Human Tonsillar Tissue Research Articles

Replicative fitness and pathogenicity of primate lentiviruses in lymphoid tissue, primary human and chimpanzee cells: relation to possible jumps to humans

Simian immunodeficiency viruses (SIV) have been jumping between non-human primates in West/Central Africa for thousands of years and yet, the HIV-1 epidemic only originated from a primate lentivirus over 100 years ago. This study examined the replicative fitness, transmission, restriction, and cytopathogenicity of 22 primate lentiviruses in primary human lymphoid tissue and both primary human and chimpanzee peripheral blood mononuclear cells. Pairwise competitions revealed that SIV from chimpanzees (cpz) had the highest replicative fitness in human or chimpanzee peripheral blood mononuclear cells, even higher fitness than HIV-1 group M strains responsible for worldwide epidemic. The SIV strains belonging to the "HIV-2 lineage" (including SIVsmm, SIVmac, SIVagm) had the lowest replicative fitness. SIVcpz strains were less inhibited by human restriction factors than the "HIV-2 lineage" strains. SIVcpz efficiently replicated in human tonsillar tissue but did not deplete CD4+ T-cells, consistent with the slow or nonpathogenic disease observed in most chimpanzees. In contrast, HIV-1 isolates and SIV of the HIV-2 lineage were pathogenic to the human tonsillar tissue, almost independent of the level of virus replication. Of all primate lentiviruses, SIV from chimpanzees appears most capable of infecting and replicating in humans, establishing HIV-1. SIV from other Old World monkeys, e.g. the progenitor of HIV-2, replicate slowly in humans due in part to restriction factors. Nonetheless, many of these SIV strains were more pathogenic than SIVcpz. Either SIVcpz evolved into a more pathogenic virus while in humans or a rare SIVcpz, possibly extinct in chimpanzees, was pathogenic immediately following the jump into human. Support for this study to E.J.A. was provided by the NIH/NIAID R01 AI49170 and CIHR project grant 385787. Infrastructure support was provided by the NIH CFAR AI36219 and Canadian CFI/Ontario ORF 36287. Efforts of J.A.B. and N.J.H. was provided by NIH AI099473 and for D.H.C., by VA and NIH AI AI080313.

Differences in the metabolomic profile of the human palatine tonsil between pediatrics and adults.

Palatine tonsils (PT) are B cell-predominant lymphoid organs that provide primary immune responses to airborne and dietary pathogens. Numerous histopathological and immunological studies have been conducted on PT, yet no investigations have been conducted on its metabolic profile. We performed high-resolution magic angle spinning nuclear magnetic resonance spectroscopy-based metabolic profiling in 35 pediatric and 28 adult human palatine tonsillar tissue samples. A total of 36 metabolites were identified, and the levels of 10 metabolites were significantly different depending on age. Among them, partial correlation analysis shows that glucose levels increased with age, whereas glycine, phosphocholine, phosphoethanolamine, and ascorbate levels decreased with age. We confirmed the decrease in immunometabolic activity in adults through metabolomic analysis, which had been anticipated from previous histological and immunological studies on the PT. These results improve our understanding of metabolic changes in the PT with aging and serve as a basis for future tonsil-related metabolomic studies.

Distinct stage-specific transcriptional states of B cells derived from human tonsillar tissue.

B cells within secondary lymphoid tissues encompass a diversity of activation states and multiple maturation processes that reflect antigen recognition and transition through the germinal center (GC) reaction, in which mature B cells differentiate into memory and antibody-secreting cells (ASCs). Here, utilizing single-cell RNA-seq, we identify a range of distinct activation and maturation states of tonsil-derived B cells. In particular, we identify what we believe is a previously uncharacterized CCL4/CCL3 chemokine-expressing B cell population with an expression pattern consistent with B cell receptor/CD40 activation. Furthermore, we present a computational method that leverages regulatory network inference and pseudotemporal modeling to identify upstream transcription factor modulation along a GC-to-ASC axis of transcriptional maturation. Our data set provides valuable insight into diverse B cell functional profiles and will be a useful resource for further studies into the B cell immune compartment.

The Progestin Medroxyprogesterone Acetate Affects HIV-1 Production in Human Lymphoid Tissue Explants in a Dose-Dependent and Glucocorticoid-like Fashion.

The association between the use of the injectable contraceptive depot medroxyprogesterone acetate and HIV-1 susceptibility has been addressed mainly in respect to the changes occurring in the female genital mucosa and blood. However, one of the main sites of HIV-1 pathogenesis is lymphoid organs. To investigate the immunoregulatory effect of medroxyprogesterone acetate (MPA) at this site, human tonsillar tissue explants were infected ex vivo with either a CCR5 (BaL) or CXCR4 (LAI) HIV-1 variant and the release of p24gag and cytokines was measured in culture supernatant. The response to MPA was compared with that elicited by treatment with progesterone (P4) and dexamethasone (DEX), which selectively binds the glucocorticoid receptor, in donor-matched explant cultures. MPA treatment reduced the replication of both tested HIV-1 strains as well as the production of the mediators of inflammation IL-1β, IL-17A and CCL5, but not CCL20, in a similar way to DEX, whereas P4 had no effect on HIV-1 replication. The magnitude of both MPA and DEX-mediated responses was proportional to the length of exposure and/or administered dose. Blockage of the progesterone and glucocorticoid receptors with mifepristone abolished all observed changes in HIV-1 and cytokine production, and was associated with increased IL-22 levels in HIV-infected explants. Our data indicate that elevated doses of MPA may affect the immune responses in lymphoid tissue in a glucocorticoid-like fashion with an immediate impact on local HIV-1 replication.

SARS-CoV-2-Reactive Mucosal B Cells in the Upper Respiratory Tract of Uninfected Individuals.

SARS-CoV-2 is a respiratory pathogen that can cause severe disease in at-risk populations but results in asymptomatic infections or a mild course of disease in the majority of cases. We report the identification of SARS-CoV-2-reactive B cells in human tonsillar tissue obtained from children who were negative for coronavirus disease 2019 prior to the pandemic and the generation of mAbs recognizing the SARS-CoV-2 Spike protein from these B cells. These Abs showed reduced binding to Spike proteins of SARS-CoV-2 variants and did not recognize Spike proteins of endemic coronaviruses, but subsets reacted with commensal microbiota and exhibited SARS-CoV-2-neutralizing potential. Our study demonstrates pre-existing SARS-CoV-2-reactive Abs in various B cell populations in the upper respiratory tract lymphoid tissue that may lead to the rapid engagement of the pathogen and contribute to prevent manifestations of symptomatic or severe disease.

A global phylogenetic analysis of Japanese tonsil-derived Epstein-Barr virus strains using viral whole-genome cloning and long-read sequencing.

Epstein-Barr virus (EBV) establishes lifelong latent infection in the majority of healthy individuals, while it is a causative agent for various diseases, including some malignancies. Recent high-throughput sequencing results indicate that there are substantial levels of viral genome heterogeneity among different EBV strains. However, the extent of EBV strain variation among asymptomatically infected individuals remains elusive. Here, we present a streamlined experimental strategy to clone and sequence EBV genomes derived from human tonsillar tissues, which are the reservoirs of asymptomatic EBV infection. Complete EBV genome sequences, including those of repetitive regions, were determined for seven tonsil-derived EBV strains. Phylogenetic analyses based on the whole viral genome sequences of worldwide non-tumour-derived EBV strains revealed that Asian EBV strains could be divided into several distinct subgroups. EBV strains derived from nasopharyngeal carcinoma-endemic areas constitute different subgroups from a subgroup of EBV strains from non-endemic areas, including Japan. The results could be consistent with biased regional distribution of EBV-associated diseases depending on the different EBV strains colonizing different regions in Asian countries.

Alternative splicing of the TNFSF13B (BAFF) pre-mRNA and expression of the BAFFX1 isoform in human immune cells.

Human B cell activating factor (TNFSF13B, BAFF) is a tumor necrosis factor superfamily member. Binding its unique receptor (TNFRSF13C, BAFF-R) mediates gene expression and cell survival in B cells via activation of NFκB pathway. Furthermore, there is data indicating a role in T cell function. A functionally inhibitory isoform (ΔBAFF) resulting from the deletion of exon 3 in the TNFSF13B pre-RNA has already been reported. However, data on the complexity of post-transcriptional regulation is scarce. Here, we report molecular cloning of nine TNFSF13B transcript variants resulting from alternative splicing of the TNFSF13B pre-mRNA including BAFFX1. This variant is characterized by a partial retention of intron 3 of the TNFSF13B gene causing the appearance of a premature stop codon. We demonstrate the expression of the corresponding BAFFX1 protein in Jurkat T cells, in ex vivo human immune cells and in human tonsillar tissue. Thereby we contribute to the understanding of TNFSF13B gene regulation and reveal that BAFF is regulated through a post-transcriptional mechanism to a greater extent than reported to date.

Extracellular vesicles from symbiotic vaginal lactobacilli inhibit HIV-1 infection of human tissues

The vaginal microbiota, dominated by Lactobacillus spp., plays a key role in preventing HIV-1 transmission. Here, we investigate whether the anti-HIV effect of lactobacilli is mediated by extracellular vesicles (EVs) released by these bacteria. Human cervico-vaginal and tonsillar tissues ex vivo, and cell lines were infected with HIV-1 and treated with EVs released by lactobacilli isolated from vaginas of healthy women. EVs released by L. crispatus BC3 and L. gasseri BC12 protect tissues ex vivo and isolated cells from HIV-1 infection. This protection is associated with a decrease of viral attachment to target cells and viral entry due to diminished exposure of Env that mediates virus-cell interactions. Inhibition of HIV-1 infection is associated with the presence in EVs of several proteins and metabolites. Our findings demonstrate that the protective effect of Lactobacillus against HIV-1 is, in part, mediated by EVs released by these symbiotic bacteria. If confirmed in vivo, this finding may lead to new strategies to prevent male-to-female sexual HIV-1 transmission.

Intracellular BH3 Profiling Reveals Shifts in Antiapoptotic Dependency in Human B Cell Maturation and Mitogen-Stimulated Proliferation.

Apoptosis is critical to B cell maturation, but studies of apoptotic regulation in primary human B cells is lacking. In this study, we sought to better understand the mechanisms of apoptotic regulation in normal and activated B cells. Using intracellular BH3 profiling, we defined the Bcl2 dependency of B cell subsets from human peripheral blood and tonsillar lymphoid tissue as well as mitogen-activated B cells. We found that naive and memory B cells were BCL-2-dependent, whereas germinal center B cells were MCL-1-dependent and plasma cells were BCL-XL-dependent. B cells stimulated to proliferate ex vivo by CpG or CD40L/IL-4 became more dependent on MCL-1 and BCL-XL As B cell lymphomas often rely on survival mechanisms derived from normal and activated B cells, these findings offer new insight into potential therapeutic strategies for lymphomas.

Virtual Screening of Acyclovir Derivatives as Potential Antiviral Agents: Design, Synthesis, and Biological Evaluation of New Acyclic Nucleoside ProTides.

Following our findings on the anti-human immunodeficiency virus (HIV) activity of acyclovir (ACV) phosphate prodrugs, we herein report the ProTide approach applied to a series of acyclic nucleosides aimed at the identification of novel and selective antiviral, in particular anti-HIV agents. Acyclic nucleoside analogues used in this study were identified through a virtual screening using HIV-reverse transcriptase (RT), adenylate/guanylate kinase, and human DNA polymerase γ. A total of 39 new phosphate prodrugs were synthesized and evaluated against HIV-1 (in vitro and ex vivo human tonsillar tissue system) and human herpes viruses. Several ProTide compounds showed substantial potency against HIV-1 at low micromolar range while the parent nucleosides were not effective. Also, pronounced inhibition of herpesvirus replication was observed. A carboxypeptidase-mediated hydrolysis study was performed for a selection of compounds to assess the formation of putative metabolites and support the biological activity observed.

Therapeutic potential of tonsil-derived mesenchymal stem cells in dextran sulfate sodium-induced experimental murine colitis.

The therapeutic potential of tonsil-derived mesenchymal stem cells (TMSC) prepared from human tonsillar tissue has been studied in animal models for several diseases such as hepatic injury, hypoparathyroidism, diabetes and muscle dystrophy. In this study, we examined the therapeutic effects of TMSC in a dextran sulfate sodium (DSS)-induced colitis model. TMSC were injected in DSS-induced colitis mice via intraperitoneal injection twice (TMSC[x2]) or four times (TMSC[x4]). Control mice were injected with either phosphate-buffered saline or human embryonic kidney 293 cells. Body weight, stool condition and disease activity index (DAI) were examined daily. Colon length, histologic grading, and mRNA expression of pro-inflammatory cytokines, interleukin 1β (IL-1β), IL-6, IL-17 and tumor necrosis factor α, and anti-inflammatory cytokines, IL-10, IL-11 and IL-13, were also measured. Our results showed a significant improvement in survival rates and body weight gain in colitis mice injected with TMSC[x2] or TMSC[x4]. Injection with TMSC also significantly decreased DAI scores throughout the experimental period; at the end of experiment, almost complete reversal of DAI scores to normal was found in colitis mice treated with TMSC[x4]. Colon length was also significantly recovered in colitis mice treated with TMSC[x4]. However, histopathological alterations induced by DSS treatment were not apparently improved by injection with TMSC. Finally, treatment with TMSC[x4] significantly reversed the mRNA levels of IL-1β and IL-6, although expression of all pro-inflammatory cytokines tested was induced in colitis mice. Under our experimental conditions, however, no apparent alterations in the mRNA levels of all the anti-inflammatory cytokines tested were found. In conclusion, our findings demonstrate that multiple injections with TMSC produced a therapeutic effect in a mouse model of DSS-induced colitis.

Mass Cytometry of Follicular Lymphoma Tumors Reveals Intrinsic Heterogeneity in Proteins Including HLA-DR and a Deficit in Nonmalignant Plasmablast and Germinal Center B-Cell Populations.

Follicular lymphoma (FL) is an indolent non-Hodgkin lymphoma that has a risk of transformation to more aggressive lymphoma. Relatively little is known about the nonmalignant B-cell and T-cell subset composition within the tumor microenvironment and whether altered phenotypes are associated with patterns of lymphoma B-cell heterogeneity. Two mass cytometry (CyTOF) panels were designed to immunophenotype B and T cells in FL tumors. Populations of malignant B cells, nonmalignant B cells, and T cells from each FL tumor were identified and their phenotypes compared to B and T cells from healthy human tonsillar tissue. Diversity in cellular phenotype between tumors was greater for the malignant B cells than for nonmalignant B or T cells. The malignant B-cell population bore little phenotypic similarity to any healthy B-cell subset, and unexpectedly clustered closer to naïve B-cell populations than GC B-cell populations. Among the nonmalignant B cells within FL tumors, a significant lack of GC and plasmablast B cells was observed relative to tonsil controls. In contrast, nonmalignant T cells in FL tumors were present at levels similar to their cognate tonsillar T-cell subsets. Mass cytometry revealed that diverse HLA-DR expression on FL cells within individual tumors contributed greatly to tumor heterogeneity. Both malignant and nonmalignant B cells in the tumor bore little phenotypic resemblance to healthy GC B cells despite the presence of T follicular helper cells in the tumor. These findings suggest that ongoing signaling interactions between malignant B cells and intra-tumor T cells shape the tumor microenvironment. © 2016 International Clinical Cytometry Society.

Expression of antimicrobial peptides in recurrent adenotonsillitis

Abstract Background Recurrent acute tonsillitis is one of the most frequent otorhinolaryngology clinic referrals, yet its pathogenesis remains poorly understood. Antimicrobial cationic peptides are components of the innate system. They are generally small, highly positively charged peptides with broad spectrum antimicrobial activity which function as the body’s “natural antibiotics”. Our aim is to investigate the role of antimicrobial cationic peptides in the susceptibility of patients to recurrent acute tonsillitis. Materials and methods The study is done with 100 children who had a history of recurrent adenotonsillitis as subject group and 100 children with adenotonsillar hypertrophy as control group. Tonsillar and adenoid tissues are dissected into parts as deep and surface epithelium and investigated semiquantitatively with immunohistochemistry. Human beta defensin (hBD) 1–3 and cathelecidin (LL-37) levels are compared with microscopically. Results Immunohistochemistry revealed a strong expression of hBD-1, hBD-2 and hBD-3 in tonsillar tissue. Quantification of hBD-1, hBD-2 and hBD-3 expressions are shown more in tonsillar tissue than in adenoids. LL-37 is one of the antimicrobial peptides found in human tonsillar tissue and adenoids, that participates in the innate immune system of these tissues. Statistically, hBD-1, hBD-3 and LL-37 expressions were different in recurrent tonsillitis tissue than control (p<0.05). Moreover hBD-2 expression was different in adenoid tissue than control (p<0.05). Conclusion Antimicrobial peptides have key role in adenotonsillar infections and this defense mechanism increases susceptibility to recurrent infections in upper respiratory tract.

CXCR5+CD8+ T cells are localized in B cell follicles and germinal centers and exhibit regulatory and anti-tumor function

Germinal centers (GC) are specialized anatomic sites where B cells undergo somatic hypermutation, clonal expansion, and affinity-based selection. Stringent regulation of GC reactions is critical for homeostatic B cell development, as well as T-dependent humoral immunity against self and foreign antigens. Follicular helper T cells (Tfh, CXCR5hiPDhiBcl-6+CD4+) that reside in GC serve as specialized B helper T cells and provide survival and selection signals to GC B cells. Despite recent understandings of Tfh function in autoimmunity, immunodeficiency, and B-cell lymphoma, mechanisms that regulate Tfh development and function remain limited. In the current study, we determined the presence of CD45RA-CXCR5+CD8+ T cells in human peripheral blood, tonsillar tissues, and follicular lymphoma (FL) tumor samples by FACS and confocal microscopy. We found that CXCR5+CD8+ T cells were present in high numbers and localized to GCs and T cell zones in the tonsillar tissues and FL, but present in low numbers in the peripheral blood. Surface marker analysis suggested that these cells displayed an activated status. Importantly, FL tumor samples had significantly more CXCR5+CD8+ T cells than tonsillar tissues. Further analysis using intracellular cytokine staining showed that CXCR5+CD8+ T cells produced higher levels of IFN-γ and TNF-α compared to CXCR5-CD8+ subset. Next, we performed T-B cell co-culture experiments to evaluate whether CXCR5+CD8+ T cells could regulate Tfh function in humans. FACS-sorted Tfh cells were co-cultured with naive or memory B cells in the presence or absence of CXCR5+CD8+ T cells. These experiments indicated that CXCR5+CD8+ T cells suppressed Tfh function, as demonstrated by reduced differentiation of naive and/or memory B cells into plasmablasts cells (CD19intCD38+). To test CXCR5+CD8+ T cell function in vivo, we employed the EG7 lymphoma model in TCR-/- mice. Briefly, ovabulmin-primed OT1-specific CXCR5+CD8+ T cells were adoptively transferred into EG7 tumor-bearing TCR-/- mice, and tumor size and survival rate were determined. In summary, our data suggest that CXCR5+CD8+ T cells may have multiple roles, as they not only inhibited Tfh function, but also exhibited strong anti-tumor immunity.

5-Arylaminouracil Derivatives as Potential Dual-Action Agents

Several 5-aminouracil derivatives that have previously been shown to inhibit Mycobacterium tuberculosis growth at concentrations of 5-40 μg/mL are demonstrated to act also as noncompetitive non-nucleoside inhibitors of HIV-1 reverse transcriptase without causing toxicity in vitro (MT-4 cells) and ex vivo (human tonsillar tissue).

Death by Firestorm: Why Bystander CD4 Cells Die During HIV Infection

Although the death of productively HIV-infected T cells is known to occur through apoptosis, the overwhelming demise of quiescent “bystander” CD4 cells, which reside mostly in lymphoid tissues, remains unexplained. The cause of heightened inflammation in the setting of HIV infection is also not well understood. Now, using fresh human tonsillar tissue, researchers in San Francisco have demonstrated that the way lymphoid CD4 cells die depends on the manner in which HIV infects them. They found that CD4 cells must be activated to permit productive HIV infection. HIV can also infect latent, nonpermissive CD4 cells, but the …

IL10 Receptor a Is a Novel Therapeutic Target That Is Epigenetically Disregulated in Low Grade Lymphomas with Plasmacytic Differentiation.

Abstract 2383Waldenstrom's Macroglobulinemia (WM) is the clinical manifestation of lymphoproliferative disorders characterized by a clonal lymphoplasmacytic proliferation, excessive IgM secretion and elevated serum viscosity.Most common B-cell non-Hodgkin lymphomas associated with WM are Lymphoplasmacytic Lymphomas (LPL) and Marginal Zone Lymphomas (MZL), especially those displaying plasmacytic differentiation (MZL-P). In order to elucidate common pathogenetic mechanisms that result in WM across lymphoma types, we profiled the methylome of 6 LPLs, 10 MZL-Ps, as well as normal B-cell subsets purified from human tonsillar tissue –germinal center B-cell controls (GCB, n=3) and plasma cell (PC, n=3) controls, using HELP assay and high-density oligonucleotide microarray from RocheNImblegen that queries DNA methylation level of 50,000 cytosine residues distributed among 14,000 gene promoters.Unsupervised hierarchical clustering approach using Ward’s method and Eucledian distances separated normal GCB and PC from the lymphoma cases and identified bi-directional changes in gene methylation: with aberrant gain or loss of methylation at specific genomic locations. We further utilized a t-test and identified 208 probesets that were differentially methylated between controls and lymphomas at p<0.01 and mean log-ratio difference between 2 groups >1.5 corresponding to 30% methylation difference. Ingenuity pathway analysis revealed CDKN1A and TGFb networks as the most aberrantly methylated in lymphomas.Remarkably, we observed that Interleukin 10 Receptor a (IL10RA), was aberrantly hypomethylated in both lymphoma subtypes, resulting in its aberrant overexpression. IL10RA is a subunit of the IL10 cytokine receptor that is known to be a key factor in terminating the inflammatory responses via signaling through the JAK/STAT pathway resulting in STAT3 activation, which also plays an integral part in GCB differentiation and commitment to the plasma cell lineage. We thus hypothesized that aberrant epigenetic upregulation of IL10RA might promote survival and expansion of lymphoma cells. We predicted that stimulating IL10RA with IL10 ligand may lead to increased cell growth, while the blockade of IL10RA would inhibit cell growth. We selected 2 anti-IL10RA antibodies, which have previously been reported to have receptor blocking properties in vivo in mice. We tested 1ug/ml, 5ug/ml and 10ug/ml of each antibody and 10 ng/ml of the stimulatory IL10 ligand and observed that ligand provided stimulatory effect on the growth rate of a panel of B-cell lymphoma, including LPL cell line, while both anti-IL10RA antibodies had marked growth inhibitory effects. We further determined that growth inhibition resulted from dramatic induction of apoptosis in cells treated with the blocking anti-IL10RA antibodies. Further studies revealed that induction of apoptosis followed specific inhibition of signaling through JAK1/2 and phosphorylation of STAT3Y705 immediately after treatment and inhibition of signaling through MAPK and phosphorylation of STAT3S727 at later treatment time points.In conclusion, we determined that IL10RA is aberrantly methylated and overexpressed in subtypes of low grade lymphomas exhibiting plasmacytic differentiation and manifesting as WM. Hence, strategies targeting this pathway should be explored as potential therapy for WM (irrespective of the underlying type of lymphoma). Disclosures:No relevant conflicts of interest to declare.

Susceptibility of human lymphoid tissue cultured ex vivo to xenotropic murine leukemia virus-related virus (XMRV) infection.

Background:Xenotropic murine leukemia virus-related virus (XMRV) was generated after a recombination event between two endogenous murine leukemia viruses during the production of a prostate cancer cell line. Although the associations of the XMRV infection with human diseases appear unlikely, the XMRV is a retrovirus of undefined pathogenic potential, able to replicate in human cells in vitro. Since recent studies using animal models for infection have yielded conflicting results, we set out an ex vivo model for XMRV infection of human tonsillar tissue to determine whether XMRV produced by 22Rv1 cells is able to replicate in human lymphoid organs. Tonsil blocks were infected and infection kinetics and its pathogenic effects were monitoredResults:XMRV, though restricted by APOBEC, enters and integrates into the tissue cells. The infection did not result in changes of T or B-cells, immune activation, nor inflammatory chemokines. Infectious viruses could be recovered from supernatants of infected tonsils by reinfecting DERSE XMRV indicator cell line, although these supernatants could not establish a new infection in fresh tonsil culture, indicating that in our model, the viral replication is controlled by innate antiviral restriction factors.Conclusions:Overall, the replication-competent retrovirus XMRV, present in a high number of laboratories, is able to infect human lymphoid tissue and produce infectious viruses, even though they were unable to establish a new infection in fresh tonsillar tissue. Hereby, laboratories working with cell lines producing XMRV should have knowledge and understanding of the potential biological biohazardous risks of this virus.

Acquisition of Host-Derived CD40L by HIV-1In Vivoand Its Functional Consequences in the B-Cell Compartment

Aberrant activation of the B-cell compartment and hypergammaglobulinemia were among the first recognized characteristics of HIV-1-infected patients in the early 1980s. It has been demonstrated previously that HIV-1 particles acquire the costimulatory molecule CD40L when budding from activated CD4(+) T cells. In this paper, we confirmed first that CD40L-bearing virions are detected in the plasma from untreated HIV-1-infected individuals. To define the biological functions of virus-associated CD40L and fully characterize its influence on the activation state of B cells, we conducted a large-scale gene expression analysis using microarray technology on B cells isolated from human tonsillar tissue. Comparative analyses of gene expression profiles revealed that CD40L-bearing virions induce a highly similar response to the one observed in samples treated with a CD40 agonist, indicating that virions bearing CD40L can efficiently activate B cells. Among modulated genes, many cytokines/chemokines (CCL17, CCL22), surface molecules (CD23, CD80, ICAM-1), members of the TNF superfamily (FAS, A20, TNIP1, CD40, lymphotoxin alpha, lymphotoxin beta), transcription factors and associated proteins (NFKB1, NFKBIA, NFKBIE), second messengers involved in CD40 signaling (TRAF1, TRAF3, MAP2K1, phosphatidylinositol 3-kinase), and the activation-induced cytidine deaminase (AID) were identified. Moreover, we show that soluble factors induced upon the exposure of B cells to CD40L-bearing virions can exert chemoattractant properties toward CD4(+) T cells. We thus propose that a positive feedback loop involving CD40L-bearing HIV-1 particles issued from CD4(+) T cells productively infected with HIV-1 play a role in the virus-induced dysfunction of humoral immunity by chronically activating B cells through sustained CD40 signaling.

Toxoplasma gondii inhibits R5 HIV-1 replication in human lymphoid tissues ex vivo

Critical events of HIV-1 pathogenesis occur in lymphoid tissues where HIV-1 is typically accompanied by infections with other pathogens (HIV co-pathogens). Co-pathogens greatly affect the clinical course of the disease and the transmission of HIV. The apicomplexan parasite Toxoplasma gondii is a common HIV co-pathogen associated with AIDS development. Here, we examined the interaction of T. gondii and HIV in coinfected human lymphoid tissue ex vivo. Both pathogens readily replicate in ex vivo infected blocks of human tonsillar tissue. Surprisingly, we found that live T. gondii preferentially inhibits R5 HIV-1 replication in coinfected tissues. This effect is reproduced by treatment of the tissue blocks with recombinant C-18, a T. gondii-encoded cyclophilin that binds to CCR5. These ex vivo findings raise the possibility that, in addition to being a co-factor in HIV disease, T. gondii may influence the outcome of viral infection by preferentially suppressing R5 variants.