title: A mutated cholera toxin without the ADP-ribosyltransferase activity induces cytokine production and inhibits apoptosis of splenocytes in mice possibly via toll-like receptor-4 signaling.
abstract: Native cholera toxin (CT) and its mutated form (CT-2*) without ADP-ribosyltransferase activity differ in their immunomodulatory effects on host cells, and the mechanisms of these differences are poorly understood. In this study, we demonstrated that CT-2* induced higher levels of cytokine production and down-regulated ex-vivo apoptosis of splenocytes from C57BL/6 mice. After exposure of the splenocytes ex-vivo to CT or CT-2* (2μg/ml) for 48h, CT-2* stimulated expression of the toll-like receptor (TLR-4) gene was much higher and the cells produced increased levels of interleukin (IL)-12, interferon (IFN)-γ, and tumor necrosis factor (TNF)-α, compared to splenocytes of mice exposed to native CT. We confirmed these findings by observing that CT-2*, induced much lower levels of IL-12, IFN-γ, and TNF-α in a TLR-4 knockout macrophage cell line derived from C57BL/6 mice. In addition, while CT is known to stimulate apoptosis in splenocytes, we observed that CT-2* significantly down-regulated apoptosis (4.2%), compared to splenocytes exposed to CT (18.7%) or PBS (negative control, 8.5%). On the contrary, we noted both native CT and CT-2* to exhibit similar levels of apoptosis in TLR-4(-/-) cell line. Overall, the evidence supports the conclusion that CT-2* modulated cytokine production and apoptosis in splenocytes of mice possibly through the TLR-4 signaling pathway.
authors.affiliation: Immunology and Tumor Research Institute, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, Shanxi 710061, China. Electronic address: xianliutie@mail.xjtu.edu.cn.
authors.forename: Tie
authors.initials: T
authors.lastname: Liu
authors.affiliation: Immunology and Tumor Research Institute, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, Shanxi 710061, China; Core Research Laboratory, The Second Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, Shanxi 710049, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Wei
authors.affiliation: Immunology and Tumor Research Institute, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, Shanxi 710061, China; Clinical Research Center, Guangdong Medical Collaege, Zhanjiang, Guangdong 524001, China.
authors.forename: Gang
authors.initials: G
authors.lastname: Liu
authors.affiliation: Immunology and Tumor Research Institute, The First Affiliated Hospital, Xi'an Jiaotong University Health Science Center, Xi'an, Shanxi 710061, China.
authors.forename: Bingyin
authors.initials: B
authors.lastname: Shi
authors.affiliation: Department of Microbiology and Immunology, University of Texas Medical Branch USA, 77555, USA.
authors.forename: Suarez
authors.initials: S
authors.lastname: Giovanni
authors.affiliation: Department of Microbiology and Immunology, University of Texas Medical Branch USA, 77555, USA; Institute for Human Infections & Immunity, USA; WHO Collaborating Center for Vaccine Development, USA.
authors.forename: Johnny W
authors.initials: JW
authors.lastname: Peterson
authors.affiliation: Department of Microbiology and Immunology, University of Texas Medical Branch USA, 77555, USA; Institute for Human Infections & Immunity, USA; WHO Collaborating Center for Vaccine Development, USA.
authors.forename: Ashok K
authors.initials: AK
authors.lastname: Chopra
mesh_terms: D036002:ADP Ribose Transferases; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000276:immunology*; D002772:Cholera Toxin / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology; D016207:Cytokines / Q000096:biosynthesis*; D005260:Female; D005434:Flow Cytometry; D055786:Gene Knockout Techniques; D008264:Macrophages / Q000187:drug effects / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation; D016133:Polymerase Chain Reaction; D015398:Signal Transduction / Q000276:immunology; D013154:Spleen / Q000187:drug effects / Q000276:immunology*; D051197:Toll-Like Receptor 4 / Q000276:immunology*
chemical_list: D016207:Cytokines; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; D002772:Cholera Toxin; D036002:ADP Ribose Transferases
keywords: Apoptosis; CT-2*; Cytokines; IL-12; Splenocyte; TLR4