title: Development of an in vitro assay and demonstration of Plasmodium berghei liver-stage inhibition by TRAP-specific CD8+ T cells.
abstract: The development of an efficacious vaccine against the Plasmodium parasite remains a top priority. Previous research has demonstrated the ability of a prime-boost virally vectored sub-unit vaccination regimen, delivering the liver-stage expressed malaria antigen TRAP, to produce high levels of antigen-specific T cells. The liver-stage of malaria is the main target of T cell-mediated immunity, yet a major challenge in assessing new T cell inducing vaccines has been the lack of a suitable pre-clinical assay. We have developed a flow-cytometry based in vitro T cell killing assay using a mouse hepatoma cell line, Hepa1-6, and Plasmodium berghei GFP expressing sporozoites. Using this assay, P. berghei TRAP-specific CD8+ T cell enriched splenocytes were shown to inhibit liver-stage parasites in an effector-to-target ratio dependent manner. Further development of this assay using human hepatocytes and P. falciparum would provide a new method to pre-clinically screen vaccine candidates and to elucidate mechanisms of protection in vitro.
authors.affiliation: The Jenner Institute, University of Oxford, Oxford, United Kingdom.
authors.forename: Rhea J
authors.initials: RJ
authors.lastname: Longley
authors.affiliation: The Jenner Institute, University of Oxford, Oxford, United Kingdom.
authors.forename: Karolis
authors.initials: K
authors.lastname: Bauza
authors.affiliation: The Jenner Institute, University of Oxford, Oxford, United Kingdom.
authors.forename: Katie J
authors.initials: KJ
authors.lastname: Ewer
authors.affiliation: The Jenner Institute, University of Oxford, Oxford, United Kingdom.
authors.forename: Adrian V S
authors.initials: AV
authors.lastname: Hill
authors.affiliation: The Jenner Institute, University of Oxford, Oxford, United Kingdom.
authors.forename: Alexandra J
authors.initials: AJ
authors.lastname: Spencer
mesh_terms: D000818:Animals; D000953:Antigens, Protozoan / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D002460:Cell Line; D005260:Female; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D066298:In Vitro Techniques; D008099:Liver / Q000276:immunology / Q000469:parasitology; D008288:Malaria / Q000276:immunology / Q000517:prevention & control; D017780:Malaria Vaccines / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D010962:Plasmodium berghei / Q000235:genetics / Q000276:immunology* / Q000472:pathogenicity; D015800:Protozoan Proteins / Q000276:immunology*; D013154:Spleen / Q000166:cytology / Q000276:immunology; D034101:Sporozoites / Q000276:immunology
chemical_list: D000953:Antigens, Protozoan; D017780:Malaria Vaccines; D015800:Protozoan Proteins; C069428:thrombospondin-related adhesive protein, protozoan; D049452:Green Fluorescent Proteins
references.citation: Infect Immun. 1994 Jul;62(7):2979-83
references.pmid: 8005684
references.citation: Mol Biochem Parasitol. 2004 Sep;137(1):23-33
references.pmid: 15279948
references.citation: Science. 2001 Jan 5;291(5501):141-4
references.pmid: 11141568
references.citation: Nature. 1989 Sep 28;341(6240):323-6
references.pmid: 2477703
references.citation: Cell. 1997 Aug 8;90(3):511-22
references.pmid: 9267031
references.citation: Exp Parasitol. 1987 Jun;63(3):345-51
references.pmid: 2438152
references.citation: Science. 1986 Jan 10;231(4734):156-9
references.pmid: 3510455
references.citation: PLoS One. 2010;5(2):e9156
references.pmid: 20161781
references.citation: J Virol. 2000 Nov;74(21):10074-80
references.pmid: 11024136
references.citation: Trans R Soc Trop Med Hyg. 1990 May-Jun;84(3):325-9
references.pmid: 2175464
references.citation: J Cell Biol. 1999 Nov 29;147(5):937-44
references.pmid: 10579715
references.citation: Cell Microbiol. 2008 Jan;10(1):218-24
references.pmid: 17697130
references.citation: J Virol Methods. 2005 Sep;128(1-2):104-12
references.pmid: 15908019
references.citation: J Immunol. 1984 Feb;132(2):909-13
references.pmid: 6317752
references.citation: Infect Immun. 2001 Jun;69(6):3845-52
references.pmid: 11349050
references.citation: J Infect Dis. 2007 Aug 15;196(4):608-16
references.pmid: 17624848
references.citation: Int J Parasitol. 2007 Feb;37(2):173-82
references.pmid: 17112526
references.citation: Methods Mol Biol. 2013;923:385-400
references.pmid: 22990793
references.citation: PLoS One. 2013 Jun 18;8(6):e65960
references.pmid: 23823332
references.citation: Am J Trop Med Hyg. 2006 May;74(5):708-15
references.pmid: 16687667
references.citation: Malar J. 2013;12:394
references.pmid: 24191920
references.citation: Int Immunol. 1991 Jun;3(6):579-85
references.pmid: 1716146
references.citation: Cell Host Microbe. 2013 Jul 17;14(1):104-15
references.pmid: 23870318
references.citation: J Immunol. 1999 Jul 15;163(2):884-92
references.pmid: 10395683
references.citation: Parassitologia. 1993 Jul;35 Suppl:69-72
references.pmid: 8233617
references.citation: Mol Ther. 2012 Aug;20(8):1633-47
references.pmid: 22354374
references.citation: Am J Trop Med Hyg. 1995 Dec;53(6):607-11
references.pmid: 8561262
references.citation: J Immunol. 1992 Sep 15;149(6):2103-9
references.pmid: 1517574
references.citation: Infect Immun. 2014 Mar;82(3):1277-86
references.pmid: 24379295
references.citation: Immunol Cell Biol. 2013 Oct;91(9):545-55
references.pmid: 24018532
references.citation: Science. 1985 Jan 25;227(4685):440-2
references.pmid: 3880923
references.citation: PLoS Pathog. 2009 Jan;5(1):e1000265
references.pmid: 19148273
references.citation: Mol Microbiol. 2002 Aug;45(3):637-51
references.pmid: 12139612
references.citation: J Hepatol. 2008 Jan;48(1):51-60
references.pmid: 17998148
references.citation: J Immunol. 2009 Nov 1;183(9):5870-8
references.pmid: 19812194
references.citation: Nat Med. 2008 Aug;14(8):819-21
references.pmid: 18660818
references.citation: Eur J Immunol. 2008 Mar;38(3):732-41
references.pmid: 18266272
references.citation: Nat Commun. 2013;4:2836
references.pmid: 24284865
references.citation: Vaccine. 2014 Apr 17;32(19):2135-8
references.pmid: 24582635
references.citation: Mol Biochem Parasitol. 2012 Dec;186(2):143-7
references.pmid: 23107927
references.citation: J Exp Med. 1994 Jul 1;180(1):353-8
references.pmid: 7516412
references.citation: Biotechnol Bioeng. 2012 Mar;109(3):719-28
references.pmid: 22252512
references.citation: J Infect Dis. 2012 Mar 1;205(5):772-81
references.pmid: 22275401
references.citation: Bull World Health Organ. 1990;68 Suppl:52-9
references.pmid: 2094592
references.citation: Gastroenterology. 2000 Oct;119(4):1104-12
references.pmid: 11040197
references.citation: J Immunol. 1993 Feb 15;150(4):1471-8
references.pmid: 7679428
references.citation: Proc Natl Acad Sci U S A. 1991 Sep 15;88(18):7963-7
references.pmid: 1680235
references.citation: J Exp Med. 1990 Mar 1;171(3):763-73
references.pmid: 1689762
references.citation: J Immunol Methods. 2003 Oct 1;281(1-2):65-78
references.pmid: 14580882
references.citation: PLoS Pathog. 2013 May;9(5):e1003303
references.pmid: 23675294
references.citation: Proc Natl Acad Sci U S A. 1988 Jan;85(2):573-6
references.pmid: 2963334
references.citation: Science. 1989 Jun 2;244(4908):1078-81
references.pmid: 2524877