarwala SS, Kirkwood JM (2000) Temozolomide, a novel alkylating agent with activity in the central nervous system, may improve the treatment of advanced metastatic melanoma. Oncologist 5(2):144–151. CrossRefPubMedGoogle Scholar
  33. 33.
    Zhou R, Zhang LZ, Wang RZ (2010) Effect of celecoxib on proliferation, apoptosis, and survivin expression in human glioma cell line U251. Chin J Cancer 29(3):294–299CrossRefGoogle Scholar
  34. 34.
    Zhang H, Gao S (2007) Temozolomide/PLGA microparticles and antitumor activity against glioma C6 cancer cells in vitro. Int J Pharm 329(1–2):122–128. CrossRefPubMedGoogle Scholar
  35. 35.
    Haghnavaz N, Asghari F, Elieh Ali Komi D, Shanehbandi D, Baradaran B, Kazemi T (2018) HER2 positivity may confer resistance to therapy with paclitaxel in breast cancer cell lines. Artif Cells Nanomed Biotechnol 46(3):518–523. CrossRefPubMedGoogle Scholar
  36. 36.
    Rieger AM, Nelson KL, Konowalchuk JD, Barreda DR (2011) Modified annexin V/propidium iodide apoptosis assay for accurate assessment of cell death. J Vis Exp. CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Huang H, Liu T, Guo J, Yu L, Wu X, He Y, Li D, Liu J, Zhang K, Zheng X, Goodin S (2017) Brefeldin A enhances docetaxel-induced growth inhibition and apoptosis in prostate cancer cells in monolayer and 3D cultures. Bioorg Med Chem Lett 27(11):2286–2291. CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Hsu EL, Chen N, Westbrook A, Wang F, Zhang R, Taylor RT, Hankinson O (2008) CXCR4 and CXCL12 down-regulation: a novel mechanism for the chemoprotection of 3,3’-diindolylmethane for breast and ovarian cancers. Cancer Lett 265(1):113–123. CrossRefPubMedGoogle Scholar
  39. 39.
    Kim HK, Kim JE, Chung J, Park KH, Han KS, Cho HI (2007) Lithium down-regulates the expression of CXCR4 in human neutrophils. J Trace Elem Med Biol 21(3):204–209. CrossRefPubMedGoogle Scholar
  40. 40.
    Gabelloni P, Da Pozzo E, Bendinelli S, Costa B, Nuti E, Casalini F, Orlandini E, Da Settimo F, Rossello A, Martini C (2010) Inhibition of metalloproteinases derived from tumours: new insights in the treatment of human glioblastoma. Neuroscience 168(2):514–522. CrossRefPubMedGoogle Scholar
  41. 41.
    Esteve PO, Tremblay P, Houde M, St-Pierre Y, Mandeville R (1998) In vitro expression of MMP-2 and MMP-9 in glioma cells following exposure to inflammatory mediators. Biochem Biophys Acta 1403(1):85–96CrossRefGoogle Scholar
  42. 42.
    Weyermann J, Lochmann D, Zimmer A (2005) A practical note on the use of cytotoxicity assays. Int J Pharm 288(2):369–376. CrossRefPubMedGoogle Scholar
  43. 43.
    Fotakis G, Timbrell JA (2006) In vitro cytotoxicity assays: comparison of LDH, neutral red, MTT and protein assay in hepatoma cell lines following exposure to cadmium chloride. Toxicol Lett 160(2):171–177. CrossRefPubMedGoogle Scholar
  44. 44.
    Geys J, Nemery B, Hoet PH (2010) Assay conditions can influence the outcome of cytotoxicity tests of nanomaterials: better assay characterization is needed to compare studies. Toxicol In Vitro 24(2):620–629. CrossRefPubMedGoogle Scholar
  45. 45.
    Ayala A, Munoz MF, Arguelles S (2014) Lipid peroxidation: production, metabolism, and signaling mechanisms of malondialdehyde and 4-hydroxy-2-nonenal. Oxid Med Cell Longev 2014:360438. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Nature B.V. 2020

Authors and Affiliations

  • Seyedsaber Mirabdaly
    • 1
  • Daniel Elieh Ali Komi
    • 2
    • 3
  • Yadollah Shakiba
    • 4
  • Ali Moini
    • 5
  • Amir Kiani
    • 4
    • 6
    Email author
  1. 1.Students Research CommitteeKermanshah University of Medical SciencesKermanshahIran
  2. 2.Immunology Research CenterTabriz University of Medical SciencesTabrizIran
  3. 3.Department of ImmunologyTabriz University of Medical SciencesTabrizIran
  4. 4.Regenerative Medicine Research Center, Kermanshah University of Medical SciencesKermanshahIran
  5. 5.Department of Internal Medicine ImamReza Hospital Kermanshah University of Medical SciencesKermanshahIran
  6. 6.Pharmaceutical Sciences Research Center, Health InstituteKermanshah University of Medical SciencesKermanshahIran

Personalised recommendations