title: Toll-like receptors 7, 8, and 9 expression and function in primary human cervical cancer Langerhans cells: evidence of anergy.
abstract: OBJECTIVE
  Human papillomavirus oncoproteins E6 and E7 down modulate Toll-like receptor (TLR) 9 expression in infected keratinocytes. We explored the status of expression and function of TLR7, TLR8, and TLR9 in primary human Langerhans cells (LCs) isolated from cervical tumors.
  
  
  METHODS
  Single-cell suspensions were made from fresh tissues of squamous cell carcinoma (International Federation of Gynecology and Obstetrics stage IB2); myeloid dendritic cells were purified using CD1c magnetic activated cell separation kits. Langerhans cells were further flow sorted into CD1a*CD207* cells. Acute monocytic leukemia cell line THP-1-derived LCs (moLCs) formed the controls. mRNA from flow-sorted LCs was reverse transcribed to cDNA and TLR7, TLR8, and TLR9 amplified. Monocyte-derived Langerhans cells and cervical tumor LCs were stimulated with TLR7, TLR8, and TLR9 ligands. Culture supernatants were assayed for interleukin (IL) 1β, IL-6, IL-10, IL-12p70, interferon (IFN) α, interferon γ, and tumor necrosis factor (TNF) α by Luminex multiplex bead array. Human papillomavirus was genotyped.
  
  
  RESULTS
  We have for the first time demonstrated that the acute monocytic leukemia cell line THP-1 can be differentiated into LCs in vitro. Although these moLCs expressed all the 3 TLRs, tumor LCs expressed TLR7 and TLR8, but uniformly lacked TLR9. Also, moLCs secreted IL-6, IL-1β, and tumor necrosis factor α to TLR8 ligand and interferon α in response to TLR9 ligand; in contrast, tumor LCs did not express any cytokine to any of the 3 TLR ligands. Human papillomavirus type 16 was one of the common human papillomavirus types in all cases.
  
  
  CONCLUSIONS
  Cervical tumor LCs lacked TLR9 expression and were functionally anergic to all the 3: TLR7, TLR8, and TLR9 ligands, which may play a crucial role in immune tolerance. The exact location of block(s) in TLR7 and TLR8 signaling needs to be investigated, which would have important immunotherapeutic implications.
authors.affiliation: Department of Microbiology, Kidwai Memorial Institute of Oncology, Bangalore, India.
authors.forename: Mahesh M
authors.initials: MM
authors.lastname: Kumar
authors.forename: Sreenivas
authors.initials: S
authors.lastname: Adurthi
authors.forename: Surya
authors.initials: S
authors.lastname: Ramachandran
authors.forename: Geetashree
authors.initials: G
authors.lastname: Mukherjee
authors.forename: Omana
authors.initials: O
authors.lastname: Joy
authors.forename: H
authors.initials: H
authors.lastname: Krishnamurthy
authors.forename: Sudhir
authors.initials: S
authors.lastname: Krishna
authors.forename: U D
authors.initials: UD
authors.lastname: Bafna
authors.forename: Devi K
authors.initials: DK
authors.lastname: Uma
authors.forename: R S
authors.initials: RS
authors.lastname: Jayshree
mesh_terms: D000328:Adult; D000368:Aged; D002294:Carcinoma, Squamous Cell / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D016022:Case-Control Studies; D017635:Clonal Anergy / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology; D006801:Humans; D007801:Langerhans Cells / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D008024:Ligands; D008875:Middle Aged; D061251:Primary Cell Culture; D051199:Toll-Like Receptor 7 / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D051200:Toll-Like Receptor 8 / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D051217:Toll-Like Receptor 9 / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D014407:Tumor Cells, Cultured; D002583:Uterine Cervical Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
chemical_list: D008024:Ligands; C495348:TLR7 protein, human; C495349:TLR8 protein, human; C495371:TLR9 protein, human; D051199:Toll-Like Receptor 7; D051200:Toll-Like Receptor 8; D051217:Toll-Like Receptor 9