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Articles by ANRS O86 PRIMOFERON A Study Group, Related Collections Immunobiology Brief Reports Social Bookmarking                   What's this?  Previous Article  |  Table of Contents  |  Next Article  Blood, 15 November 2001, Vol. 98, No. 10, pp. 3169-3171 BRIEF REPORT Preferential and persistent depletion of CCR5+ T-helper lymphocytes with nonlymphoid homing potential despite early treatment of primary HIV infection Roman Krzysiek, Annick Rudent, Laurence Bouchet-Delbos, Arnaud Foussat, Claudie Boutillon, Alain Portier, Didier Ingrand, Daniel Sereni, Pierre Galanaud, Liliane Grangeot-Keros, Dominique Emilie, and the ANRS O86 PRIMOFERON A Study Group From the INSERM U131 and Service de Médecine Interne et d'Immunologie Clinique, Service de Microbiologie-Immunologie, Hôpital Antoine Béclère, Institut Paris-Sud sur les Cytokines, Clamart, France, and the Service de Médecine Interne, Hôpital Saint-Louis, Paris, France.     Abstract Top Abstract Introduction Study design Results and discussion References Strains of human immunodeficiency virus (HIV) transmitted between individuals use the CCR5 coreceptor, but no preferential depletion of particular Th-lymphocyte subpopulations has been reported during primary HIV infection (PHI). In contrast, gut-associated Th lymphocytes are preferentially depleted in macaques recently infected by simian immunodeficiency virus. The expression of CCR5 and the intestinal homing receptor integrin 47 on subpopulations of Th lymphocytes was studied in 12 patients with PHI. There was a profound decrease of circulating 47+ Th lymphocytes and CCR5+ memory Th lymphocytes with nonlymphoid homing potential (CD62LCD45RO+). Unlike other Th lymphocytes, this cell population remained depleted despite early control of viral replication under antiretroviral treatment. Therefore, HIV preferentially targets a specific CCR5+ subpopulation of Th lymphocytes early during infection, inducing its persistent depletion despite treatment. Protective immunity in vivo depends on Th lymphocytes carrying homing capacity to nonlymphoid tissue, and therefore these data may explain the persistent abnormalities of immune functions in patients infected with HIV. (Blood. 2001;98:3169-3171) © 2001 by The American Society of Hematology.     Introduction Top Abstract Introduction Study design Results and discussion References Strains of human immunodeficiency virus (HIV) transmitted between individuals use the CCR5 coreceptor. However, the decline in the circulating T-helper (Th) lymphocyte count during primary HIV infection (PHI) involves CCR5 cells, but not CCR5+ cells.1 The rapid recovery of CCR5 Th lymphocyte counts under highly active antiretroviral treatment (HAART) and the limited depletion of Th lymphocytes in lymph nodes of patients with PHI suggest that sequestration of CCR5 Th lymphocytes to lymphoid organs makes a large contribution to their early loss from the blood.1-3 CCR5 is only expressed by memory Th lymphocytes.4 Memory Th lymphocytes can be divided into 2 subpopulations depending on their expression of the CD62L lymph node homing receptor: CD62L+ cells with the lymphoid tissue homing potential (LHP) and CD62L cells with the nonlymphoid tissue homing potential (NLHP).5 Intestinal mucosa-associated Th lymphocytes belong to the NLHP subpopulation and express the intestinal homing receptor integrin 47. Most Th lymphocytes in the gut lamina propria also express CCR5.6 Because the gastrointestinal tract is the major site of viral replication and loss of Th lymphocytes during primary infection with simian immunodeficiency virus (SIV), HIV may similarly target CCR5+ NLHP Th lymphocytes during PHI.7-10 Therefore, analysis of this particular fraction of Th lymphocytes may better reflect HIV-induced depletion of Th lymphocytes than analysis of the entire CD4+ T lymphocyte population. We show that the counts of circulating 47+ and CCR5+ NLHP Th lymphocytes are substantially decreased during PHI. Moreover, the abnormally low counts of CCR5+ NLHP Th lymphocytes persist unchanged for 48 weeks despite HAART, indicating that their early loss is due to early depletion rather than redistribution.     Study design Top Abstract Introduction Study design Results and discussion References Study patients After informed consent, 12 patients with clinical symptoms of PHI were enrolled in the ANRS 086 Primoferon A study.11 The following criteria were used for inclusion: anti-HIV antibodies detectable by enzyme-linked immunosorbent assay or detectable plasma HIV RNA, and 3 bands or fewer in a Western blot test for anti-HIV antibodies. Median plasma HIV RNA concentration at inclusion was 5.8 log10 copies/mL. Antiretroviral treatment was initiated 1 ± 0.5 day (mean ± SEM) after inclusion, combining stavudine (60-80 mg/d), didanosine (300-400 mg/d), and nelfinavir (2500 mg/d) until the end of the follow-up, and pegylated-interferon (IFN)-2b (PEG-Intron, Schering-Plough, Research Institute, Kenilworth, NJ) 1 µg/kg per week for 14 consecutive weeks. Immunofluorescent staining and FACS analysis Peripheral blood mononuclear cells (PBMCs) were isolated at inclusion and after 5, 13, 24, and 48 weeks. They were stained with the following monoclonal antibodies: allophycocyanin (APC)-conjugated anti-CD4 (Becton Dickinson, San Jose, CA), fluorescein isothiocyanate (FITC)-conjugated anti-CCR5, and phycoerythrin (PE)-conjugated anti 7 (both from Pharmingen, San Diego, CA) and PE-conjugated anti-CD45RO, PC5-conjugated anti-CD62L, FITC-conjugated anti-CD49d/4 (all from Immunotech, Marseilles, France) or control immunoglobulins. Multiparametric FACS analysis of lymphocyte subpopulations was performed on 10 000 cells after gating for lymphocytes using dual-laser FACScan and CellQuest software (Becton Dickinson). Statistical analysis Results are median and interquartile range (IQR) values unless otherwise indicated. The unpaired Student t test was used to examine differences between PHI patients and age-matched healthy controls. P values less than .05 were considered as statistically significant.     Results and discussion Top Abstract Introduction Study design Results and discussion References At inclusion, CD4+ cell counts of PHI patients were significantly lower than those of controls (P = .0001; Figure 1A). Both naive (CD62L+CD45RO) and memory (CD45RO+) Th lymphocyte counts were lower in patients (P = .0018 and P = .0001, respectively).12-14 Among memory Th lymphocytes, the counts of both the CD62L+ and CD62L subpopulations were decreased (Figure 1B,C). View larger version (40K): [in this window] [in a new window]   Figure 1. Th lymphocyte subpopulations in PHI patients. Absolute cell counts of CD4+ (A), LHP (CD62L+CD45RO+) Th cells (B), and NLHP (CD62LCD45RO+) Th cells (C) and the percentage of CCR5-expressing LHP Th cells (D), CCR5-expressing NLHP Th cells (E), and 47-expressing Th cells (F) were determined at inclusion in 12 PHI patients (hollow boxes) and 12 age-matched healthy controls (solid boxes). Box plots depict median (horizontal line within box), 75% to 25% IQR (upper and lower limits of the box, respectively), and range (upper and lower horizontal bars outside the box). (G) Dot plots of 7-integrin staining of CD4+ T cells of a representative control individual and a selected PHI patient. *P < .05; **P < .005 by 2-tailed Student t test. The proportion of CD62L+CD45RO+ Th lymphocytes expressing CCR5 was in the same range in patients and controls (Figure 1D). Therefore, CD62L+CD45RO+ Th lymphocyte counts are decreased during PHI independent of their expression of CCR5. In contrast, the percentage of CD62LCD45RO+ Th cells expressing CCR5 was lower in patients than in controls (14.5% versus 25.3%, respectively; P = .036; Figure 1E). The preferential decrease of CD62LCD45RO+ Th lymphocytes expressing CCR5 was associated with a decrease of Th lymphocytes expressing 47+. The 47+ cells made up 27.3% of Th cells in controls, but only 15.8% in patients (P = .0009; Figure 1F). The absolute number of circulating 47+ Th lymphocytes was only 48 × 106/L in patients and 240 × 106/L in healthy controls (P = .0001). In controls, circulating Th lymphocytes expressing high levels of the 7 integrin were enriched in CCR5+ cells as compared to 7 Th lymphocytes (median percentage: 20.8 versus 6.2, respectively; P = .02). There were almost no Th lymphocytes expressing high levels of 7 in patients (P = .0078, as compared to controls; Figure 1G). The HAART combination administered in this study led to an early control of HIV replication and a rapid recovery of peripheral Th cells (Figure 2A).11 We determined whether this restoration involved all subpopulations of Th lymphocytes. The recovery was rapid for naive and CD62L+CD45RO+ Th lymphocytes (not shown), as already reported in patients treated at the chronic phase of the infection.15,16 These findings are consistent with a mechanism of sequestration in lymphoid organs causing the early decrease of most Th lymphocytes.1,17 This recovery was also rapid for CD62LCD45RO+ Th lymphocytes. The proportion of CCR5-expressing cells among CD62L+CD45RO+ Th cells remained within the normal range for 48 weeks (not shown). In contrast, the proportion of CCR5-expressing cells among CD62LCD45RO+ Th lymphocytes remained low, with no tendency for recovery. On week 48, the fraction of CCR5+ cells among CD62LCD45RO+ Th lymphocytes remained significantly lower than that in healthy individuals (P = .012, Figure 2B), and the absolute number of circulating CCR5+CD62LCD45RO+ Th cells was only 30% that of controls (P = .013; Figure 2C). The 47+ Th lymphocyte counts increased during HAART, but their absolute number did not fully recover (P = .01 on week 24 as compared to controls; Figure 2D). View larger version (30K): [in this window] [in a new window]   Figure 2. Persistent depletion of CCR5-expressing NLHP Th lymphocytes in PHI patients undergoing HAART treatment. The median (IQR) CD4+ T cell count (A), the median (IQR) percentage of CCR5-expressing NLHP Th cells (B), the median (IQR) CCR5-expressing NLHP Th cell count (C), and the median (IQR) 47+ Th cell count (D) were followed in 12 PHI patients treated with HAART. Shaded areas represent the 75% to 25% IQR values for healthy controls. The solid symbol in the shaded area represents median for the controls. *P < .05; **P < .005 by 2-tailed Student t test. These results show that despite CCR5 coreceptor usage by primary HIV isolates, HIV does not lead to a homogenous decrease of CCR5-expressing Th lymphocytes during PHI. Rather, it preferentially targets Th lymphocytes expressing CCR5 in the NLHP subpopulation of memory Th lymphocytes. The absence of recovery of this CCR5+ cell population under treatment indicates that, in contrast to other populations of Th lymphocytes, sequestration and redistribution cannot account for its early changes. Rather, CCR5+ NLHP Th cells are lost during PHI, and they are not replaced during the following year despite control of viral replication. Because the vast majority of lamina propria Th lymphocytes express CCR5, depletion of CCR5+ NLHP Th lymphocytes may reflect a preferential compartmentalization of the Th lymphocyte depletion in the gastrointestinal tract at the early stage of HIV infection, as reported in SIV primary infection of macaques.8,9 This would be consistent with the substantial loss of 47+ Th lymphocytes during PHI. The partial recovery of 47+ Th cells under treatment may reflect expansion of these cells or conversion from 47 Th lymphocytes. Such processes do not allow recovery of CCR5+ NLHP Th lymphocytes. The selective and persistent depletion of CCR5+ NLHP Th lymphocytes after PHI is of particular interest in view of recent data demonstrating redistribution of antigen-experienced Th cells from lymphoid organs to nonlymphoid tissues during primary immune responses.18 Moreover, the nonlymphoid compartment seems to promote survival of memory Th lymphocytes. It remains to be determined whether the loss of CCR5+ NLHP memory Th lymphocytes accounts for the loss of anti-HIV-specific memory Th lymphocytes during PHI. NLHP memory Th lymphocytes are richer than LHP in Th1 lymphocytes.19-21 In addition, CCR5 is a marker of the Th1 subpopulation. The depletion of CCR5+ NLHP may thus contribute to the impairment of Th1 responses in HIV-infected patients.22 Our data also suggest the need to reconsider the relevance of markers currently used to evaluate immune reconstitution in HIV-infected patients. It may be valuable to include analysis of CCR5 expression on particular subpopulations of Th lymphocytes, such as NLHP memory Th lymphocytes.     Acknowledgments R.K. and A.R. contributed equally to this work. Ghislaine Lubart and Anne-Marie Delavalle are acknowledged for their technical assistance. The Primoferon A Study Group is composed of the authors and Catherine Lemonnier (Schering-Plough France); F. Duchenne and B. Hoen (Centre Hospitalier-Universitaire [CHU] Besançon); P. Goubin, R. Verdon, and C. Bazin (CHU Caen); J. L. Touraine and J. M. Livrozet (Hôpital E Herriot, Lyon); V. Baillat and J. Reynes (CHU Montpellier); F. Grihon (Hôpital de Noyon); F. Cartier and F. Souala (CHU Rennes); D. Sissoko and Y. Mouton (CHU Tourcoing); C. Yung, P. Lesprita, and Y. Levy (CHU Henri Mondor, Créteil); J. M. Molina, S. Balkan, D. Ponscarme, and C. Lascoux-Combe (CHU Saint-Louis, Paris); C. Rouzioux and M. Burgard (CHU Necker-Enfants-Malades, Paris); C. Pignon (Hôpital A Béclère, Clamart); and G. Chene (Bordeaux), all in France; and M. Laughlin, Schering-Plough Research Institute, Kenilworth, NJ.     Footnotes Submitted May 7, 2001; accepted July 16, 2001. Supported by the Agence Nationale de Recherches sur le SIDA (ANRS). The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734. Reprints: Dominique Emilie, Institut Paris-Sud sur les Cytokines, 32, rue des Carnets, 92 140 Clamart, France; e-mail: emilie{at}ipsc.u-psud.fr.     References Top Abstract Introduction Study design Results and discussion References 1. Zaunders JJ, Kaufmann GR, Cunningham PH, et al. Increased turnover of CCR5+ and redistribution of CCR5 CD4 T lymphocytes during primary human immunodeficiency virus type 1 infection. J Infect Dis. 2001;183:736-743[CrossRef][Medline] [Order article via Infotrieve]. 2. Schacker T, Little S, Connick E, et al. Productive infection of T cells in lymphoid tissues during primary and early human immunodeficiency virus infection. J Infect Dis. 2001;183:555-562[CrossRef][Medline] [Order article via Infotrieve]. 3. Fleury S, Rizzardi GP, Chapuis A, et al. Long-term kinetics of T cell production in HIV-infected subjects treated with highly active antiretroviral therapy. Proc Natl Acad Sci U S A. 2000;97:5393-5398[Abstract/Free Full Text]. 4. Zou W, Foussat A, Houhou S, et al. Acute upregulation of CCR5 expression by CD4+ T lymphocytes in HIV-infected patients treated with interleukin-2. ANRS 048 IL-2 Study Group. AIDS. 1999;13:455-463[CrossRef][Medline] [Order article via Infotrieve]. 5. Butcher EC, Picker LJ. Lymphocyte homing and homeostasis. Science. 1996;272:60-66[Abstract]. 6. Anton PA, Elliott J, Poles MA, et al. Enhanced levels of functional HIV-1 co-receptors on human mucosal T cells demonstrated using intestinal tissue. AIDS. 2000;14:1761-1765[CrossRef][Medline] [Order article via Infotrieve]. 7. Vajdy M, Veazey RS, Knight HK, Lackner AA, Neutra MR. Differential effects of simian immunodeficiency virus infection on immune inductive and effector sites in the rectal mucosa of rhesus macaques. Am J Pathol. 2000;157:485-495[Abstract/Free Full Text]. 8. Veazey RS, Tham IC, Mansfield KG, et al. Identifying the target cell in primary simian immunodeficiency virus (SIV) infection: highly activated memory CD4+ T cells are rapidly eliminated in early SIV infection in vivo. J Virol. 2000;74:57-64[Abstract/Free Full Text]. 9. Veazey RS, DeMaria M, Chalifoux LV, et al. Gastrointestinal tract as a major site of CD4+ T cell depletion and viral replication in SIV infection. Science. 1998;280:427-431[Abstract/Free Full Text]. 10. Smit-McBride Z, Mattapallil JJ, McChesney M, Ferrick D, Dandekar S. Gastrointestinal T lymphocytes retain high potential for cytokine responses but have severe CD4+ T-cell depletion at all stages of simian immunodeficiency virus infection compared to peripheral lymphocytes. J Virol. 1998;72:6646-6656[Abstract/Free Full Text]. 11. Emilie D, Burgard M, Lascoux-Combe C, et al. Early control of replication in primary HIV-1 infection treated with antiretroviral drugs and pegylated interferon alpha. Results from the PRIMOFERON A (ANRS 086) Study. AIDS. 2001;15:1435-1437[CrossRef][Medline] [Order article via Infotrieve]. 12. Carcelain G, Blanc C, Leibowitch J, et al. T cell changes after combined nucleoside analogue therapy in HIV primary infection. AIDS. 1999;13:1077-1081[CrossRef][Medline] [Order article via Infotrieve]. 13. Kaufman GR, Zaunders JJ, Cunningham P. Rapid restoration of CD4 T cell subsets in subjects receiving antiretroviral therapy during primary HIV-1 infection. AIDS. 2000;14:2643-2651[CrossRef][Medline] [Order article via Infotrieve]. 14. Zaunders JJ, Cunningham PH, Kelleher AD. Potent antiretroviral therapy of primary human immunodeficiency virus type 1 (HIV-1) infection: partial normalization of T lymphocyte subsets and limited reduction of HIV-1 DNA despite clearance of plasma viremia. J Infect Dis. 1999;180:320-329[CrossRef][Medline] [Order article via Infotrieve]. 15. Gray CM, Schapiro JM, Winters MA, Merigan TC. Changes in CD4+ and CD8+ T cell subsets in response to highly active antiretroviral therapy in HIV type 1-infected patients with prior protease inhibitor experience. AIDS Res Hum Retroviruses. 1998;14:561-569[Medline] [Order article via Infotrieve]. 16. Hengel RL, Jones BM, Kennedy MS, Hubbard MR, McDougal JS. Markers of lymphocyte homing distinguish CD4 T cell subsets that turn over in response to HIV-1 infection in humans. J Immunol. 1999;163:3539-3548[Abstract/Free Full Text]. 17. Schacker T, Little S, Connick E, et al. Productive infection of T cells in lymphoid tissues during primary and early human immunodeficiency virus infection. J Infect Dis. 2001;183:555-562. 18. Reinhardt RL, Khoruts A, Merica R, Zell T, Jenkins MK. Visualizing the generation of memory CD4 T cells in the whole body. Nature. 2001;410:101-105[CrossRef][Medline] [Order article via Infotrieve]. 19. Sallusto F, Mackay CR, Lanzavecchia A. The role of chemokine receptors in primary, effector, and memory immune responses. Annu Rev Immunol. 2000;18:593-620[CrossRef][Medline] [Order article via Infotrieve]. 20. Iezzi G, Scheidegger D, Lanzavecchia A. Migration and function of antigen-primed nonpolarized T lymphocytes in vivo. J Exp Med. 2001;93:987-994. 21. Hladik F, Lentz G, Delpit E, McElroy A, McElrath MJ. Coexpression of CCR5 and IL-2 in human genital but not blood T cells: implications for the ontogeny of the CCR5+ Th1 phenotype. J Immunol. 1999;163:2306-2313[Abstract/Free Full Text]. 22. Clerici M, Lucey DR, Berzofsky JA, et al. Restoration of HIV-specific cell-mediated immune responses by interleukin-12 in vitro. Science. 1993;262:1721-1724[Abstract/Free Full Text]. © 2001 by The American Society of Hematology.   CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this? This article has been cited by other articles: P. U. Cameron, A. J. Handley, D. C. Baylis, A. E. Solomon, N. Bernard, D. F. J. Purcell, and S. R. Lewin Preferential Infection of Dendritic Cells during Human Immunodeficiency Virus Type 1 Infection of Blood Leukocytes J. Virol., March 1, 2007; 81(5): 2297 - 2306. [Abstract] [Full Text] [PDF] R. Soares, R. Foxall, A. Albuquerque, C. Cortesao, M. Garcia, R. M. M. Victorino, and A. E. Sousa Increased Frequency of Circulating CCR5+ CD4+ T Cells in Human Immunodeficiency Virus Type 2 Infection J. Virol., December 15, 2006; 80(24): 12425 - 12429. [Abstract] [Full Text] [PDF] J. J. Zaunders, S. Ip, M. L. Munier, D. E. Kaufmann, K. Suzuki, C. Brereton, S. C. Sasson, N. Seddiki, K. Koelsch, A. Landay, et al. 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