Quantitation of simian cytokine and beta-chemokine mRNAs, using real-time reverse transcriptase-polymerase chain reaction: variations in expression during chronic primate lentivirus infection.
Cytokines and beta-chemokines are important mediators of the immune system and are expressed in many infectious diseases. To study cytokine and beta-chemokine profiles during pathogenesis of lentiviral infection and progression to AIDS in rhesus macaques, we established new quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assays based on TaqMan chemistry. Using synthetic RNA standards, we quantified mRNAs of IL-2, IL-4, IL-6, IL-10, IL-12 p40, interferon gamma (IFN-gamma), tumor necrosis factor alpha (TNF-alpha), RANTES, macrophage inflammatory protein 1 alpha (MIP-1 alpha), and MIP-1 beta in unstimulated peripheral blood mononuclear cells (PBMCs) and lymph nodes from macaques chronically infected with SIV or SHIV. Viremic monkeys with decreased CD4(+) T cell counts (<500 cells/microl) had significantly higher IL-10 mRNA expression than uninfected controls, which parallels the findings in HIV-1-infected humans. In addition, MIP-1 alpha, MIP-1 beta, and RANTES mRNA expression increased in viremic monkeys with decreased CD4(+) T cell counts; gene expression was inversely correlated with CD4(+) T cell counts, but not viral load. The newly established quantitative real-time RT-PCR assays will allow the determination of cytokine and beta-chemokine patterns in rhesus macaques in studies of microbial pathogenesis or vaccine development.
Hofmann-Lehmann
Regina
R
Dana-Farber Cancer Institute and Department of Medicine, Harvard Medical School, Boston, Massachusetts 02115, USA.
Williams
Alison L
AL
Swenerton
Ryan K
RK
Li
Pei-Lin
PL
Rasmussen
Robert A
RA
Chenine
Agnès-Laurence
AL
McClure
Harold M
HM
Ruprecht
Ruth M
RM
D000818:Animals; D018791:CD4 Lymphocyte Count; D054405:Chemokine CCL3; D054407:Chemokine CCL4; D019742:Chemokines, CC / Q000032:analysis* / Q000235:genetics; D016207:Cytokines / Q000032:analysis* / Q000235:genetics; D018450:Disease Progression; D015870:Gene Expression; D016753:Interleukin-10 / Q000032:analysis / Q000235:genetics; D017958:Lentiviruses, Primate; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008198:Lymph Nodes / Q000166:cytology / Q000378:metabolism; D008253:Macaca mulatta; D019402:Macrophage Inflammatory Proteins / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016097:Simian Acquired Immunodeficiency Syndrome / Q000276:immunology / Q000378:metabolism*; D019562:Viral Load
D054405:Chemokine CCL3; D054407:Chemokine CCL4; D019742:Chemokines, CC; D016207:Cytokines; D019402:Macrophage Inflammatory Proteins; D012333:RNA, Messenger; D016753:Interleukin-10