Relationships between IL-17(+) subsets, Tregs and pDCs that distinguish among SIV infected elite controllers, low, medium and high viral load rhesus macaques

Abstract

Comprehensive studies of the frequencies and absolute numbers of the various cell lineages that synthesize IL-17 in the blood and corresponding gastrointestinal (GI) tissues, their correlation with CD4(+) Tregs, CD8(+) Tregs, total and IFN-α synthesizing plasmacytoid dendritic cells (pDC) relative to plasma viral load in SIV infection has been lacking. The unique availability of SIV infected rhesus macaques (RM) classified as Elite Controllers (EC), and those with Low, Intermediate and High Viral Loads (HVL) provided a unique opportunity to address this issue. Results of these studies showed that EC demonstrated a remarkable ability to reverse changes that are induced acutely by SIV in the various cell lineages. Highlights of the differences between EC and HVL RM within Gastro-intestinal tissues (GIT) was the maintenance and/or increases in the levels of IL-17 synthesizing CD4, CD8, and NK cells and pDCs associated with slight decreases in the levels of CD4(+) Tregs and IFN-α synthesizing pDCs in EC as compared with decreases in the levels of IL-17 synthesizing CD4, CD8 and NK cells associated with increases in pDCs and IFN-α synthesizing pDCs in HVL monkeys. A previously underappreciated role for CD8(+) Tregs was also noted with a moderate increase in ECs but further increases of CD8(+) Tregs with increasing VL in viremic monkeys. Positive correlations between plasma VL and decreases in the levels of Th17, Tc17, NK-17, CD4(+) Tregs and increases in the levels of CD8(+) Tregs, total and IFN-α synthesizing pDCs were also noted. This study also identified 2 additional IL-17(+) subsets in GIT as CD3(-/)CD8(+)/NKG2a(-) and CD3(+)/CD8(+)/NKG2a(+) subsets. Studies also suggest a limited role for IFN-α synthesizing pDCs in chronic immune activation despite persistent up-regulation of ISGs. Finally, elevated persistent innate immune responses appear associated with poor prognosis. These findings provide an initial foundation for markers important to follow for vaccine design.

Figure 1. Analysis of Th17⁺ cells in PBMC and corresponding colo-rectal biopsies from SIV infected nonhuman primates.

Frequencies (%) and absolute numbers (#) of CD4⁺ Th17 cells in the PBMC and colorectal biopsies of groups of uninfected (Uninf), and SIV infected rhesus macaques classified as Elite Controllers (EC), those with plasma levels of Low Viral Loads (LVL), Intermediate Viral Loads (IVL), and High Viral Loads (HVL) in the left panels. The right panels reflect the same measurements in SIV seronegative (Negative) and SIV seropositive (Positive) sooty mangabeys assayed in parallel. A) Reflects the data on % CD4⁺ Th17 cells in PBMC B) the absolute number of CD4⁺ Th17 cells C) the % CD4⁺ Th17 cells in colorectal biopsy tissues and D) the % CD4⁺ T cells in the same aliquot of biopsies. Statistically significant data are reflected by the number of asterisks with p<0.02 (*). p<0.05 (**), p<0.01 (***), p<0.001 (****) and p<0.0001 (*****) for all figures.

Figure 2. Analysis of Tc17 cells in PBMC and corresponding colo-rectal biopsies from SIV infected nonhuman primates.

Nomenclature identical to Fig. 1 except the data reflects values for CD8⁺ IL-17 cells (Tc-17).

Figure 3. Analysis of NK-17 cells in PBMC and corresponding colo-rectal biopsies from SIV infected nonhuman primates.

Nomenclature identical to Fig. 1 except the data reflects values for CD3⁻, CD8⁺, NKG2a⁺ (NK) cells.

Figure 4. Analysis of novel IL-17⁺ cell subsets cells in colo-rectal biopsies from SIV infected nonhuman primates.

Frequencies (%) of CD3⁻, CD8⁺, NKG2a⁻ and the CD3⁺, CD8⁺, NKG2a⁺ subsets of cells in the colo-rectal biopsy tissues from groups of uninfected rhesus macaques, SIV infected rhesus macaques classified as Elite Controllers (EC), those with LVL, IVL and HVL in the left panels. The right panels reflect data on seronegative (Negative) and SIV infected seropositive (Positive) sooty mangabeys. A) Frequencies of the CD3⁻, CD8⁺, NKG2a⁻ subset of NK cells that are IL-17⁺ B) Frequencies of the parent CD3⁻, CD8⁺, NKG2a⁻ cells from the same specimen. C) Frequencies of CD3⁺, CD8⁺, NKG2a⁺ that are IL-17⁺ D) Frequencies of the parent CD3⁺, CD8⁺, NKG2a⁺ cells from the same specimen.

Figure 5. Analysis of CD4⁺ Tregs cells in PBMC and corresponding colo-rectal biopsies from SIV infected nonhuman primates.

Nomenclature identical to Fig. 1 except the data reflects values for CD4⁺ Tregs. Please note D) reflects the frequencies of total CD4⁺ T cells in the same sample.

Figure 6. Analysis of CD8-Tregs cells in PBMC and corresponding colo-rectal biopsies from SIV infected nonhuman primates.

Nomenclature identical to Fig. 1 except the data reflects values for CD8⁺ Tregs.

Figure 7. Analysis of pDCs in the PBMC and corresponding colo-rectal biopsies from SIV infected nonhuman primates.

The frequencies and absolute number of CD123⁺ on the gated population of HLA-DR⁺ Lin⁻ cells (pDCs) in the PBMC and colo-rectal biopsy tissues of the same group of animals as outlined in Fig. 1. A) The frequencies of pDCs in the PBMC B) Absolute number of pDCs in the PBMC C) The frequencies of pDCs in the colo-rectal biopsy tissues from the same animals.

Figure 8. Analysis of IFN-α⁺ pDC cells in PBMC and corresponding colo-rectal biopsies from SIV infected nonhuman primates.

The frequencies of IFN-α synthesizing pDCs in the PBMC (A) and colo-rectal biopsy tissues (B) of the same group of animals as depicted in Fig. 1. The blank bars reflect the frequencies of pDCs that constitutively express IFN-α and the dark bars reflect the frequencies of pDCs that are induced to express IFN-α.

Figure 9. Kinetics of total and IFN-α⁺ pDCs in PBMC and colo-rectal biopsies from SIV infected macaques.

The kinetics of the frequencies of A) pDCs in the PBMC, B) pDCs in the corresponding colo-rectal biopsies (C) the frequencies of the pDC shown in (A) that were IFN-α⁺ in the PBMC and D) the frequencies of pDCs that were shown in (B) that were IFN-α positive in the colo-rectal biopsy tissues. Data was obtained on a cohort of rhesus macaques prior to (Uninfected) and post intravenous infection with 1000 TCID50 of SIVmac239 collected on days 2–4, 7, 14, 21, 28 and 56 p.i. The monkeys were retrospectively classified as EC, LVL, IVL and HVL after they reached viral load set point and reached the chronic stage.