A T-cell subset, defined as CD4+CD25hi (regulatory T-cells [Treg cells]), was recently shown to suppress T-cell activation. We demonstrate that human Treg cells isolated from healthy donors express the HIV-coreceptor CCR5 and are highly susceptible to HIV infection and replication. Because Treg cells are present in very few numbers and are difficult to expand in vitro, we genetically modified conventional human T-cells to generate Treg cells in vitro by ectopic expression of FoxP3, a transcription factor associated with reprogramming T-cells into a Treg subset. Overexpression of FoxP3 in naïve human CD4+ T-cells recapitulated the hyporesponsiveness and suppressive function of naturally occurring Treg cells. However, FoxP3 was less efficient in reprogramming memory T-cell subset into regulatory cells. In addition, FoxP3-transduced T-cells also became more susceptible to HIV infection. Remarkably, a portion of HIV-positive individuals with a low percentage of CD4+ and higher levels of activated T-cells have greatly reduced levels of FoxP3+CD4+CD25hi T-cells, suggesting disruption of the Treg cells during HIV infection. Targeting and disruption of the T-cell regulatory system by HIV may contribute to hyperactivation of conventional T-cells, a characteristic of HIV disease progression. Moreover, the ability to reprogram human T-cells into Treg cells in vitro will greatly aid in decoding their mechanism of suppression, their enhanced susceptibility to HIV infection, and the unique markers expressed by this subset.

The chytrid fungus, Batrachochytrium dendrobatidis, causes chytridiomycosis and is a major contributor to global amphibian declines. Although amphibians have robust immune defenses, clearance of this pathogen is impaired. Because inhibition of host immunity is a common survival strategy of pathogenic fungi, we hypothesized that B. dendrobatidis evades clearance by inhibiting immune functions. We found that B. dendrobatidis cells and supernantants impaired lymphocyte proliferation and induced apoptosis; however, fungal recognition and phagocytosis by macrophages and neutrophils was not impaired. Fungal inhibitory factors were resistant to heat, acid, and protease. Their production was absent in zoospores and reduced by nikkomycin Z, suggesting that they may be components of the cell wall. Evasion of host immunity may explain why this pathogen has devastated amphibian populations worldwide.

Background Necrotising enterocolitis (NEC) is the most common gastrointestinal emergency in premature infants. Immaturity of gastrointestinal immune regulation may predispose preterm infants to NEC as FOXP3 T regulatory cells (Treg) are critical for intestinal immune homoeostasis. Objective To investigate the hypothesis that abnormal developmental regulation of lamina propria Treg would define premature infants with NEC. Design Lamina propria mononuclear cell populations from surgically resected ileum from 18 patients with NEC and 30 gestational age-matched non-NEC surgical controls were prospectively isolated. Polychromatic flow cytometry was performed to phenotype and analyse lamina propria T cell populations. The cytokine gene expression profile in NEC tissue was compared with that of non-NEC controls. Results The total number of Treg, CD4, or CD8 T cells in each ileum section was independent of gestational age, age or postmenstrual age and similar between patients with NEC and controls. In contrast, the ratio of Treg to CD4 T cells or Treg to CD8 T cells was significantly lower in NEC ileum than in infants without NEC (medians 2.9% vs 6.6%, p=0.001 and medians 6.6% vs 25.9%, p<0.001, respectively). For any given number of CD4 or CD8 T cells, Treg were, on average, 60% lower in NEC ileum than in controls. NEC tissue cytokine gene expression profiles were characteristic of inhibited Treg development or function. Treg/CD4 and Treg/CD8 ratios recovered between initial resection for NEC and reanastomosis. Conclusion The proportion of lamina propria Treg is significantly reduced in the ileum of premature infants with NEC and may contribute to the excessive inflammatory state of this disease.

Rationale: Effective therapeutic interventions for chronic, idiopathic lung diseases remain elusive. Normalized T-cell function is an important contributor to spontaneous resolution of pulmonary sarcoidosis. Up-regulation of inhibitor receptors, such as programmed death-1 (PD-1) and its ligand, PD-L1, are important inhibitors of T-cell function.Objectives: To determine the effects of PD-1 pathway blockade on sarcoidosis CD41 T-cell proliferative capacity.Methods: Gene expression profiles of sarcoidosis and healthy control peripheral blood mononuclear cells were analyzed at baseline and follow-up. Flow cytometry was used to measure ex vivo expression of PD-1 and PD-L1 on systemic and bronchoalveolar lavage-derived cells of subjects with sarcoidosis and control subjects, as well as the effects of PD-1 pathway blockade on cellular proliferation after T-cell receptor stimulation. Immunohistochemistry analysis for PD-1/PD-L1 expression was conducted on sarcoidosis, malignant, and healthy control lung specimens.

Sarcoidosis pathogenesis is characterized by peripheral anergy and an exaggerated, pulmonary CD4+ Th1 response. In this study, we demonstrate that CD4+ anergic responses to polyclonal TCR stimulation are present peripherally and within the lungs of sarcoid patients. Consistent with prior observations, spontaneous release of IL-2 was noted in sarcoidosis bronchoalveolar lavage CD4+ T cells. However, in contrast to spontaneous hyperactive responses reported previously, the cells displayed anergic responses to polyclonal TCR stimulation. The anergic responses correlated with diminished expression of the Src kinase Lck, PKC-θ and NF-κB, key mediators of IL-2 transcription. Although Tregs were increased in sarcoid patients, Treg depletion from the CD4+ T cell population of sarcoidosis patients did not rescue IL-2 and IFN-γ production; whereas, restoration of the IL-2 signaling cascade, via PKC-θ overexpression, did. Furthermore, sarcoidosis Tregs displayed poor suppressive capacity indicating that T cell dysfunction was a global CD4+ manifestation. Analyses of patients with spontaneous clinical resolution revealed that restoration of CD4+ Th1 and Treg cell function was associated with resolution. Conversely, disease progression exhibited decreased Th1 cytokine secretion and proliferative capacity, and reduced Lck expression. These findings implicate normalized CD4+ T cell function as a potential therapeutic target for sarcoidosis resolution.

Topical antimicrobicides hold great promise in reducing human immunodeficiency virus (HIV) transmission. Amphibian skin provides a rich source of broad-spectrum antimicrobial peptides including some that have antiviral activity. We tested 14 peptides derived from diverse amphibian species for the capacity to inhibit HIV infection. Three peptides (caerin 1.1, caerin 1.9, and maculatin 1.1) completely inhibited HIV infection of T cells within minutes of exposure to virus at concentrations that were not toxic to target cells. These peptides also suppressed infection by murine leukemia virus but not by reovirus, a structurally unrelated nonenveloped virus. Preincubation with peptides prevented viral fusion to target cells and disrupted the HIV envelope. Remarkably, these amphibian peptides also were highly effective in inhibiting the transfer of HIV by dendritic cells (DCs) to T cells, even when DCs were transiently exposed to peptides 8 h after virus capture. These data suggest that amphibian-derived peptides can access DC-sequestered HIV and destroy the virus before it can be transferred to T cells. Thus, amphibian-derived antimicrobial peptides show promise as topical inhibitors of mucosal HIV transmission and provide novel tools to understand the complex biology of HIV capture by DCs.

CD4+CD25+ human regulatory T cells (Treg cells), which express the transcription factor FoxP3, suppress T cell activation. In this study, we sought to define cellular and molecular mechanisms of human Treg cell differentiation. A subset of human naive CD4+ T cells that are CD25+ express high levels of FoxP3. We show that upon activation through the TCR, these FoxP3-expressing naive T cells (termed TNreg cells) greatly expand in vitro. Expanded TNreg cells acquire a full Treg phenotype with potent suppressive activity and display low IL-2 production upon TCR stimulation. TNreg cells in which FoxP3 expression was reduced through RNA interference lost their suppressive activity, but retained their low IL-2 secretion in response to TCR stimulation. Furthermore, in support of the notion that TNreg cells represent a separate lineage of naive cells, we found that they were more susceptible to HIV infection as compared with naive CD4+ T cells. Based on these findings, we propose that TNreg cells are precursors for human Treg cells and that these cells require a high level of FoxP3 expression to maintain their suppressive function. Accordingly, modulation of TNreg cell numbers during infections such as HIV may disrupt human Treg cell development, and contribute to chronic immune activation.

Considerable evidence supports the concept that CD4؉ T cells are important in sarcoidosis pathogenesis, but the antigens responsible for the observed Th1 immunophenotype remain elusive. The epidemiologic association with bioaerosols and the presence of granulomatous inflammation support consideration of mycobacterial antigens. To explore the role of mycobacterial antigens in sarcoidosis immunopathogenesis, we assessed the immune recognition of mycobacterial antigens, the 6-kDa early secreted antigenic protein (ESAT-6) and catalase-peroxidase (KatG), by T cells derived from bronchoalveolar lavage (BAL) fluid obtained during diagnostic bronchoscopy. We report the presence of antigen-specific recognition of ESAT-6 and KatG in T cells from BAL fluid of 32/44 sarcoidosis subjects, compared to 1/27 controls (P < 0.0001). CD4