Regulatory T cell abnormalities associated with aberrant CD4+ T-cell responses in HIV+ patients with immune reconstitution disease (IRD) Nabila SEDDIKI National Centre in HIV Epidemiology and Clinical Research, The University of NSW, Centre For Immunology

Immune Reconstitution Disease Anti-retroviral therapy (ART) may have adverse effects in HIV patients who start when very immunodeficient (usually within two months); this results in an excessive inflammatory response referred to immune reconstitution disease (IRD) IRD occurs more frequently in HIV-infected patients : Who start ART with less than 100 CD4 (cells/L) Who have more than 1 log10 viral load decline from baseline (< 2 months) With prior opportunistic infection history (some patients with NO OI have been reported) IRD has been associated with bacteria, viruses and fungi but mycobacterial IRD has been defined as an important cause of mortality We postulated that deficits in T regulatory cell (Treg) numbers/function may be defective in these patients and are not able to turn off the excessive T cell responses

Patients characteristics In a cross-sectional study including HIV+ patients with less than 50 CD4 (cells/l) before ART: 8 with mycobacterial IRD and 6 without IRD at wk0, wk4 and wk48 after starting ART

IRD CD4+ T cells secrete high levels of IL-2 and IFN- Intra cellular staining for cytokine ELISpot IFN-

High % of Foxp3+CD25+CD127lo CD4+ Treg in IRD vs healthy

Treg in IRD proliferate extensively ex vivo In vitro *High IL-2 secretion in IRD may help Treg survival

High proportion of CD4+ CD127lo cells in IRD vs HIV+ patients w/o IRD CD4+ CD25+ CD127lo CD4+ CD25- CD127lo CD25 CD4+ CD45RO+ CD25+ CD127lo CD25 CD4+ CD45RO+ CD25+ CD127hi

High IL-7, IL-6 and IL-4 cytokine level in IRD vs controls - High IL-7 level might be due to the absence or decrease of its receptor IL-7R (CD127)

IL-7 level remains high in IRD after immune reconstitution

Treg in IRD do not suppress efficiently CD4+ T cell proliferation Healthy control Patient - Irr APCs - anti-CD3 + 72h culture 3HTdR T cell proliferation CD25 Sp:Rc Sc:Rc Sc:Rp Sp:Rp CD127 - Treg from 3 patients have a defective function - CD4+ effectors from patient #3 are resistant to suppression

Treg in IRD do not suppress efficiently inflammatory cytokine secretion In accordance with suppression of proliferation data … residual cytokine secretion (pg/ml) - Treg from 3 patients have a defective function - CD4+ effectors from patient #3 are resistant to suppression Abstract #: MOPEA020

Treg in IRD do not produce sufficient IL-10 to suppress high inflammatory cytokine secretion residual cytokine secretion (pg/ml) Sufficient IL-10, regulatory cytokine, production is compromised in IRD patients probably due to the presence of high effector/inflammatory cytokine (IFN-, IL-6 and TNF-)

IL-7, IL-4 and IL-15 inhibit suppressive Treg function Increased levels of IL-2, IL-4 and IL-7 found in the plasma of patients with IRD may help the pathogenic high proliferation of CD4+ T cell observed in these patients

Summary Significant expansion of CD127loFoxp3+CD25+Treg in HIV+ patients with IRD compared to HIV+ w/o IRD higher ratio Treg to effector/memory subsets in IRD compared to patients w/o IRD High proportion of CD4+ T cells with low IL-7R expression explaining probably the high IL-7 level in the plasma of these patients In vitro suppression assays demonstrated abnormalities in the functional capacity of both suppressors and responders from these patients, and interestingly IL-10 secretion was diminished in patients Increased levels of IL-4, IL-6 and IL-7 in plasma. Some of these cytokines strongly inhibited Treg suppression in vitro Despite the high Treg expansion in IRD, their ability to induce suppression and down regulate aberrant immune responses is compromised, probably due to an imbalance in regulatory versus effector cytokine (IFN- to IL-10) Abstract #: MOPEA020

Acknowledgements NHMRC Centre For Immunology and NCHECR Sarah Sasson John Zaunders Mee Ling Munier David van Bockel Suzanna Ip David Cooper Anthony Kelleher - St Vincents Hospital Patients Sarah Pett Debbie Mariott - Nepean Hospital Brigitte Nanan NHMRC