title: Immunosuppressive factors secreted by human amniotic epithelial cells.
abstract: OBJECTIVE
  Amniotic membrane has been applied to the ocular surface to restore corneal function. The beneficial effect of amniotic membrane transplantation may be due to the immunosuppressive effects of amniotic epithelial cells. The purpose of this study was to determine whether amniotic epithelial cells (AECs) secrete anti-inflammatory and antiproliferative factors that affect the chemotaxis of neutrophils and macrophages and suppress both T- and B-cell proliferation in vitro.
  
  
  METHODS
  Human amniotic cells were isolated from human amniotic membrane and cultured in vitro. The supernatants from AEC cultures were collected after 48 hours of incubation. Neutrophil and macrophage chemotactic activity was tested in the presence of AEC supernatant, using 24-well migration assay chambers. Lymphocyte proliferation was tested by H(3)-thymidine incorporation. Apoptosis was examined by caspase-3 and annexin V assays, and expression of cytokines was assessed by RT-PCR.
  
  
  RESULTS
  AEC supernatant significantly inhibited the chemotactic activity of neutrophils and macrophages toward macrophage inflammatory protein (MIP)-2 (P < 0.05). The supernatant significantly reduced the proliferation of both T and B cells after mitogenic stimulation (P < 0.05). Caspase-3 assays revealed that the supernatant induced apoptosis of T and B cells, but not of corneal epithelial cells and liver cells. In contrast to lymphocytes, macrophages and neutrophils were resistant to apoptosis induced by AEC supernatant. The AECs expressed message for TNFalpha, Fas ligand (FasL), TRAIL (tumor necrosis factor-related apoptosis-inducing ligand), TGFbeta, and macrophage migration-inhibitory factor (MIF). However, AEC induction of apoptosis was inhibited (50%) by anti-FasL antibody but not by anti-TRAIL or anti-TNFalpha antibodies. Moreover, AEC supernatant inhibited macrophage migration in vitro.
  
  
  CONCLUSIONS
  AECs secrete soluble factors that inhibit cells in both the innate and adaptive immune systems.
authors.affiliation: Department of Ophthalmology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Haochuan
authors.initials: H
authors.lastname: Li
authors.forename: Jerry Y
authors.initials: JY
authors.lastname: Niederkorn
authors.forename: Sudha
authors.initials: S
authors.lastname: Neelam
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Mayhew
authors.forename: R Ann
authors.initials: RA
authors.lastname: Word
authors.forename: James P
authors.initials: JP
authors.lastname: McCulley
authors.forename: Hassan
authors.initials: H
authors.lastname: Alizadeh
mesh_terms: D000650:Amnion / Q000166:cytology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001402:B-Lymphocytes / Q000187:drug effects / Q000473:pathology; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D002633:Chemotaxis / Q000187:drug effects; D004847:Epithelial Cells / Q000276:immunology*; D006801:Humans; D007166:Immunosuppressive Agents / Q000302:isolation & purification* / Q000494:pharmacology; D019169:Jurkat Cells; D008213:Lymphocyte Activation / Q000187:drug effects; D017737:Macrophages, Peritoneal / Q000502:physiology; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D009504:Neutrophils / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013601:T-Lymphocytes / Q000187:drug effects / Q000473:pathology
chemical_list: D007166:Immunosuppressive Agents; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases