Cancer Immunology, Immunotherapy

, Volume 58, Issue 9, pp 1375–1385 | Cite as

Toll-like receptor expression in normal ovary and ovarian tumors

  • Mingfu Zhou
  • Molly M. McFarland-Mancini
  • Holly M. Funk
  • Nader Husseinzadeh
  • Taofic Mounajjed
  • Angela F. Drew
Original Article


Recent studies have implicated inflammation in the initiation and progression of ovarian cancer, though the mechanisms underlying this effect are still not clear. Toll-like receptors (TLRs) allow immune cells to recognize pathogens and to trigger inflammatory responses. Tumor cell expression of TLRs can promote inflammation and cell survival in the tumor microenvironment. Here we sought to characterize the expression of TLRs in normal human ovaries, benign and malignant ovarian tumors from patients, and in established ovarian tumor cell lines. We report that TLR2, TLR3, TLR4, and TLR5 are strongly expressed on the surface epithelium of normal ovaries. In contrast to previous studies of uterus and endocervix, we found no cyclic variation in TLR expression occurred in murine ovaries. TLR2, TLR3, TLR4, and TLR5 are expressed in benign conditions, epithelial tumors, and in ovarian cancer cell lines. Variable expression of TLR6 and TLR8 was seen in benign and malignant epithelium of some patients, while expression of TLR1, TLR7, and TLR9 was weak. Normal and malignant ovarian stroma were negative for TLR expression. Vascular endothelial cells, macrophages, and occasional fibroblasts in tumors were positive. Functional activity for TLRs was demonstrated by stimulation of cell lines with specific ligands and subsequent activation and translocation of NFκB and release of the proinflammatory cytokines interleukin-6 and CCL-2. These studies demonstrate expression of multiple TLRs in the epithelium of normal ovaries and in ovarian tumor cells, and may indicate a mechanism by which epithelial tumors manipulate inflammatory pathways to facilitate tumor progression.


Toll-like receptor Ovary Ovarian tumor Inflammation Epithelium NFκB 



We gratefully acknowledge the expert assistance of Glenn Doerman and Maryellen Daston with graphics and editing of the manuscript, respectively. The 3EconAluc plasmid was kindly provided by Dr. Diaz-Meco, University of Cincinnati. This study was supported by a pilot project grant from the University of Cincinnati Cancer Center (AFD) and RSG-0614101CSM from the American Cancer Society (AFD).


  1. 1.
    DeNardo DG, Johansson M, Coussens LM (2008) Immune cells as mediators of solid tumor metastasis. Cancer Metastasis Rev 27:11–18PubMedCrossRefGoogle Scholar
  2. 2.
    Karin M, Greten FR (2005) NF-kappaB: linking inflammation and immunity to cancer development and progression. Nat Rev Immunol 5:749–759PubMedCrossRefGoogle Scholar
  3. 3.
    Burke F, Relf M, Negus R, Balkwill F (1996) A cytokine profile of normal and malignant ovary. Cytokine 8:578–585PubMedCrossRefGoogle Scholar
  4. 4.
    Haskill S, Becker S, Fowler W, Walton L (1982) Mononuclear-cell infiltration in ovarian cancer I. Inflammatory-cell infiltrates from tumour and ascites material. Br J Cancer 45:728–736PubMedGoogle Scholar
  5. 5.
    Penson RT, Kronish K, Duan Z, Feller AJ, Stark P, Cook SE, Duska LR, Fuller AF, Goodman AK, Nikrui N, MacNeill KM, Matulonis UA, Preffer FI, Seiden MV (2000) Cytokines IL-1beta, IL-2, IL-6, IL-8, MCP-1, GM-CSF and TNF-alpha in patients with epithelial ovarian cancer and their relationship to treatment with paclitaxel. Int J Gynecol Cancer 10:33–41PubMedCrossRefGoogle Scholar
  6. 6.
    Punnonen R, Teisala K, Kuoppala T, Bennett B, Punnonen J (1998) Cytokine production profiles in the peritoneal fluids of patients with malignant or benign gynecologic tumors. Cancer 83:788–796PubMedCrossRefGoogle Scholar
  7. 7.
    Freedman RS, Deavers M, Liu J, Wang E (2004) Peritoneal inflammation—a microenvironment for Epithelial Ovarian Cancer (EOC). J Transl Med 2:23PubMedCrossRefGoogle Scholar
  8. 8.
    Hagemann T, Wilson J, Burke F, Kulbe H, Li NF, Pluddemann A, Charles K, Gordon S, Balkwill FR (2006) Ovarian cancer cells polarize macrophages toward a tumor-associated phenotype. J Immunol 176:5023–5032PubMedGoogle Scholar
  9. 9.
    Wang X, Deavers M, Patenia R, Bassett RL Jr, Mueller P, Ma Q, Wang E, Freedman RS (2006) Monocyte/macrophage and T-cell infiltrates in peritoneum of patients with ovarian cancer or benign pelvic disease. J Transl Med 4:30–40PubMedCrossRefGoogle Scholar
  10. 10.
    Robinson-Smith TM, Isaacsohn I, Mercer CA, Zhou M, Van Rooijen N, Husseinzadeh N, McFarland-Mancini MM, Drew AF (2007) Macrophages mediate inflammation-enhanced metastasis of ovarian tumors in mice. Cancer Res 67:5708–5716PubMedCrossRefGoogle Scholar
  11. 11.
    Yu L, Chen S (2008) Toll-like receptors expressed in tumor cells: targets for therapy. Cancer Immunol Immunother 57:1271–1278PubMedCrossRefGoogle Scholar
  12. 12.
    Kelly MG, Alvero AB, Chen R, Silasi DA, Abrahams VM, Chan S, Visintin I, Rutherford T, Mor G (2006) TLR-4 signaling promotes tumor growth and paclitaxel chemoresistance in ovarian cancer. Cancer Res 66:3859–3868PubMedCrossRefGoogle Scholar
  13. 13.
    Fichorova RN, Cronin AO, Lien E, Anderson DJ, Ingalls RR (2002) Response to Neisseria gonorrhoeae by cervicovaginal epithelial cells occurs in the absence of Toll-Like receptor 4-mediated signaling. J Immunol 168:2424–2432PubMedGoogle Scholar
  14. 14.
    Fazeli A, Bruce C, Anumba DO (2005) Characterization of Toll-Like receptors in the female reproductive tract in humans. Hum Reprod 20:1372–1378PubMedCrossRefGoogle Scholar
  15. 15.
    Pioli PA, Amiel E, Schaefer TM, Connolly JE, Wira CR, Guyre PM (2004) Differential expression of Toll-Like receptors 2 and 4 in tissues of the human female reproductive tract. Infect Immun 72:5799–5806PubMedCrossRefGoogle Scholar
  16. 16.
    Aflatoonian R, Tuckerman E, Elliott SL, Bruce C, Aflatoonian A, Li TC, Fazeli A (2007) Menstrual cycle-dependent changes of Toll-like receptors in endometrium. Hum Reprod 22:586–593PubMedCrossRefGoogle Scholar
  17. 17.
    Pivarcsi A, Nagy I, Koreck A, Kis K, Kenderessy-Szabo A, Szell M, Dobozy A, Kemeny L (2005) Microbial compounds induce the expression of pro-inflammatory cytokines, chemokines and human beta-defensin-2 in vaginal epithelial cells. Microbes Infect 7:1117–1127PubMedCrossRefGoogle Scholar
  18. 18.
    Itoh H, Nasu K, Nishida M, Matsumoto H, Yuge A, Narahara H (2006) Human oviductal stromal fibroblasts, but not oviductal epithelial cells, express Toll-like receptor 4: the site-specific mucosal immunity of the human fallopian tube against bacterial infection. Am J Reprod Immunol 56:91–101PubMedCrossRefGoogle Scholar
  19. 19.
    Gonzalez JM, Xu H, Ofori E, Elovitz MA (2007) Toll-like receptors in the uterus, cervix, and placenta: is pregnancy an immunosuppressed state? Am J Obstet Gynecol 197:296 e1–6PubMedCrossRefGoogle Scholar
  20. 20.
    Shimada M, Hernandez-Gonzalez I, Gonzalez-Robanya I, Richards JS (2006) Induced expression of pattern recognition receptors in cumulus oocyte complexes: novel evidence for innate immune-like functions during ovulation. Mol Endocrinol 20:3228–3239PubMedCrossRefGoogle Scholar
  21. 21.
    Girling JE, Hedger MP (2007) Toll-like receptors in the gonads and reproductive tract: emerging roles in reproductive physiology and pathology. Immunol Cell Biol 85(6):481–489PubMedCrossRefGoogle Scholar
  22. 22.
    Rock FL, Hardiman G, Timans JC, Kastelein RA, Bazan JF (1998) A family of human receptors structurally related to Drosophila Toll. Proc Natl Acad Sci USA 95:588–593PubMedCrossRefGoogle Scholar
  23. 23.
    Takeuchi O, Kawai T, Sanjo H, Copeland NG, Gilbert DJ, Jenkins NA, Takeda K, Akira S (1999) TLR6: a novel member of an expanding toll-like receptor family. Gene 231:59–65PubMedCrossRefGoogle Scholar
  24. 24.
    Zarember KA, Godowski PJ (2002) Tissue expression of human Toll-like receptors and differential regulation of Toll-like receptor mRNAs in leukocytes in response to microbes, their products, and cytokines. J Immunol 168:554–561PubMedGoogle Scholar
  25. 25.
    Herath S, Williams EJ, Lilly ST, Gilbert RO, Dobson H, Bryant CE, Sheldon IM (2007) Ovarian follicular cells have innate immune capabilities that modulate their endocrine function. Reproduction 134:683–693PubMedCrossRefGoogle Scholar
  26. 26.
    Molteni M, Marabella D, Orlandi C, Rossetti C (2006) Melanoma cell lines are responsive in vitro to lipopolysaccharide and express TLR-4. Cancer Lett 235:75–83PubMedCrossRefGoogle Scholar
  27. 27.
    Krinke GJ (2004) Normative histology of organs. In: Hedrich HJ, Bullock G (eds) The laboratory mouse. Elsevier, LondonGoogle Scholar
  28. 28.
    Diaz-Meco MT, Municio MM, Sanchez P, Lozano J, Moscat J (1996) Lambda-interacting protein, a novel protein that specifically interacts with the zinc finger domain of the atypical protein kinase C isotype lambda/iota and stimulates its kinase activity in vitro and in vivo. Mol Cell Biol 16:105–114PubMedGoogle Scholar
  29. 29.
    Palazzo M, Balsari A, Rossini A, Selleri S, Calcaterra C, Gariboldi S, Zanobbio L, Arnaboldi F, Shirai YF, Serrao G, Rumio C (2007) Activation of enteroendocrine cells via TLRs induces hormone, chemokine, and defensin secretion. J Immunol 178:4296–4303PubMedGoogle Scholar
  30. 30.
    Takeuchi O, Hoshino K, Kawai T, Sanjo H, Takada H, Ogawa T, Takeda K, Akira S (1999) Differential roles of TLR2 and TLR4 in recognition of gram-negative and gram-positive bacterial cell wall components. Immunity 11:443–451PubMedCrossRefGoogle Scholar
  31. 31.
    Adachi O, Kawai T, Takeda K, Matsumoto M, Tsutsui H, Sakagami M, Nakanishi K, Akira S (1998) Targeted disruption of the MyD88 gene results in loss of IL-1- and IL-18-mediated function. Immunity 9:143–150PubMedCrossRefGoogle Scholar
  32. 32.
    Alexopoulou L, Holt AC, Medzhitov R, Flavell RA (2001) Recognition of double-stranded RNA and activation of NF-kappaB by Toll-like receptor 3. Nature 413:732–738PubMedCrossRefGoogle Scholar
  33. 33.
    Yang Y, Liu B, Dai J, Srivastava PK, Zammit DJ, Lefrancois L, Li Z (2007) Heat shock protein gp96 is a master chaperone for toll-like receptors and is important in the innate function of macrophages. Immunity 26:215–226PubMedCrossRefGoogle Scholar
  34. 34.
    Feuillet V, Medjane S, Mondor I, Demaria O, Pagni PP, Galan JE, Flavell RA, Alexopoulou L (2006) Involvement of Toll-like receptor 5 in the recognition of flagellated bacteria. Proc Natl Acad Sci USA 103:12487–12492PubMedCrossRefGoogle Scholar
  35. 35.
    Fukata M, Michelsen KS, Eri R, Thomas LS, Hu B, Lukasek K, Nast CC, Lechago J, Xu R, Naiki Y, Soliman A, Arditi M, Abreu MT (2005) Toll-like receptor-4 is required for intestinal response to epithelial injury and limiting bacterial translocation in a murine model of acute colitis. Am J Physiol Gastrointest Liver Physiol 288:G1055–G1065PubMedCrossRefGoogle Scholar
  36. 36.
    Macedo L, Pinhal-Enfield G, Alshits V, Elson G, Cronstein BN, Leibovich SJ (2007) Wound healing is impaired in MyD88-deficient mice: a role for MyD88 in the regulation of wound healing by adenosine A2A receptors. Am J Pathol 171:1774–1788PubMedCrossRefGoogle Scholar
  37. 37.
    Shimada M, Yanai Y, Okazaki T, Noma N, Kawashima I, Mori T, Richards JS (2008) Hyaluronan fragments generated by sperm-secreted hyaluronidase stimulate cytokine/chemokine production via the TLR2 and TLR4 pathway in cumulus cells of ovulated COCs, which may enhance fertilization. Development 135:2001–2011PubMedCrossRefGoogle Scholar
  38. 38.
    Bouman A, Moes H, Heineman MJ, de Leij LF, Faas MM (2001) The immune response during the luteal phase of the ovarian cycle: increasing sensitivity of human monocytes to endotoxin. Fertil Steril 76:555–559PubMedCrossRefGoogle Scholar
  39. 39.
    Moeinpour F, Choudhry MA, Kawasaki T, Timares L, Schwacha MG, Bland KI, Chaudry IH (2007) 17beta-Estradiol normalizes Toll receptor 4, mitogen activated protein kinases and inflammatory response in epidermal keratinocytes following trauma-hemorrhage. Mol Immunol 44:3317–3323PubMedCrossRefGoogle Scholar
  40. 40.
    Paimela T, Ryhanen T, Mannermaa E, Ojala J, Kalesnykas G, Salminen A, Kaarniranta K (2007) The effect of 17beta-estradiol on IL-6 secretion and NF-kappaB DNA-binding activity in human retinal pigment epithelial cells. Immunol Lett 110:139–144PubMedCrossRefGoogle Scholar
  41. 41.
    Vegeto E, Ghisletti S, Meda C, Etteri S, Belcredito S, Maggi A (2004) Regulation of the lipopolysaccharide signal transduction pathway by 17beta-estradiol in macrophage cells. J Steroid Biochem Mol Biol 91:59–66PubMedCrossRefGoogle Scholar
  42. 42.
    Schaefer TM, Fahey JV, Wright JA, Wira CR (2005) Innate immunity in the human female reproductive tract: antiviral response of uterine epithelial cells to the TLR3 agonist poly(I:C). J Immunol 174:992–1002PubMedGoogle Scholar
  43. 43.
    Ghosh M, Schaefer TM, Fahey JV, Wright JA, Wira CR (2008) Antiviral responses of human Fallopian tube epithelial cells to toll-like receptor 3 agonist poly(I:C). Fertil Steril 89:1497–1506PubMedCrossRefGoogle Scholar
  44. 44.
    Lee JW, Choi JJ, Seo ES, Kim MJ, Kim WY, Choi CH, Kim TJ, Kim BG, Song SY, Bae DS (2007) Increased toll-like receptor 9 expression in cervical neoplasia. Mol Carcinog 46:941–947PubMedCrossRefGoogle Scholar
  45. 45.
    Kim WY, Lee JW, Choi JJ, Choi CH, Kim TJ, Kim BG, Song SY, Bae DS (2008) Increased expression of Toll-like receptor 5 during progression of cervical neoplasia. Int J Gynecol Cancer 18:300–305PubMedCrossRefGoogle Scholar
  46. 46.
    Akazawa T, Ebihara T, Okuno M, Okuda Y, Shingai M, Tsujimura K, Takahashi T, Ikawa M, Okabe M, Inoue N, Okamoto-Tanaka M, Ishizaki H, Miyoshi J, Matsumoto M, Seya T (2007) Antitumor NK activation induced by the Toll-like receptor 3-TICAM-1 (TRIF) pathway in myeloid dendritic cells. Proc Natl Acad Sci USA 104:252–257PubMedCrossRefGoogle Scholar
  47. 47.
    El Andaloussi A, Sonabend AM, Han Y, Lesniak MS (2006) Stimulation of TLR9 with CpG ODN enhances apoptosis of glioma and prolongs the survival of mice with experimental brain tumors. Glia 54:526–535PubMedCrossRefGoogle Scholar
  48. 48.
    Scheel B, Aulwurm S, Probst J, Stitz L, Hoerr I, Rammensee HG, Weller M, Pascolo S (2006) Therapeutic anti-tumor immunity triggered by injections of immunostimulating single-stranded RNA. Eur J Immunol 36:2807–2816PubMedCrossRefGoogle Scholar
  49. 49.
    Zhu X, Nishimura F, Sasaki K, Fujita M, Dusak JE, Eguchi J, Fellows-Mayle W, Storkus WJ, Walker PR, Salazar AM, Okada H (2007) Toll like receptor-3 ligand poly-ICLC promotes the efficacy of peripheral vaccinations with tumor antigen-derived peptide epitopes in murine CNS tumor models. J Transl Med 5:10PubMedCrossRefGoogle Scholar
  50. 50.
    Rhee SH, Im E, Pothoulakis C (2008) Toll-like receptor 5 engagement modulates tumor development and growth in a mouse xenograft model of human colon cancer. Gastroenterology 135(2):518–528PubMedCrossRefGoogle Scholar
  51. 51.
    Harmey JH, Bucana CD, Lu W, Byrne AM, McDonnell S, Lynch C, Bouchier-Hayes D, Dong Z (2002) Lipopolysaccharide-induced metastatic growth is associated with increased angiogenesis, vascular permeability and tumor cell invasion. Int J Cancer 101:415–422PubMedCrossRefGoogle Scholar
  52. 52.
    Huang B, Zhao J, Li H, He KL, Chen Y, Chen SH, Mayer L, Unkeless JC, Xiong H (2005) Toll-like receptors on tumor cells facilitate evasion of immune surveillance. Cancer Res 65:5009–5014PubMedCrossRefGoogle Scholar
  53. 53.
    Huang B, Zhao J, Shen S, Li H, He KL, Shen GX, Mayer L, Unkeless J, Li D, Yuan Y, Zhang GM, Xiong H, Feng ZH (2007) Listeria monocytogenes promotes tumor growth via tumor cell toll-like receptor 2 signaling. Cancer Res 67:4346–4352PubMedCrossRefGoogle Scholar
  54. 54.
    Sfondrini L, Rossini A, Besusso D, Merlo A, Tagliabue E, Menard S, Balsari A (2006) Antitumor activity of the TLR-5 ligand flagellin in mouse models of cancer. J Immunol 176:6624–6630PubMedGoogle Scholar
  55. 55.
    Naugler WE, Sakurai T, Kim S, Maeda S, Kim K, Elsharkawy AM, Karin M (2007) Gender disparity in liver cancer due to sex differences in MyD88-dependent IL-6 production. Science 317:121–124PubMedCrossRefGoogle Scholar
  56. 56.
    Rakoff-Nahoum S, Medzhitov R (2007) Regulation of spontaneous intestinal tumorigenesis through the adaptor protein MyD88. Science 317:124–127PubMedCrossRefGoogle Scholar
  57. 57.
    Risch HA, Howe GR (1995) Pelvic inflammatory disease and the risk of epithelial ovarian cancer. Cancer Epidemiol Biomarkers Prev 4:447–451PubMedGoogle Scholar
  58. 58.
    Ness RB, Grisso JA, Cottreau C, Klapper J, Vergona R, Wheeler JE, Morgan M, Schlesselman JJ (2000) Factors related to inflammation of the ovarian epithelium and risk of ovarian cancer. Epidemiology 11:111–117PubMedCrossRefGoogle Scholar
  59. 59.
    Wong A, Maclean AB, Furrows SJ, Ridgway GL, Hardiman PJ, Perrett CW (2007) Could epithelial ovarian cancer be associated with chlamydial infection? Eur J Gynaecol Oncol 28:117–120PubMedGoogle Scholar
  60. 60.
    Ness RB, Goodman MT, Shen C, Brunham RC (2003) Serologic evidence of past infection with Chlamydia trachomatis, in relation to ovarian cancer. J Infect Dis 187:1147–1152PubMedCrossRefGoogle Scholar
  61. 61.
    Apetoh L, Ghiringhelli F, Tesniere A, Criollo A, Ortiz C, Lidereau R, Mariette C, Chaput N, Mira JP, Delaloge S, Andre F, Tursz T, Kroemer G, Zitvogel L (2007) The interaction between HMGB1 and TLR4 dictates the outcome of anticancer chemotherapy and radiotherapy. Immunol Rev 220:47–59PubMedCrossRefGoogle Scholar
  62. 62.
    Gobert AP, Bambou JC, Werts C, Balloy V, Chignard M, Moran AP, Ferrero RL (2004) Helicobacter pylori heat shock protein 60 mediates interleukin-6 production by macrophages via a toll-like receptor (TLR)-2-, TLR-4-, and myeloid differentiation factor 88-independent mechanism. J Biol Chem 279:245–250PubMedCrossRefGoogle Scholar
  63. 63.
    Termeer C, Benedix F, Sleeman J, Fieber C, Voith U, Ahrens T, Miyake K, Freudenberg M, Galanos C, Simon JC (2002) Oligosaccharides of Hyaluronan activate dendritic cells via toll-like receptor 4. J Exp Med 195:99–111PubMedCrossRefGoogle Scholar
  64. 64.
    Okamura Y, Watari M, Jerud ES, Young DW, Ishizaka ST, Rose J, Chow JC, Strauss JF 3rd (2001) The extra domain A of fibronectin activates Toll-like receptor 4. J Biol Chem 276:10229–10233PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Mingfu Zhou
    • 1
  • Molly M. McFarland-Mancini
    • 1
  • Holly M. Funk
    • 1
  • Nader Husseinzadeh
    • 2
  • Taofic Mounajjed
    • 3
  • Angela F. Drew
    • 1
  1. 1.Department of Cancer and Cell BiologyUniversity of CincinnatiCincinnatiUSA
  2. 2.Division of Gynecologic OncologyUniversity of CincinnatiCincinnatiUSA
  3. 3.Department of Pathology and Laboratory MedicineUniversity of CincinnatiCincinnatiUSA

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