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| Status |
Public on Mar 08, 2010 |
| Title |
Lymphatic Tissues of Sooty Mangabeys and Rhesus Macaques in Early SIV Infection |
| Platform organism |
Macaca mulatta |
| Sample organisms |
Cercocebus atys; Macaca mulatta |
| Experiment type |
Expression profiling by array
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| Summary |
Transcriptional Profiling Reveals Distinguishing Features of Immune Activation in the Lymphatic Tissues of Sooty Mangabeys and Rhesus Macaques in Early SIV Infection
Immune activation in the chronic stages of HIV-1 and SIV infection of rhesus macaques is thought to play a critical role in CD4+ T cell depletion and progression to AIDS. Conversely, the lack of immune activation in the chronic stages of SIV infection of sooty mangabeys is thought to protect this species from a pathogenic outcome. This critical difference in immune activation in rhesus macaques and sooty mangabeys has recently been shown to be established in the early stages of SIV infection where there is immune activation in both species, but in sooty mangabeys immune activation is subsequently resolved. Here we report the results of a microarray analysis of early SIV infection in the lymphatic tissues of rhesus macaques and sooty mangabeys that provides a comprehensive view of the distinguishing features of immune activation and host defenses in the two species. We confirm immune activation in sooty mangabeys, but show that it is less robust than in rhesus macaques, where positive feedback loops involving cytosolic pattern recognition receptors, and chemokines and their ligands amplify the response to early SIV infection. We confirm resolution of immune activation in sooty mangabeys, but show it to be partial and selective, identifying the interferon system and CD38 as key correlates of outcome. We also identify potentially novel immunoregulatory mechanisms that mediate resolution, and defensins as host defenses sooty mangabeys employ as expression of other innate immune system effectors declines.
The studies we report here were undertaken in a collaborative longitudinal analysis of early SIV infection in SMs described in Bosinger et al and previously reported studies (Estes et al., J Immunol 1008, 180, 6798-6807.). Briefly, SMs were inoculated i.v. with 1 ml of plasma from an experimentally SIVsmm-infected SM sampled at day 11 post infection, with a viral load 107 copies/ml. In our studies we analyzed axillary or inguinal lymph node biopsies that had been obtained from two SMs prior to infection; two SMs at the peak of viral replication at 14 dpi; and two SMs at 30 dpi as viral loads were decreasing to set point. We analyzed axillary or inguinal LNs from 4 RMs without SIV infection and LNs from four RMs infected intravaginally with 2x105 TCID50 of SIVmac239 and sacrificed at 14 dpi the peak of viral replication from two cross-sectional studies described in (Miller et al., The Journal of Virology 2005, 79, 9217-9227.). For lymph node (LN) biopsies, animals were anesthetized with Ketamine or Telazol; the skin over the axillary or inguinal region was clipped and surgically prepped. An incision was made over the LN, which was exposed by blunt dissection and excised over clamps. A portion of the lymph node biopsy for microarray analysis was snap frozen in liquid N2. All animals were housed and cared for at the Yerkes National Primate Research Center in Atlanta, Georgia in accordance with the regulations of the American Association of Accreditation of Laboratory Animal Care standards. These studies were approved by the Emory University and University of Pennsylvania Institutional Animal Care and Usage Committees (IACUC).
Microarray Analysis
RNA extractions, synthesis of biotin-labeled cRNA probes, microarray hybridization, and data analysis followed previously published procedures (Li et al., The Journal of infectious diseases 2004, 189, 572-582; Li et al., J Immunol 2009, 2009, 183: 1975–1982.). Briefly, snap-frozen lymph node was homogenized in TRIzol. Total RNA was isolated and further purified. Double stranded cDNA and biotin-labeled cRNA probes were synthesized, column purified and fragmented. Fifteen micrograms of fragmented cRNA was hybridized to an Affymetrix GeneChip® Rhesus Macaque Genome Array. After hybridization, chips were washed, stained with streptavidin-phycoerythrin, and scanned with GeneChip Operating Software at the Biomedical Genomics Center at the University of Minnesota. Preparation of cRNA probes and microarray hybridizations were done in duplicate for each RNA sample.
Cel. files were uploaded into the Expressionist program (Genedata, Pro version 5.1) and the expression level for each of the 47,000 transcripts in the arrays were analyzed using the RMA algorithm. The expression levels from duplicate microarrays of the same animal’s RNA were correlated and averaged. Tests for differences between the before and after infection at various time points were conducted using the 2-sample Wilcoxon signed-rank test. Fold differences in the level of gene expression between after infection and before infection were calculated with the ratio of the means. After statistical analysis, data was sorted based on these transcript cutoffs: p-value of < 0.05 and fold change ≥ 2.0. Significantly changed genes and transcripts were uploaded into NetAffix Analysis Center (http://www.affymetrix.com/analysis/index.affx) to query gene ontology information and into Ingenuity Pathways Analysis (Ingenuity® Systems, www.ingenuity.com) for gene annotation and pathway analysis. Hierarchical clustering analysis was carried out by using Spotfire for DecisionSite for Functional Genomics.
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| Overall design |
We used affymetrix rhesus microarrays to profile the host transcriptional response to SIV in lymph nodes obtained from SM (Sooty mangabeys, Cercocebus atys)) and RMs ( Rhesus Macaques, Macaca mulatta) in the first two weeks of infection in studies described by Bosinger et al. (GEO accession number GSE16147) of the transcriptional profiles in peripheral blood. We confirm in these genome-wide comparisons that immune activation is a shared feature of early infection in the two species, but show that the breadth of the changes in RMs is much greater than in SMs, with evidence of positive feedback loops that amplify innate and cellular responses in RMs. We also confirm evidence of resolution of immune activation in SMs, but were surprised to find little evidence at the transcriptional level of increased expression of genes that might mediate that resolution. Lastly, we show that as expression of genes involved in host defenses decline in SMs, there is increased expression of defensins that again may represent a species-specific host defense mechanism. Samples: 12 Sooty Mangabeys (uninfected, 14 dpi, and 30 dpi) and 16 Rhesus Macaques (uninfected and 14 dpi).
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| Contributor(s) |
Li Q, Duan L, Smith AJ, Zeng M, Carlis JV, Silvestri G, Haase AT |
| Citation(s) |
19959874 |
| |
| Submission date |
Aug 13, 2009 |
| Last update date |
Jul 18, 2012 |
| Contact name |
qingsheng li |
| E-mail(s) |
lixxx076@umn.edu
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| Phone |
612-624-9130
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| Fax |
612-626-0623
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| Organization name |
University of Minnesota
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| Department |
Microbiology
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| Lab |
Ashley T. Haase
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| Street address |
Box 196, 420 Delaware ST SE
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| City |
minneapolis |
| State/province |
MN |
| ZIP/Postal code |
55455 |
| Country |
USA |
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| Platforms (1) |
| GPL3535 |
[Rhesus] Affymetrix Rhesus Macaque Genome Array |
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| Samples (28)
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| GSM440057 |
Cercocebus atys uninfected LN-FRs-replicate-1 |
| GSM440058 |
Cercocebus atys uninfected LN-FRs-replicate-2 |
| GSM440059 |
Cercocebus atys uninfected LN-Fuv-replicate-1 |
| GSM440060 |
Cercocebus atys uninfected LN-Fuv-replicate-2 |
| GSM440061 |
Cercocebus atys 14 dpi SIVsmm LN-FRs-replicate-1 |
| GSM440062 |
Cercocebus atys 14 dpi SIVsmm LN-FRs-replicate-2 |
| GSM440063 |
Cercocebus atys 14 dpi SIVsmm LN-FWv-replicate-1 |
| GSM440064 |
Cercocebus atys 14 dpi SIVsmm LN-FWv-replicate-2 |
| GSM440065 |
Cercocebus atys 30 dpi SIVsmm LN-Fuv-replicate-1 |
| GSM440066 |
Cercocebus atys 30 dpi SIVsmm LN-Fuv-replicate-2 |
| GSM440067 |
Cercocebus atys 30 dpi SIVsmm LN-FWv-replicate-1 |
| GSM440068 |
Cercocebus atys 30 dpi SIVsmm LN-FWv-replicate-2 |
| GSM440069 |
Macaca mulatta uninfected LN-RYe8-replicate-1 |
| GSM440070 |
Macaca mulatta uninfected LN-RYe8-replicate-2 |
| GSM440071 |
Macaca mulatta uninfected LN-RF8-replicate-1 |
| GSM440072 |
Macaca mulatta uninfected LN-RF8-replicate-2 |
| GSM440073 |
Macaca mulatta uninfected LN-34550-replicate-1 |
| GSM440074 |
Macaca mulatta uninfected LN-34550-replicate-2 |
| GSM440075 |
Macaca mulatta uninfected LN-31391-replicate-1 |
| GSM440076 |
Macaca mulatta uninfected LN-31391-replicate-2 |
| GSM440077 |
Macaca mulatta 14 dpi SIVmac239 LN-R00034-replicate-1 |
| GSM440078 |
Macaca mulatta 14 dpi SIVmac239 LN-R00034-replicate-2 |
| GSM440079 |
Macaca mulatta 14 dpi SIVmac239 LN-24037-replicate-1 |
| GSM440080 |
Macaca mulatta 14 dpi SIVmac239 LN-24037-replicate-2 |
| GSM440081 |
Macaca mulatta 14 dpi SIVmac239 LN-R93047-replicate-1 |
| GSM440082 |
Macaca mulatta 14 dpi SIVmac239 LN-R93047-replicate-2 |
| GSM440083 |
Macaca mulatta 14 dpi SIVmac239 LN-R98032-replicate-1 |
| GSM440084 |
Macaca mulatta 14 dpi SIVmac239 LN-R98032-replicate-2 |
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| Relations |
| BioProject |
PRJNA118617 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE17626_RAW.tar |
237.5 Mb |
(http)(custom) |
TAR (of CEL) |
| Processed data included within Sample table |
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