European Journal of Medical Research - Deutsche AIDS ...

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June 27, 2007 EUROPEAN JOURNAL OF MEDICAL RESEARCH 123 phase of infection, their importance has remained unclear so far. Studies of CD8 T cell responses based on interferon-g production have shown persistence of these in progressive infection and failed to demonstrate a difference between controllers and progressors. We therefore studied different effector functions as well as activation status, proliferation capacity and apoptosis markers of CD8 T cell responses in individuals at different stages of HIV infection. Methods: 28 subjects with chronic, untreated HIV infection stratified into 5 groups according to viral load were studied. Intracellular cytokine staining using synthetic peptides as antigenic stimulus was used to study production of interferong, TNF-a and IL-2. Tetramer staining was used for content of perforin and granzyme B as well as CD38, HLA-DR, CD57, Ki67, cell cycle analysis, Bcl-2 and CD95. Results: We did not find a significant difference between the five groups for antigen specific production of interferon-g, TNF-a and IL-2. Neither was there a difference in granzyme B and perforin content as outread for the effector function cytolysis. We found highly significant differences between controllers and the four progressor groups for immune activation by CD38 staining (p < 0.05 for all groups) with a VL>200,000 cp/ml having the most CD38 positive cells. The difference was even more pronounced for HIV specific CD8 T cells than for bulk CD8 T cells (p < 0.05). Immune activation of HIV specific cells according to CD38 was correlated with viral load (r2 = 0.51, p < 0.0001) and inversely correlated with CD4 counts (r2 = 0.52, p < 0.0001). We then failed to find differences between markers for cell proliferation as well as for protection of apoptosis (Bcl-2) or death receptors (CD95) of HIV specific cells. Conclusion: Our study showed a highly significant difference in immune activation as outlined by CD38 staining. But neither enhanced proliferation nor the aptitude for apoptosis can explain the consequences of immune activation and therefore the reason for the difference in immune activation between controllers and progressors. F.12 (Vortrag) Interleukin-2 augments dendritic cells numbers in lymphoid tissue Stellbrink H.-J. 1 , Tenner-Racz K. 2 , van Lunzen J. 3 , Schneider C. 4 , Racz P. 2 1 ICH Hamburg, Hamburg, Germany, 2 Bernhard-Nocht- Institut, Abt. für Pathologie, Hamburg, Germany, 3 Ambulanzzentrum des Universitätsklinikums Hamburg- Eppendorf, Bereich Infektiologie, Hamburg, Germany, 4 Universitätsklinikum Hamburg-Eppendorf, Klinik und Poliklinik für Allgemein-, Viszeral- und Thoraxchirurgie, Hamburg, Germany Background: Autologous dendritic cells have been used for therapeutic vaccination in HIV infected individuals. IL-2 represents a potential adjuvant for therapeutic vaccination in this setting. We therefore investigated if HAART +/- IL-2 impacts on immature (CD1a+ or CD207 [Langerin]+) as well as mature (CD208 [DC-LAMP]+ or CD83+) dendritic cell numbers in the lymphoid tissue. Methods: 15 chronically HIV-1 infected, antiretroviral naive subjects (group 1: HAART, n=8; group 2: HAART + interleukin-2, n=7; all male) were enrolled in a prospective, randomised, controlled clinical trial on HAART +/- IL-2 (COS- MIC). Serial axillary lymph node biopsies were performed at baseline and after at least six months of undetectable plasma viremia. Subjects were selected according to the availability of suitable frozen tissue for the analysis. CD1a+ or Langerin+ immature as well as CD83+ or DC-LAMP+ mature DCs were detected on frozen sections by immunohistochemistry. They were enumerated by PC-based image analysis and expressed as cells per unit area, based on the analysis of 10 non-overlapping fields. Results were analysed by t-test, Mann-Whitney U test, and MANOVA, where appropriate. Results: Peripheral blood CD4+ T cells were 603 ±165 and 477 ± 176 in groups 1 and 2, resp. (p=n.s.) Plasma viremia was 4.3 ±0.61 and 4.53 ± 0.55 log10, resp. (p=n.s.). CD4+ cells increased in both groups (p=0.00004) and were higher in the IL-2 group 2 at the 2nd biopsy (p=0.04, MANOVA). Time on treatment until 2nd biopsy (370 vs. 365 days) did not differ between the groups (p=0.86, t-test). Baseline CD1a+ and DC- LAMP+ cell numbers were higher in group 2 (p=0.0031 and 0.04, resp., t-test). CD1a+, Langerin+, DC-LAMP+, and CD83+ cells increased in both groups (all p
124 EUROPEAN JOURNAL OF MEDICAL RESEARCH June 27, 2007 probably plays an important role in the pathological expression of this chemokine. (1) Lehmann MH, Masanetz S, Kramer S, Erfle V. (2006). J. Cell Sci. 119(21): 4520-4530. (2) Eugenin, E. A., Osiecki, K., Lopez, L., Goldstein, H., Calderon, T. M. and Berman, J. W. (2006). J. Neurosci. 26, 1098-1106. F.14 (Vortrag) Suppressive HAART abrogates HIV-induced intestinal barrier impairment and mucosal immune activation Epple H.-J. 1 , Schneider T. 1 , Tröger H. 1 , Kunkel D. 1 , Allers K. 1 , Moos V. 1 , Fromm M. 2 , Zeitz M. 1 , Schulzke J.-D. 1 1 Charité, Campus Benjamin Franklin, Med. Klin. I, Gastroenterologie, Infektiologie, Rheumatologie, Berlin, Germany, 2 Charité, Campus Benjamin Franklin, Institut für Klinische Physiologie, Berlin, Germany Objective: A barrier defect of the intestinal mucosa has been suggested as central mechanism of the hyper immune activation characteristic for chronic HIV infection. However, although highly effective antiretroviral therapy (HAART) reduces the general immune activation in HIV-infected patients, the effect of HAART on the HIV-induced epithelial barrier defect is unknown as are its underlying mechanisms. In this study we investigated the effect of suppressive HAART on the HIV-induced intestinal barrier defect and mucosal cytokine secretion. Methods: Epithelial barrier function was characterized in duodenal biopsies obtained from 11 untreated, 8 suppressively treated HIV-infected patients, and 9 HIV-seronegative controls by impedance spectroscopy and 3H-mannitol fluxes. Villus/crypt ratio was determined microscopically. Epithelial apoptotic rate was analyzed by TUNEL staining. Expression of tight junction proteins was quantified by densitometric analysis of immunoblots. Mucosal cytokine production was determined by cytometric bead array. Results: Untreated but not treated HIV infection was associated with a marked reduction of the epithelial resistance, an increase of mannitol permeability and pronounced villus atrophy. Accordingly, mucosal production of interleukin (IL)-2, interferon-g, IL-4, IL-5, and tumor necrosis factor-a was increased in untreated but not in treated HIV patients. Furthermore, in treated patients epithelial apoptosis was increased, the expression of the sealing tight junction protein claudin-1 was reduced and that of the pore-forming claudin-2 was increased. Conclusions: The HIV-induced intestinal barrier defect and villus atrophy are reversed by suppressive HAART. Accordingly, suppressive HAART strongly reduces the mucosal production of inflammatory cytokines which is highly increased in untreated HIV-infected patients. Finally, we identified increased epithelial apoptosis and an altered tight junction composition as mechanisms of the HIV-induced barrier defect. Our data indicate that interruption of the self-perpetuating cycle of mucosal immune activation by suppressive HAART leads to reconstitution of the epithelial barrier function and may thus contribute to the reduction of systemic immune activation. F.15 (Poster) Pronounced increase of mucosal FoxP3+ regulatory CD4+ T cells early after the start of ART in chronically SIV infected macaques Allers K. 1 , Loddenkemper C. 2 , Epple H.-J. 1 , Stahl-Hennig C. 3 , Schrod A. 3 , Sauermann U. 3 , Verena M. 1 , Schneider T. 1 1 Medical Clinic I, Charité - Campus Benjamin Franklin, Gastroenterology, Infectious Diseases and Rheumatology, Berlin, Germany, 2 Medical Clinic I, Charité - Campus Benjamin Franklin, Institute of Pathology, Berlin, Germany, 3 German Primate Centre, Department of Virology and Immunology, Goettingen, Germany HIV infection is associated with chronic immune activation and progressive loss in the number and function of CD4+ T cells. Antiretroviral therapy (ART) generally results in a reconstitution of T cells. However, in a small fraction of patients receiving ART a hyperinflammatory disorder occurs despite decreased viral load and reconstitution of T cells. The mechanisms responsible for this immune inflammatory reconstitution syndrom (IRIS) are unknown. Naturally occurring regulatory CD4+ T cells (Treg) constitutively express CD25 and FoxP3 and have recently been described as a subset of T cells with immunosuppressive properties. Previously we found evidences for an important role of mucosal Treg in untreated HIV infection and observed a return of Treg numbers to normal levels after ART. However, the dynamics of Treg normalization following ART are unknown. In this study, we analyzed the peripheral and mucosal lymphocyte subset distribution in the course of untreated and treated SIVmac251 infection in six rhesus macaques by flow cytometry. Quantifications were performed by immunohistochemistry and BD TruCountTM. In addition, functional tests were performed. Peripheral Treg decreased during untreated SIV infection and normalized after start of ART. In contrast, mucosal Treg increased and reached highest levels early after initiation of ART. We did not find significant correlations between Th1 or Th2 cytokines in the gut mucosa and increasing Treg. Thus, one regulatory function of Treg could consist in sustaining levels of Th1 and Th2 cytokines that hold immune activation and inflammation in a non-detrimental state. CD4+ T cell response against SIV peptides was restricted to the acute phase of infection and not restored after start of ART. Interestingly, there was a highly significant inverse correlation between the SIV specific response in the peripheral blood and the frequency of mucosal Treg. Therefore, we suggest that Treg located at lymphatic sites play an important role in suppressing immune responses. Taken together, our results indicate that Treg accumulate at sites of high immune activation to suppress detrimental hyperactivation. We hypothesize that a loss of Treg induces hyperinflammatory disorder resulting in IRIS in patients receiving ART.
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