CD8αα+ T cells show amoeboid shape and frequent morphological change in vitro, and localize to small intestinal intraepithelial region in vivo.

Intraepithelial lymphocytes (IELs) are very unique in the intestinal immune system. They include γδT cells and CD4-CD8-TCRαß+T cells (double negative: DNT), both of which are specific for the intestine, in addition to CD4+ and CD8+ T cells. IELs exist within the monolayer of the intestinal epithelial cells and dynamically move between lamina propria (LP) and intraepithelial (IE) region. The localization and movement patterns of IEL subsets and the regulatory factors have been unknown. Here, we developed a novel in vitro live imaging system and quantified the motility and morphological changes among subsets of IELs. We identified CD8αα as the key regulatory factor. IELs, especially γδ and DNT cells, showed amoeboid shape and frequent morphological change, while most T cells in MLN or SP showed round shape in vitro. TCR signal, IL-15, gut microbes, CCL25, and integrin αEß7 expression were non-essential for IEL movement in vitro. CD8αα+ cells showed higher motility and larger morphological changes than CD8αα- cells. Adoptive transferred CD8αα+CD4-IELs localized to IE region of recipient NSG mice, while CD8αα-CD4-IELs localized to the LP. Our results showed that the CD8αα/TL signal is essential for the localization of IELs to IE region in vivo. CD8αα/TL may be an effective target to increase the number of IELs, which protects against intestinal infection, allergy, tumorigenesis or inflammation.

Ultrasensitive DNA Immune Repertoire Sequencing Using Unique Molecular Identifiers.

BACKGROUND: Immune repertoire sequencing of the T-cell receptor can identify clonotypes that have expanded as a result of antigen recognition or hematological malignancies. However, current sequencing protocols display limitations with nonuniform amplification and polymerase-induced errors during sequencing. Here, we developed a sequencing method that overcame these issues and applied it to γδ T cells, a cell type that plays a unique role in immunity, autoimmunity, homeostasis of intestine, skin, adipose tissue, and cancer biology. METHODS: The ultrasensitive immune repertoire sequencing method used PCR-introduced unique molecular identifiers. We constructed a 32-panel assay that captured the full diversity of the recombined T-cell receptor delta loci in γδ T cells. The protocol was validated on synthetic reference molecules and blood samples of healthy individuals. RESULTS: The 32-panel assay displayed wide dynamic range, high reproducibility, and analytical sensitivity with single-nucleotide resolution. The method corrected for sequencing-depended quantification bias and polymerase-induced errors and could be applied to both enriched and nonenriched cells. Healthy donors displayed oligoclonal expansion of γδ T cells and similar frequencies of clonotypes were detected in both enrichment and nonenriched samples. CONCLUSIONS: Ultrasensitive immune repertoire sequencing strategy enables quantification of individual and specific clonotypes in a background that can be applied to clinical as well as basic application areas. Our approach is simple, flexible, and can easily be implemented in any molecular laboratory.

Proposal for the designation of the natural killer antigens-positive γδ T-cell subset as γδ NKT-cells: nomenclature based on immunoprofile.

Natural killer T (NKT)-cells with both T- and NK-cell antigens can be classified into αß or γδ type according to the TCR gene expression. The WHO classification of lymphoid neoplasms did not further subdivide the above-mentioned NKT-cell malignancies according to the expression of these TCR types. γδ T-cells can be stimulated and expanded by Zoledronic acid, usually carrying Vγ9 Vδ2 TCR and various NK-associated receptors (NKR) such as CD56, CD94, CD158a, CD158b, CD161, etc. In contrast, αß T-type NKT-cells are positive for Vα24 Vß11 TCR. NKR positive γδ T-cells have clearly different features than the NKT-cells with Vα24 Vß11 TCR type, αß NKT. NKT-cells carrying γδ TCR should be classified and named as γδ NKT-cells to distinguish the cells explicitly from αß NKT-cells.

The Dual Roles of Human γδ T Cells: Anti-Tumor or Tumor-Promoting.

γδ T cells are the unique T cell subgroup with their T cell receptors composed of γ chain and δ chain. Unlike αß T cells, γδ T cells are non-MHC-restricted in recognizing tumor antigens, and therefore defined as innate immune cells. Activated γδ T cells can promote the anti-tumor function of adaptive immune cells. They are considered as a bridge between adaptive immunity and innate immunity. However, several other studies have shown that γδ T cells can also promote tumor progression by inhibiting anti-tumor response. Therefore, γδ T cells may have both anti-tumor and tumor-promoting effects. In order to clarify this contradiction, in this review, we summarized the functions of the main subsets of human γδ T cells in how they exhibit their respective anti-tumor or pro-tumor effects in cancer. Then, we reviewed recent γδ T cell-based anti-tumor immunotherapy. Finally, we summarized the existing problems and prospect of this immunotherapy.