DOI QR코드

DOI QR Code

Comparative Studies to Evaluate Relative in vitro Potency of Luteolin in Inducing Cell Cycle Arrest and Apoptosis in HaCaT and A375 Cells

  • Published : 2013.02.28

Abstract

Luteolin is a naturally occurring flavonoid present in many plants with diverse applications in pharmacology. Despite several studies elucidating its significant anti-cancer activity against various cancer cells, the mechanism of action in skin cancer is not well addressed. Hence, we investigated the effects of luteolin in HaCaT (human immortalized keratinocytes) and A375 (human melanoma) cells. The radical scavenging abilities of luteolin were determined spectrophotometrically, prior to a cytotoxic study (XTT assay). Inhibitory effects were assessed by colony formation assay. Further, the capability of luteolin to induce cell cycle arrest and apoptosis were demonstrated by flow cytometry and cellular DNA fragmentation ELISA, respectively. The results revealed that luteolin possesses considerable cytotoxicity against both HaCaT and A375 cells with $IC_{50}$ values of 37.1 ${\mu}M$ and 115.1 ${\mu}M$, respectively. Luteolin also inhibited colony formation and induced apoptosis in a dose and time-dependent manner by disturbing cellular integrity as evident from morphological evaluation by Wright-Giemsa staining. Accumulation of cells in G2/M (0.83-8.14%) phase for HaCaT cells and G0/G1 (60.4-72.6%) phase for A375 cells after 24 h treatment indicated cell cycle arresting potential of this flavonoid. These data suggest that luteolin inhibits cell proliferation and promotes cell cycle arrest and apoptosis in skin cancer cells with possible involvement of programmed cell death, providing a substantial basis for it to be developed into a potent chemopreventive template for skin cancer.

Keywords

References

  1. Adinarayan M, Krishnamurthy SP (2011). Clinicopathological evaluation of nonmelanoma skin cancer. Indian J Dermatol, 56, 670-2. https://doi.org/10.4103/0019-5154.91826
  2. Benzie IF, Strain JJ (1996). The ferric reducing ability of plasma (FRAP) as a measure of "antioxidant power": the FRAP assay. Anal Biochem, 239, 70-6. https://doi.org/10.1006/abio.1996.0292
  3. Brady MS, Kaushal A, KO C, et al (2011). Melanoma and other skin cancers. Cancer Management, 14, 1-32.
  4. Chahar KM, Sharma N, Dobhal MP, et al (2011). Flavonoids: a versatile source of anticancer drugs. Pharmacognosy Rev, 5, 1-12. https://doi.org/10.4103/0973-7847.79093
  5. Chang J, Hsu Y, Kuo P, et al (2005). Increase of Bax/ Bcl-XL ratio and arrest of cell cycle by luteolin in immortalized human hepatoma cell line. Life Sci, 76, 1883-93. https://doi.org/10.1016/j.lfs.2004.11.003
  6. Chen T, Wong YS (2009). Selenocystine induces reactive oxygen species mediated apoptosis in human cancer cells. Biomed Pharmacother, 63, 105-13. https://doi.org/10.1016/j.biopha.2008.03.009
  7. Chiang CT, Way TD, Lin JK (2007). Sensitizing HER2-overexpressing cancer cells to luteolin-induced apoptosis through suppressing p21(WAF1/CIP1) expression with rapamycin. Mol Cancer Ther, 6, 2127-38. https://doi.org/10.1158/1535-7163.MCT-07-0107
  8. Chung H, Suh EK, Han IO, et al (2011). Keratinocyte-derived laminin-332 promotes adhesion and migration in melanocytes and melanoma. J Biol Chem, 15, 13438-47.
  9. Deorukhkar A, Krishnan S, Sethi G (2007). Back to basics: how natural products can provide the basis for new therapeutics. Expert Opin Investig Drugs, 16, 1753-73. https://doi.org/10.1517/13543784.16.11.1753
  10. Dua P, Gude RP (2006). Antiproliferative and antiproteolytic activity of pentoxifylline in cultures of B16F10 melanoma cells. Cancer Chemother Pharmacol, 58, 195-202. https://doi.org/10.1007/s00280-005-0155-8
  11. Fabbrocini G, Triassi M, Mauriello MC, et al (2010). Epidemiology of skin cancer: role of some environmental factors. Cancers, 2, 1980-9. https://doi.org/10.3390/cancers2041980
  12. Fecker LF, Geilen CC, Hossini AM, et al (2005). Selective induction of apoptosis in melanoma cells by tyrosinase promoter-controlled CD95 ligand overexpression. J Invest Dermatol, 124, 221-8. https://doi.org/10.1111/j.0022-202X.2004.23572.x
  13. Garbe C, Eigentler TK, Keilholz U, et al (2011). Systematic review of medical treatment in melanoma: current status and future prospects. The Oncologist, 16, 5-24. https://doi.org/10.1634/theoncologist.2010-0190
  14. George VC, Kumar DRN, Rajkumar V, et al (2012). Quantitative assessment of the relative antineoplastic potential of the n-butanolic leaf extract of Annona muricata Linn. in normal and immortalized human cell lines. Asian Pac J Cancer Prev, 13, 699-704. https://doi.org/10.7314/APJCP.2012.13.2.699
  15. Gibellini L, Pinti M, Nasi M, et al (2010). Interfering with ROS metabolism in cancer cells: the potential role of quercetin. Cancers, 2, 1288-311. https://doi.org/10.3390/cancers2021288
  16. Girschik J, Fritschi L, Threlfall T, et al (2008). Deaths from non-melanoma skin cancer in eastern Australia. Cancer Causes Control, 19, 879-85. https://doi.org/10.1007/s10552-008-9150-9
  17. Green DR, Reed JC (1998). Mitochondria and apoptosis. Science, 281, 1309-12. https://doi.org/10.1126/science.281.5381.1309
  18. Gu YH, Belury MA (2005). Selective induction of apoptosis in murine skin carcinoma cells (CH72) by an ethanol extract of Lentinula edodes. Cancer Lett, 220, 21-8. https://doi.org/10.1016/j.canlet.2004.06.037
  19. Hong WK, Sporn MB (1997). Recent advances in chemoprevention of cancer. Science, 278, 1073-7. https://doi.org/10.1126/science.278.5340.1073
  20. Hsan KM, Chen CC, Shyur LF (2010). Current research and development of chemotherapeutic agents for melanoma. Cancers, 2, 397-419. https://doi.org/10.3390/cancers2020397
  21. Kajstura M, Halicka HD, Pryjma J, et al (2007). Discontinuous fragmentation of nuclear DNA during apoptosis revealed by discrete "sub-G1" peaks on DNA content histograms. Cytometry, 71, 125-31.
  22. Kamran MZ, Gude RP (2012). Preclinical evaluation of the antimetastatic efficacy of Pentoxifylline on A375 human melanoma cell line. Biomed Pharmacother, 10, 6.
  23. Kang N, Zhang JH, Qiu F, et al (2010). Induction of G(2)/M phase arrest and apoptosis by oridonin in human laryngeal carcinoma cells. J Nat Prod, 73, 1058-63. https://doi.org/10.1021/np9008199
  24. Kang OH, Choi JG, Lee JH, et al (2010). Luteolin isolated from the flowers of Lonicera japonica suppresses inflammatory mediator release by blocking NF-${\kappa}B$ and MAPKs activation pathways in HMC-1 cells. Molecules, 15, 385-98. https://doi.org/10.3390/molecules15010385
  25. Kim MJ, Woo JS, Kwon CH, et al (2012). Luteolin induces apoptotic cell death through AIF nuclear translocation mediated by activation of ERK and p38 in human breast cancer cell lines. Cell Biol Int, 36, 339-44. https://doi.org/10.1042/CBI20110394
  26. Kobayashi T, Nakata T, Kuzumaki T (2002). Effect of flavonoids on cell cycle progression in prostate cancer cells. Cancer Lett, 176, 17-23. https://doi.org/10.1016/S0304-3835(01)00738-8
  27. Krishnamurthy R, Maly DJ (2010). Biochemical mechanisms of resistance to small-molecule protein kinase inhibitors. ACS Chem Biol, 5, 121-38. https://doi.org/10.1021/cb9002656
  28. Kuo JH, Chu YL, Yang JS, et al (2010). Cantharidin induces apoptosis in human bladder cancer TSGH 8301 cells through mitochondria-dependent signal pathways. Int J Oncol, 37, 1243-50.
  29. Lo C, Lai TY, Yang JS, et al (2011). Gallic acid inhibits the migration and invasion of A375.S2 human melanoma cells through the inhibition of matrix metalloproteinase-2 and Ras. Melanoma Res, 21, 267-73. https://doi.org/10.1097/CMR.0b013e3283414444
  30. Maioli E, Torricelli C, Fortino V, et al (2009). Critical appraisal of the MTT assay in the presence of rottlerin and uncouplers. Biol Proced Online, 11, 227-40. https://doi.org/10.1007/s12575-009-9020-1
  31. Middleton EJ, Kandaswami C, Theoharides TC (2000). Effects of plant flavonoids on mammalian cells:implications for inflammation, heart disease and cancer. Pharmacol Rev, 52, 673-751.
  32. Miimohammadsadegh A, Hassan M, Bardenheuer W, et al (2006). STAT5 phosphorylation in malignant melanoma is important for survival and is mediated through SRC and JAK1 kinases. J Invest Dermatol, 126, 2272-80. https://doi.org/10.1038/sj.jid.5700385
  33. Miimohammadsadegh A, Mota R, Gustrau A, et al (2007). ERK1/2 is highly phosphorylated in melanoma metastases and protects melanoma cells from cisplatin-mediated apoptosis. J Invest Dermatol, 127, 2207-15. https://doi.org/10.1038/sj.jid.5700870
  34. Ozgen U, Mavi A, Terzi Z, et al (2011). Relationship between chemical structure and antioxidant activity of luteolin and its glycosides isolated from Thymus. Rec Nat Prod, 5, 12-21.
  35. Pelicano H, Carney D, Huang P (2004). ROS stress in cancer cells and therapeutic implications. Drug Resist Updat, 7, 97-110. https://doi.org/10.1016/j.drup.2004.01.004
  36. Ross JA, Kasum CM (2002). Dietary flavonoids: bioavailability, metabolic effects, and safety. Annu Rev Nutr, 22, 19-34. https://doi.org/10.1146/annurev.nutr.22.111401.144957
  37. Seelinger G, Merfort I, Schempp CM (2008). Anti-oxidant, anti-inflammatory and anti-allergic activities of luteolin. Planta Med, 74, 1667-77. https://doi.org/10.1055/s-0028-1088314
  38. Shahidi F (1997). Natural antioxidants: An overview. In natural antioxidants, chemistry, health effects and applications. Edited by Shahidi F. Champaign, AOCS Press: 1-11.
  39. Shi J, Wei L (2007). Rho kinase in the regulation of cell death and survival. Arch Immunol Ther Exp, 55, 61-75. https://doi.org/10.1007/s00005-007-0009-7
  40. Shi RX, Ong CN, Shen HM (2004). Luteolin sensitizes tumor necrosis factor-alpha-induced apoptosis in human tumor cells. Oncogene, 23, 7712-21. https://doi.org/10.1038/sj.onc.1208046
  41. Shimada K, Fujikawa K, Yahara K, et al (1992). Antioxidative properties of xanthin and autooxidation of soybean oil in cyclodextrin emulsion. J Agric Food Chem, 40, 945-8. https://doi.org/10.1021/jf00018a005
  42. Soares JR, Dins TCP, Cunha AP, et al (1997). Antioxidant activity of some extracts of Thymus zygis. Free Radic Res, 26, 469-78. https://doi.org/10.3109/10715769709084484
  43. Ueda H, Yamazaki C, Yamazaki M (2002). Luteolin as an anti-inflammatory and anti-allergic constituent of Perilla frutescens. Biol Pharm Bull, 25, 1197-202. https://doi.org/10.1248/bpb.25.1197
  44. Voland C, Bord A, Peleraux A, et al (2006). Repression of cell cycle-related proteins by oxaliplatin but not cisplatin in human colon cancer cells. Mol Cancer Ther, 5, 2149-57. https://doi.org/10.1158/1535-7163.MCT-05-0212
  45. Wang HY, Quan K, Jiang YL, et al (2010). Effect of Luteolin and its combination with chemotherapeutic drugs on cytotoxicity of cancer cells. J Zhejiang Uni Medi Sci, 39, 30-6.
  46. Wang LM, Xie KP, Huo HN, et al (2012). Luteolin inhibits proliferation induced by IGF-1 pathway dependent ER$\alpha$ in human breast cancer MCF-7 cells. Asian Pac J Cancer Prev, 13, 1431-7. https://doi.org/10.7314/APJCP.2012.13.4.1431
  47. Wang Z, Tang X, Zhang Y, et al (2012). Lobaplatin induces apoptosis and arrests cell cycle progression in a human cholangio carcinoma cell line RBE. Biomed Pharmacother, 66, 161-6. https://doi.org/10.1016/j.biopha.2011.09.008
  48. Weislow OS, Kiser R, Fine DL, et al (1989). New soluble-formazan assay for HIV-1 cytopathic effects: application to high-flux screening of synthetic and natural products for AIDS-antiviral activity. J Natl Cancer Inst, 81, 577-86. https://doi.org/10.1093/jnci/81.8.577
  49. Xu K, Liu B, Ma Y, et al (2009). Physicochemical properties and antioxidant activities of luteolin-phospholipid complex. Molecules, 14, 3486-93. https://doi.org/10.3390/molecules14093486

Cited by

  1. Phytoestrogens as natural prodrugs in cancer prevention: towards a mechanistic model vol.13, pp.4, 2014, https://doi.org/10.1007/s11101-014-9355-3
  2. Luteolin Arrests Cell Cycling, Induces Apoptosis and Inhibits the JAK/STAT3 Pathway in Human Cholangiocarcinoma Cells vol.15, pp.12, 2014, https://doi.org/10.7314/APJCP.2014.15.12.5071
  3. Luteolin-loaded Phytosomes Sensitize Human Breast Carcinoma MDA-MB 231 Cells to Doxorubicin by Suppressing Nrf2 Mediated Signalling vol.15, pp.13, 2014, https://doi.org/10.7314/APJCP.2014.15.13.5311
  4. Antioxidant value and Antiproliferative Efficacy of Mitragynine and a Silane Reduced Analogue vol.15, pp.14, 2014, https://doi.org/10.7314/APJCP.2014.15.14.5659
  5. Luteolin Suppresses Cancer Cell Proliferation by Targeting Vaccinia-Related Kinase 1 vol.9, pp.10, 2014, https://doi.org/10.1371/journal.pone.0109655
  6. Anti-Proliferative Effect of Rosmarinus officinalis L. Extract on Human Melanoma A375 Cells vol.10, pp.7, 2015, https://doi.org/10.1371/journal.pone.0132439
  7. Luteolin induces caspase-14-mediated terminal differentiation in human epidermal keratinocytes vol.51, pp.10, 2015, https://doi.org/10.1007/s11626-015-9936-5
  8. Molecular mechanisms of flavonoids in melanin synthesis and the potential for the prevention and treatment of melanoma vol.60, pp.6, 2016, https://doi.org/10.1002/mnfr.201500822
  9. Novel Structurally Related Flavones Augment Cell Death Induced by rhsTRAIL vol.18, pp.6, 2017, https://doi.org/10.3390/ijms18061211
  10. Apple Flavonoids Suppress Carcinogen-Induced DNA Damage in Normal Human Bronchial Epithelial Cells vol.2017, pp.1942-0994, 2017, https://doi.org/10.1155/2017/1767198
  11. -Methyl Transferase Inhibition on Anti-Inflammatory Activity of Luteolin Metabolites vol.82, pp.2, 2017, https://doi.org/10.1111/1750-3841.13620
  12. Enhanced Induction of Apoptosis in HaCaT Cells by Luteolin Encapsulated in PEGylated Liposomes—Role of Caspase-3/Caspase-14 pp.1559-0291, 2018, https://doi.org/10.1007/s12010-018-2907-z