T-tropic HIV-1 Strains Research Articles

CD40-Mediated induction of CD4 and CXCR4 on B lymphocytes correlates with restricted susceptibility to human immunodeficiency virus type 1 infection: potential role of B lymphocytes as a viral reservoir.

Human immunodeficiency virus type 1 (HIV-1) replicates primarily in lymphoid tissues where it has ready access to activated immune competent cells. We used one of the major pathways of immune activation, namely, CD40-CD40L interactions, to study the infectability of B lymphocytes isolated from peripheral blood mononuclear cells. Highly enriched populations of B lymphocytes generated in the presence of interleukin-4 and oligomeric soluble CD40L upregulated costimulatory and activation markers, as well as HIV-1 receptors CD4 and CXCR4, but not CCR5. By using single-round competent luciferase viruses complemented with either amphotropic or HIV-derived envelopes, we found a direct correlation between upregulation of HIV-1 receptors and the susceptibility of the B lymphocytes to infection with dual-tropic and T-tropic strains of HIV-1; in contrast, cells were resistant to M-tropic strains of HIV-1. HIV-1 envelope-mediated infection was completely abolished with either an anti-CD4 monoclonal antibody or a peptide known to directly block CXCR4 usage and partially blocked with stromal cell-derived factor 1, all of which had no effect on the entry of virus pseudotyped with amphotropic envelope. Full virus replication kinetics confirmed that infection depends on CXCR4 usage. Furthermore, productive cycles of virus replication occurred rapidly yet under most conditions, without the appearance of syncytia. Thus, an activated immunological environment may induce the expression of HIV-1 receptors on B lymphocytes, priming them for infection with selective strains of HIV-1 and allowing them to serve as a potential viral reservoir.

Human intestinal lamina propria lymphocytes are naturally permissive to HIV-1 infection.

The presence of HIV-1 in the intestinal mucosa of AIDS patients has been reported and human intestinal lamina propria lymphocytes (LPL) have been proposed as important targets for HIV-1 infection. However, little information is available concerning the permissiveness of human intestinal CD4+ T lymphocytes to HIV-1 infection. Here, we show that human LPL, in contrast to autologous peripheral blood lymphocytes (PBL), are permissive to both X4 T-tropic and R5 M-tropic strains of HIV-1, as well as to clinical isolates, in the absence of exogenous stimuli. Flow cytometry showed that the vast majority of T LPL were CD45RO+ and CD69+, and that CD4+ T LPL highly expressed CC chemokine receptor 5 (CCR5) as compared to PBL, while CX chemokine receptor 4 was equally expressed on LPL and PBL. Exogenous RANTES and macrophage inflammatory protein-1alpha (natural CCR5 ligands) virtually abolished the entry of the R5 M-tropic strain HIV-1 into human LPL. Thus, we infer that human intestinal CD4+ T lymphocytes are naturally susceptible to HIV-1 infection, due to their physiological state of activation and to marked expression of HIV-1 coreceptors, independently of the route of primary (either mucosal or parental) infection and the shifts of the virus phenotype occurring during the course of AIDS.

Fusion of monocytes and macrophages with HIV-1 correlates with biochemical properties of CXCR4 and CCR5.

Human macrophages can be infected more efficiently by M-tropic than by T-tropic HIV-1 strains, despite surface expression of both CXCR4 and CCR5 co-receptors. Western blot analyses of total cell extracts and surface proteins from multiple sets of monocytes and macrophages demonstrated substantial differences between CXCR4 molecules. CXCR4 was mainly a monomer in monocytes, but was mainly a species of higher molecular weight (90 kDa) on the surface of macrophages. CCR5 was monomeric in both cell types. A constitutive association between CD4 and the co-receptors was seen in monocytes and macrophages. However, CD4 co-precipitated with CCR5 and CXCR4 monomers, but not with the high-molecular-weight forms of CXCR4, indicating that the high-molecular-weight CXCR4 species in macrophages are not available for association with CD4, which may contribute to the inefficient entry of T-tropic strains into mature macrophages.

A Low-Molecular-Weight Inhibitor against the Chemokine Receptor CXCR4: A Strong Anti-HIV Peptide T140

T22 ([Tyr5,12, Lys7]-polyphemusin II) is an 18-residue peptide amide, which has strong anti-HIV activity. T22 inhibits the T cell line-tropic (T-tropic) HIV-1 infection through its specific binding to a chemokine receptor CXCR4, which serves as a coreceptor for the entry of T-tropic HIV-1 strains. Herein, we report our finding of novel 14-residue CXCR4 inhibitors, T134 and T140, on the basis of the T22 structure. In the assays we examined, T140 showed the highest inhibitory activity against HIV-1 entry and the strongest inhibitory effect on the binding of an anti-CXCR4 monoclonal antibody (12G5) to CXCR4 among all the CXCR4 inhibitors that have been reported up to now.

IL-4 and a glucocorticoid up-regulate CXCR4 expression on human CD4+ T lymphocytes and enhance HIV-1 replication.

CXCR4 is a key co-receptor required for the infection of T-tropic HIV-1 strain of CD4+ T lymphocytes. The regulation of this chemokine receptor was therefore studied. Th2 polarized cells expressed more CXCR4 than Th1 cells. Among a panel of cytokines and stimulants, a Th2 type cytokine interleukin-4 (IL-4) selectively up-regulated the mRNA level as well as surface protein expression of CXCR4 within 16 h. In addition, CXCR4 was also up-regulated by a glucocorticoid, dexamethasone. These treated cells became more responsive in transendothelial migration assays to the specific CXCR4 ligand, SDF-1alpha. Furthermore, up-regulation of CXCR4 was also associated with the enhancement of HIV replication in human CD4+ T lymphocytes. This study indicates the enhanced T-tropic HIV-1 infection to CD4+ T lymphocytes through up-regulation of CXCR4 by several immunomodulating agents, IL-4, and a glucocorticoid. These findings may explain the shift to T-tropic HIV-1 dominance during AIDS progression when Th2 comes to predominate.

Enhanced HIV expression during Th2-oriented responses explained by the opposite regulatory effect of IL-4 and IFN-gamma of fusin/CXCR4.

The human alpha-chemokine receptor fusin/CXCR4 is an important cofactor for entry of T lymphocyte-tropic HIV-1 strains. We investigated the possible regulatory role of T cell cytokine patterns on CXCR4 as well as HIV expression by using in vitro models of both secondary and primary immune responses. Antigen-specific memory CD4+ T cells infected with a T-tropic HIV-1 strain showed significantly higher CXCR4 and HIV-1 expression in Th0/2-oriented responses in comparison with Th1-oriented responses. Similarly, in naive CD4+ T cells activated in the presence of IL-4 or IL-12 and infected with the same T-tropic strain, IL-4 up-regulated whereas IL-12 down-regulated both CXCR4 and HIV-1 expression. The down-regulatory effect of IL-12 on CXCR4 expression was found to be dependent on its capacity to induce IFN-gamma production. These observations can account for the higher risk of progression in HIV-1-infected individuals undergoing Th0/2-oriented immune responses.

CC-chemokines enhance the replication of T-tropic strains of HIV-1 in CD4(+) T cells: role of signal transduction.

This study demonstrates that several CC-chemokines, including those that inhibit entry and replication of macrophage-tropic strains of HIV, increase the replication of T cell (T)-tropic strains in CD4(+) T cells. Enhancement of T-tropic HIV replication is observed at early stages of replication, requires signaling through inhibitory guanine nucleotide-binding regulatory (Gi) proteins, and is associated with increased cell surface colocalization of CD4 and the T-tropic HIV coreceptor CXCR4. These findings may further our understanding of the factors that influence the replication and spread of T-tropic strains of HIV in vivo and suggest that the use of cell signaling CC-chemokines as therapeutic agents for the purpose of limiting HIV replication in vivo should be approached with caution.

Cutting Edge: Enhanced Anti-HIV-1 Activity and Altered Chemotactic Potency of NH2-Terminally Processed Macrophage-Derived Chemokine (MDC) Imply an Additional MDC Receptor

AbstractPosttranslational processing of chemokines increases (IL-8) or decreases (monocyte chemotactic protein-1) their chemotactic potency. Macrophage-derived chemokine (MDC) attracts monocytes, dendritic cells, activated lymphocytes, and NK cells and has reportedly anti-HIV-1 activity. Here we report that truncation of MDC by deletion of two NH2-terminal residues resulted in impaired binding to CC chemokine receptor (CCR)4, the only identified MDC receptor sofar. Truncated MDC(3-69) failed to desensitize calcium mobilization by MDC(1-69) or thymus- and activation-regulated chemokine (TARC), another CCR4 ligand. MDC(3-69) lacked HUT-78 T cell chemotactic activity but retained its capacity to attract monocytes and to desensitize chemotaxis. Compared with MDC(1-69), MDC(3-69) had weak but enhanced antiviral activity against M- and T-tropic HIV-1 strains. Furthermore, both MDC forms failed to signal through the orphan receptors Bonzo/STRL33 and BOB/GPR15 and to desensitize RANTES and stromal cell-derived factor (SDF)-1 responses in CCR5-transfected and CXC chemokine receptor (CXCR)4-transfected cells, respectively. These findings suggest that MDC recognizes another, yet unidentified, receptor. We conclude that minimal NH2-terminal truncation of MDC differentially affects its various immunologic functions.

CXCR4 and CCR5 on Human Thymocytes: Biological Function and Role in HIV-1 Infection

Thymocyte infection with HIV-1 is associated with thymic involution and impaired thymopoiesis, particularly in pediatric patients. To define mechanisms of thymocyte infection, we examined human thymocytes for expression and function of CXCR4 and CCR5, the major cell entry coreceptors for T cell line-tropic (T-tropic) and macrophage-tropic (M-tropic) strains of HIV-1, respectively. CXCR4 was detected on the surface of all thymocytes. CXCR4 expression on mature, high level TCR thymocytes was similar to that on peripheral blood T cells, but was much lower than that on immature thymocytes, including CD34+ thymic progenitors. Consistent with this, stroma-derived factor-1 (SDF-1) induced calcium flux primarily in immature thymocytes, with CD34+ progenitors giving the strongest response. In addition, SDF-1 mRNA was detected in thymic-derived stromal cells, and SDF-1 induced chemotaxis of thymocytes, suggesting that CXCR4 may play a role in thymocyte migration. Infection of immature thymocytes by the T-tropic HIV-1 strain LAI was 10-fold more efficient than that in mature thymocytes, consistent with their relative CXCR4 surface expression. Anti-CXCR4 antiserum or SDF-1 blocked fusion of thymocytes with cells expressing the LAI envelope. In contrast to CXCR4, CCR5 was detected at low levels on thymocytes, and CCR5 agonists did not induce calcium flux or chemotaxis in thymocytes. However, CD4+ mature thymocytes were productively infected with the CCR5-tropic strain Ba-L, and this infection was specifically inhibited with the CCR5 agonist, macrophage inflammatory protein-1beta. Our data provide strong evidence that CXCR4 and CCR5 function as coreceptors for HIV-1 infection of human thymocytes.

Processing by CD26/dipeptidyl-peptidase IV reduces the chemotactic and anti-HIV-1 activity of stromal-cell-derived factor-1α

The chemokine stromal-cell-derived factor-1α (SDF-1α) chemoattracts lymphocytes and CD34 + haematopoietic progenitors and is the ligand for CXCR4 (CXC chemokine receptor 4), the main co-receptor for T-tropic HIV-1 strains. SDF-1α was NH 2-terminally cleaved to SDF-1α(3-68) by dipeptidyl-peptidase IV (CD26/DPP IV), which is present in blood in soluble and membrane-bound form. SDF-1α(3-68) lost both lymphocyte chemotactic and CXCR4-signaling properties. However, SDF-1α(3-68) still desensitized the SDF-1α(1-68)-induced Ca 2+ response. In contrast to CD26/DPP IV-processed RANTES(3-68), SDF-1α(3-68) had diminished potency to inhibit HIV-1 infection. Thus, CD26/DPP IV impairs the inflammatory and haematopoietic potency of chemokines but plays a dual role in AIDS.

Crystal structure of chemically synthesized [N33A] stromal cell-derived factor 1alpha, a potent ligand for the HIV-1 "fusin" coreceptor.

Stromal cell-derived factor-1alpha (SDF-1alpha ) is a member of the chemokine superfamily and functions as a growth factor and chemoattractant through activation of CXCR4/LESTR/Fusin, a G protein-coupled receptor. This receptor also functions as a coreceptor for T-tropic syncytium-inducing strains of HIV-1. SDF-1alpha antagonizes infectivity of these strains by competing with gp120 for binding to the receptor. The crystal structure of a variant SDF-1alpha ([N33A]SDF-1alpha ) prepared by total chemical synthesis has been refined to 2.2-A resolution. Although SDF-1alpha adopts a typical chemokine beta-beta-beta-alpha topology, the packing of the alpha-helix against the beta-sheet is strikingly different. Comparison of SDF-1alpha with other chemokine structures confirms the hypothesis that SDF-1alpha may be either an ancestral protein from which all other chemokines evolved or the chemokine that is the least divergent from a primordial chemokine. The structure of SDF-1alpha reveals a positively charged surface ideal for binding to the negatively charged extracellular loops of the CXCR4 HIV-1 coreceptor. This ionic complementarity is likely to promote the interaction of the mobile N-terminal segment of SDF-1alpha with interhelical sites of the receptor, resulting in a biological response.

Productive infection of neonatal CD8+ T lymphocytes by HIV-1.

CD8+ T lymphocytes confer significant but ultimately insufficient protection against HIV infection. Here we report that activated neonatal CD8+ T cells can be productively infected in vitro by macrophage-tropic (M-tropic) HIV-1 isolates, which are responsible for disease transmission, whereas they are resistant to T cell-tropic (T-tropic) HIV strains. Physiological activation of CD8-alpha/beta+ CD4- T cell receptor-alpha/beta+ neonatal T cells, including activation by allogeneic dendritic cells, induces the accumulation of CD4 messenger RNA and the expression of CD4 Ag on the cell surface. The large majority of anti-CD3/B7.1-activated cord blood CD8+ T cells coexpress CD4, the primary HIV receptor, as well as CCR5 and CXCR4, the coreceptors used by M- and T-tropic HIV-1 strains, respectively, to enter target cells. These findings are relevant to the rapid progression of neonatal HIV infection. Infection of primary HIV-specific CD8+ T cells may compromise their survival and thus significantly contribute to the failure of the immune system to control the infection. Furthermore, these results indicate a previously unsuspected level of plasticity in the neonatal immune system in the regulation of CD4 expression by costimulation.

Differentiation of Promonocytic U937 Subclones into Macrophagelike Phenotypes Regulates a Cellular Factor(s) Which Modulates Fusion/Entry of Macrophagetropic Human Immunodeficiency Virus Type 1

Monocytes/macrophages (M/M) and CD4+ T cells are two important targets of human immunodeficiency virus (HIV) infection. Different strains of HIV-1 vary markedly in their abilities to infect cells belonging to the M/M lineage. Macrophagetropic (M-tropic) HIV-1 strains replicate well in primary lymphocytes as well as in primary macrophages; however, they generally infect T-cell lines poorly, if at all. Although promonocytic cell lines such as U937 have been used as in vitro models of HIV-1 infection of M/M, these cell lines are susceptible to certain T-cell-tropic (T-tropic) HIV-1 strains but are resistant to M-tropic HIV-1. In this study, we demonstrate that (i) certain U937 clones ("plus" clones), which are susceptible only to T-tropic HIV-1, become highly susceptible to M-tropic HIV-1 upon differentiation with retinoic acid (RA); (ii) other U937 clones ("minus" clones), which are resistant to both T- and M-tropic HIV-1, remain resistant to both viruses; and (iii) RA treatment induces expression of CCR5, a fusion/entry cofactor for M-tropic HIV-1 in both types of U937 clones, and yet enhances the fusogenicity of the plus clones, but not the minus clones, with M-tropic Env's. These results indicate that the major restriction of M-tropic HIV-1 infection in promonocytic cells occurs at the fusion/entry level, that differentiation into macrophage-like phenotypes renders some of these cells highly susceptible to infection with M-tropic HIV-1, and that CD4 and CCR5 may not be the only determinants of fusion/entry of M-tropic HIV-1 in these cells.

Embryonic expression of the mRNA for the rat homologue of the fusin/CXCR-4 HIV-1 co-receptor

We have previously cloned a human receptor recently shown to be a cofactor for entry of T-tropic HIV-1 strains into CD4+ cells, now named fusin. Stromal derived factor-1 (SDF-1) is an endogenous ligand for fusin, also called CXCR-4. Here we show the distribution of fusin/CXCR-4 mRNA during ontogeny in the rat. The onset of mRNA expression is around embryonic day 9 and the mRNA expression is high in the thymus as well as proliferative areas of the brain during development. Our results suggest: (1) that fusin/CXCR-4 might have a dual role in both brain development and the immune system; (2) that SDF-1 has a role in brain development or that additional physiological ligands exist for this receptor; (3) co-expression of CD4 and fusin/CXCR-4 may make fetuses susceptible to HIV infection during development.