Mucosal Gammadelta Research Articles

Dysregulation of IL-17/IL-22 Effector Functions in Blood and Gut Mucosal Gamma Delta T Cells Correlates With Increase in Circulating Leaky Gut and Inflammatory Markers During cART-Treated Chronic SIV Infection in Macaques.

HIV-associated inflammation has been implicated in the premature aging and increased risk of age-associated comorbidities in cART-treated individuals. However, the immune mechanisms underlying the chronic inflammatory state of cART-suppressed HIV infection remain unclear. Here, we investigated the role of γδT cells, a group of innate IL-17 producing T lymphocytes, in the development of systemic inflammation and leaky gut phenotype during cART-suppressed SIV infection of macaques. Plasma levels of inflammatory mediators, intestinal epithelial barrier disruption (IEBD) and microbial translocation (MT) biomarkers, and Th1/Th17-type cytokine functions were longitudinally assessed in blood and gut mucosa of SIV-infected, cART-suppressed macaques. Among the various gut mucosal IL-17/IL-22-producing T lymphocyte subsets including Th17, γδT, CD161+ CD8+ T, and MAIT cells, a specific decline in the Vδ2 subset of γδT cells and impaired IL-17/IL-22 production in γδT cells significantly correlated with the subsequent increase in plasma IEBD/MT markers (IFABP, LPS-binding protein, and sCD14) and pro-inflammatory cytokines (IL-6, IL-1β, IP10, etc.) despite continued viral suppression during long-term cART. Further, the plasma inflammatory cytokine signature during long-term cART was distinct from acute SIV infection and resembled the inflammatory cytokine profile of uninfected aging (inflammaging) macaques. Overall, our data suggest that during cART-suppressed chronic SIV infection, dysregulation of IL-17/IL-22 cytokine effector functions and decline of Vδ2 γδT cell subsets may contribute to gut epithelial barrier disruption and development of a distinct plasma inflammatory signature characteristic of inflammaging. Our results advance the current understanding of the impact of chronic HIV/SIV infection on γδT cell functions and demonstrate that in the setting of long-term cART, the loss of epithelial barrier-protective functions of Vδ2 T cells and ensuing IEBD/MT occurs before the hallmark expansion of Vδ1 subsets and skewed Vδ2/Vδ1 ratio. Thus, our work suggests that novel therapeutic approaches toward restoring IL-17/IL-22 cytokine functions of intestinal Vδ2 T cells may be beneficial in preserving gut epithelial barrier function and reducing chronic inflammation in HIV-infected individuals.

Isolation and Flow Cytometric Analysis of Human Endocervical Gamma Delta T Cells.

The female reproductive tract (FRT) mucosal immune system serves as the first line of defense. Better knowledge of the genital mucosa is therefore essential for understanding pathogenicity of different pathogens including HIV. Gamma delta (GD) T cells are the prototype of 'unconventional' T cells and represent a relatively small subset of T cells defined by their expression of heterodimeric T-cell receptors (TCRs) composed of gamma and delta chains. This sets them apart from the classical and much better known CD4+ helper T cells and CD8+ cytotoxic T cells that are defined by alpha-beta TCRs. GD T cells often show tissue-specific localization and are enriched in epithelium. GD T cells orchestrate immune responses in inflammation, tumor surveillance, infectious disease, and autoimmunity. Here, we present a method to reproducibly isolate and analyze human endocervical intraepithelial GD T lymphocytes. We have used endocervical cytobrush samples from women participating in the Women's Interagency HIV Infection Study (WIHS). Knowledge about GD T cells interactions during conditions in which there is an insult to the vaginal mucosal could be applied to any clinical study in which mucosal vulnerability is addressed, including the development of vaginal microbicides.In addition, knowledge about mucosal GD T cell responses has potential for application of GD T cell-based immune therapy in treating infectious diseases.

The protective effect of oral colitis-derived proteins in a murine model of inflammatory bowel disease is associated with an increase in γδ T cells in large intestinal mucosa

Oral tolerance has previously been shown effective in preventing several immune-mediated disorders in animal models. The aims of this study were to investigate the effect of oral colitis-extracted proteins (CEP) on dextran sulfate sodium (DSS)-induced colitis in BALB/c mice and to explore the relative role of the intestinal mucosal gammadelta T cells. The effect of five low oral doses of CEP on colitis was evaluated by clinical manifestation and histological lesions. Serum cytokines were measured by enzyme-linked immunosorbent assay. The percentages of the intestinal mucosal gammadelta T cells were evaluated by flow cytometry. CEP-fed colitis mice showed less severe symptoms and histological injury than bovine serum albumin (BSA)-fed control mice. Tolerized mice developed an increase in TGF-beta1 and no change in IFN-gamma serum levels. Increases in TCRgammadelta(+) T cells and CD8alpha(+)TCRgammadelta(+) T cells in small intestinal mucosal lymphocytes and no quantitative change in large intestinal mucosal lymphocytes were demonstrated in colitis mice compared to untreated mice. The proportions of TCRgammadelta(+) T cells and CD8alpha(+)TCRgammadelta(+) T cells in large intestinal mucosal lymphocytes from CEP-fed colitis mice were significantly higher compared to BSA-fed controls. The disease activity index negatively correlated with the percentages of large intestinal mucosal gammadelta T cells. Furthermore, mucosal repair in repair-period mice was also accompanied by increases in TCRgammadelta(+) T cells and CD8alpha(+)TCRgammadelta(+) T cells in large intestinal mucosal lymphocytes. Improvement of DSS-induced colitis that resulted from oral administration of colitis-extracted proteins is associated with an increase in gammadelta T cells in large intestinal mucosa.

Response of γδ T Cells to Plant-Derived Tannins

Many pharmaceutical drugs arc isolated from plants used in traditional medicines, and new plant-derived pharmaceutical drugs continue to be identified. Relevant to this review, different plant-derived agonists for gammadelta T cells are described that impart effector functions upon distinct subsets of these cells. Recently, plant tannins have been defined as one class of gammadelta T cell agonist and appear to preferentially activate the mucosal population. Mucosal gammadelta T cells function to modulate tissue immune responses and induce epithelium repair. Select tannins, isolated from apple peel, rapidly induce immune gene transcription in gammadelta T cells, leading to cytokinc production and increased responsiveness to secondary signals. Activity of these tannin preparations tracks to the procyanidin fraction, with the procyanidin trimer (C1) having the most robust activity defined to date. The response to the procyanidins is evolutionarily conserved in that responses are seen with human, bovine, and murine gammadelta T cells, although human cells show less selectivity. Procyanidin-induced responses described in this review likely account for the expansion of mucosal gammadelta T cells seen in mice and rats fed soluble extracts of tannins. Use of procyanidins to activate gammadelta T cells may represent a novel approach for the treatment of tissue damage and autoimmune diseases.

γδ T cells increase with gastric mucosal interleukin (IL)‐7, IL‐1β, and Helicobacter pylori urease specific immunoglobulin levels via CCR2 upregulation in Helicobacter pylori gastritis

The purpose of this study was to investigate possible factors that could impact on gammadelta T cell accumulation in the gastric mucosa. Subjects were 22 Helicobacter pylori (H. pylori)-free and 75 H. pylori-infected mucosa biopsies classified into grades I approximately III gastritis as per our previous study. The number of gammadelta- and 45 RO-positive T cells were determined by immunostaining. Gastric mucosal anti-H. pylori urease specific antibodies and interleukin (IL)-1beta, IL-2, 4, 7, 10 and IL-12 levels were assayed by enzyme-linked immunosorbent assay (ELISA). CC chemokine receptor 2 (CCR2) expression levels, migration, and cytokine production in gammadelta T cells stimulated by H. pylori urease were also evaluated. The gammadelta T cell count was significantly higher in grade III gastritis which exhibits strong immunoglobulin (Ig)A and IgG responses to H. pylori urease with lymphoid follicles than in other groups. gammadelta T cell count was significantly correlated with IL-1beta and interleukin-7 (IL-7) levels in the gastric mucosa. H. pylori urease immunoreactivity was detected in lamina propria of grade III gastritis, along with many gammadelta T cells. After H. pylori eradication therapy, the gammadelta T cell count in grade III gastritis significantly decreased. H. pylori urease stimulated significant increases in CCR2 expression levels, although to a lesser degree than those induced by IL-7 stimulation in both peripheral and mucosal gammadelta T cells. Interferon (IFN)-gamma and IL-10 production was also stimulated by H. pylori urease in peripheral gammadelta T cells. Gastric mucosal increases in IL-7 and IL-1beta closely corresponded to the accumulation of gammadelta T cells in gastric mucosa. An association was also seen between gammadelta T cell accumulation and H. pylori urease-specific Ig levels.

Studies in vivo of ω-gliadins in gluten sensitivity (coeliac sprue disease)

1. Highly purified omega-gliadins from wheat were used to challenge gluten-sensitized individuals. Characteristic responses by mucosal CD3(+) and gamma delta+ lymphocytes were demonstrated. Each lymphocyte subset showed an increase within 8-12 h post-challenge, indicating a specific response by the rectal mucosa to this gliadin species.2. Available sequence data for the omega-gliadins and homologous proteins from barley and rye indicate a common repeating octapeptide motif (consensus PQQPFPQQ). The results indicate, therefore, that the octapeptide repeat, or a contained sequence such as PQQP, plays an important role in the mucosal immunopathology of gluten sensitivity.

Expression of the mucosal γδ T cell receptor V region repertoire in patients with IgA nephropathy

IgA nephropathy (IgAN) is characterized by the deposition of IgA in the glomerular mesangium and often leads to progressive renal dysfunction and kidney failure. We have previously shown that the mesangial IgA is likely to derive from the bone marrow plasma cells, and suggested that a primary abnormality within the mucosal immune system may underly the pathogenesis of IgAN. This study has analyzed the T cell receptor (TCR) variable (V) region expression by gamma delta T cells in the intestinal mucosa of patients with IgAN. The V gamma and V delta usage of TCR transcripts was determined using a semiquantitative reverse transcriptase-PCR protocol. Primers specific for the four human V gamma and six V delta subfamilies were used each with a constant (C) gamma or C delta specific primer, and the PCR-amplified TCR transcripts were detected by Southern blotting and oligonucleotide hybridization. gamma delta TCR V region expression was determined in gut biopsies and peripheral blood of 11 patients with IgAN, and the TCR V gamma and V delta repertoires were compared to those in gut and peripheral blood of 11 control individuals. gamma delta T cells in normal blood predominantly expressed V gamma 2 (V gamma 9 gene) and V delta 2 gene segments whereas those in normal gut mainly expressed V gamma 3 and V delta 3. In IgAN patients, V delta 2 was also the predominant V delta gene utilized by peripheral blood gamma delta T cells, however, we observed a predominance of V gamma 3 and reduced V gamma 2 usage by these cells. gamma delta T cells in the gut of IgAN patients mainly used V gamma 3 and V delta 1. While the gamma and delta TCR V region repertoires did not differ significantly between the peripheral blood of patients and controls, there were significant differences in V gamma and V delta repertoire expression between IgAN and control gut biopsies. V gamma 3 gene expression was significantly decreased in IgAN gut compared to control gut (P = 0.023). In addition, there was a significant decrease in V delta 3 gene expression in IgAN gut compared to control gut (P = 0.043). These findings indicate that a subpopulation of gamma delta T cells, which represent the majority of gamma delta T cells in normal gut mucosa, are significantly diminished in the gut of patients with IgAN. This suggests that a "hole" in the mucosal gamma delta T cell repertoire may play a fundamental role in contributing to the pathogenesis of IgAN.

Changes in Human Mucosal γδ T Cell Repertoire and Function Associated with the Disease Process in Inflammatory Bowel Disease

Although gamma delta T cells are a major component of the human intestinal mucosa, it is not clear what role they play in mucosal immunity or if they are involved in the disease process of inflammatory bowel disease (IBD). Flow cytometry and reverse transcriptase-polymerase chain reaction (RT-PCR) assays were used to identify quantitative and qualitative changes in the repertoire of gamma delta T cells present in surgical and/or biopsy samples or normal and inflamed colon from individual patients with ulcerative colitis (UC) or Crohn's disease (CD). Cytokine production and the ability to adhere to and interact with colonic fibroblasts were used to compare the functional properties of gamma delta T cells isolated from the normal and diseased colonic mucosa. Increased numbers of gamma delta T cells localized in areas of inflammation and tissue injury were found in the majority of patients, irrespective of the type of IBD present. This expansion was attributable to an increase in V delta 1+ cells expressing a V delta 1-(D delta 3)-J delta 1-encoded T cell receptor and was seen in patients with severe disease as well as those with newly diagnosed or less severe forms of IBD. Among T cells present in the inflamed mucosa of patients with CD, gamma delta T cells, particularly V delta 1+ cells, were a major source of the proinflammatory cytokine interferon-gamma and could interact with colonic fibroblasts. Our results demonstrate that the chronic inflammatory immune response characteristic of IBD is associated with distinct changes in the number, distribution, composition, and function of mucosal gamma delta T cells. Through the production of cytokines and physical interaction with other cells, gamma delta T cells can perform an immunoregulatory function and contribute to the pathophysiology of IBDs.

Phenotypic and molecular characteristics of nasal mucosal gamma delta T cells in allergic and infectious rhinitis.

T cells expressing the T-cell receptor (TCR) gamma delta home in on various epithelia and may play an important role in local immunity to foreign antigens. The nasal mucosa is a potential site for chronic inflammatory diseases, yet little is known about the characteristics of nasal mucosal gamma delta T cells. Using flow cytometry, immunohistochemistry, and RT-PCR, we elucidated the characteristics of nasal mucosal gamma delta T cells in patients with perennial allergic rhinitis (PAR), chronic infective rhinitis (CIR), and seasonal allergic rhinitis (SAR) and in normal subjects. The gamma delta T cells were significantly increased in the nasal mucosa of patients with PAR (PAR, 24.3 +/- 4.5%; CIR, 12.9+/- 2.7%, p < 0.01), unrelated to those in autologous peripheral blood (PAR, 5.6 +/- 0.8%; CIR, 9.6 +/- 2.8%), and they were preferentially distributed in the epithelial compartment (26.7 +/- 2.3%) rather than in the lamina propria (5.4 +/- 2.5%). Of the expanded population of nasal mucosal gamma delta T cells in patients with PAR, CD4+ and CD4-8- gamma delta T cells were selectively increased (p < 0.01). Although nasal intraepithelial gamma delta T cells from all groups of patients and normal subjects dominantly expressed the V gamma 1/V delta 1 genes, and a bias for V gamma 3 gene expression was noted in those of patients with PAR, a significantly larger fraction of nasal mucosal gamma delta T cells in patients with PAR expressed the V gamma 1/V delta 1 TCR (p, 0.01), whereas those of the peripheral blood expressed the V gamma 2/V delta 2 TCR. More than 60% of V gamma 1/V delta 1 TCR+ cells in patients with PAR, were CD45RO+ ("memory cells"), independent of those in their peripheral blood (p < 0.01). Furthermore, a substantial proportion of nasal mucosal gamma delta T cells in patients with PAR synthesized IL-4 and IL-5 but negligible amounts of IFN-gamma. These observations of an increase in the proportions and activation of distinct subsets of nasal mucosal gamma delta T cells and their Th2-type cytokine profile in patients with PAR, unrelated to those in autologous peripheral blood suggest an important role for the oligoclonally expanded expanded nasal mucosal gamma delta T cells in the pathogenesis of PAR.

Role of gamma delta T cells in the regulation of mucosal IgA response and oral tolerance.

In this short review, we first described experiments that show that prolonged oral immunization with a protein vaccine, such as DT, induced systemic unresponsiveness in the presence of antigen-specific mucosal IgA responses. Mucosal T cells, such as IEL, may play an important role for the maintenance of antigen-specific IgA responses because these T cells are able to respond to stimulation signals provided by antigen even when T-cell unresponsiveness was induced in systemic tissue, such as spleen of mice orally tolerized with the protein DT. Inasmuch as IEL contain a high frequency of gamma delta T cells, it was logical to postulate that mucosal gamma delta T cells are essential regulatory T cells for the induction of IgA responses in oral tolerance. To this end, our previous studies showed that adoptive transfer of mucosal gamma delta T cells from IEL of mice orally tolerized with SRBC to the recipient mice with systemic unresponsiveness to the same antigen resulted in the abrogation of unresponsiveness to Ig synthesis, including those of IgA isotype. In this regard, when the mucosal immune system of TCR-delta-/- and their control mice was examined, lower numbers of IgA antibody-producing cells were noted in TCR-delta-/- mice in comparison to control background mice. Further, the level of IgA in fecal extracts was also low in TCR-delta-/- mice. These findings suggested that loss of gamma delta T cells in down-regulation of IgA B-cell responses.

PATHOGENIC MECHANISMS OF HELICOBACTER PYLORI

There is general agreement that motility, urease activity, and association with gastric mucosal cells are important virulence factors of H. pylori. Urease activity is perhaps the best characterized of these factors. Presumably, urease activity creates a "cloud" of ammonia around the bacterium, thus neutralizing the lethal effects of gastric acid. Motility allows the bacterium to penetrate the mucus layer and promotes specific association of the bacteria with epithelial cells, further allowing evasion of gastric acidity. The association between gastrin levels and H. pylori infection is currently the most thoroughly studied feature relating to pathogenesis in vivo. Prolonged hypergastrinemia associated with H. pylori infection may contribute to increased parietal cell mass and chronically increased secretion of gastric acid; however, long-term studies are needed to validate this hypothesis. The identification of mucosal gamma delta T cells and immunologic cross-reactivity between H. pylori and gastric cells implies that the immune response contributes significantly to the pathogenesis of H. pylori. The role of the immune system in modulating H. pylori infection requires further study. Although many putative pathogenic factors have been identified on the basis of in vitro phenomena alone, their significance in vivo is not known. Ultimately, it will be necessary to evaluate the significance of these factors in animal models by using isogenic strains of H. pylori that differ only in a single genotypic characteristic.