Direct enumeration ofBorrelia-reactive CD4 T cellsex vivoby using MHC class II tetramers

Abstract

We characterized antigen-specific CD4 ؉ T cells in six patients with treatment-resistant Lyme arthritis, using an HLA-DRB1*0401 major histocompatibility complex (MHC) class II tetramer covalently loaded with OspA164-175, an immunodominant epitope of Borrelia burgdorferi. Direct analysis of OspA-tetramer binding CD4 ؉ cells in patients expressing the HLA-DRB1*0401 allele revealed frequencies of between <0.005 and 0.1% in peripheral blood (n ‫؍‬ 6), and between <0.005 and 3.1% in synovial fluid (n ‫؍‬ 3). OspAtetramer ؉ CD4 ؉ cells were directly cloned at 1 cell per well and expanded by mitogen and IL-2 on allogeneic feeder cells. As measured by [ 3 H]thymidine incorporation, 95% of 168 T cell clones from synovial fluid binding the OspA-tetramer were antigenreactive. Clones generated from peripheral blood revealed a different pattern of responsiveness when compared with clones generated from synovial fluid, as measured by proliferation, IFN-␥, and IL-13 secretion. These clones, selected on the basis of their peptide binding, also responded to whole protein, but with a different cytokine profile. Our studies demonstrate that MHC class II tetramers can be used in humans to directly identify, isolate, and characterize antigen-reactive T cells from an inflammatory compartment.

Therapy for Lyme arthritis: Strategies for the treatment of antibiotic‐refractory arthritis

“…Moreover, patients with antibiotic-refractory arthritis often have T cell recognition of OspA 163-175 (29)(30)(31)(32)(33). Because this epitope does not occur in the more common causative agents of Lyme borreliosis in Europe, Borrelia afzelii and Borrelia garinii (32), this mechanism may apply primarily to American patients with antibiotic-refractory Lyme arthritis.…”

Efficient Immunization and Cross-Priming by Vaccine Adjuvants Containing TLR3 or TLR9 Agonists Complexed to Cationic Liposomes

“…Negative controls included spleens from naive mice and use of tetramers prepared with irrelevant peptides. For analysis of Ag-specific CD4 T cells, MHC class II tetramers were produced with a covalent linkage to the gp61-80 peptide of LCMV (I-A b -gp61) as described previously (49). After incubation with tetramers, cells were washed and stained with Abs to surface determinants, then washed and stained with appropriate streptavidin conjugates for 15 min, then washed and resuspended in FACS buffer or fixed in 1% paraformaldehyde before analysis.…”

MHC class II Tetramer Guided Detection of Mycobacterium tuberculosis‐specific CD4+ T Cells in Peripheral Blood from Patients with Pulmonary Tuberculosis

“…A number of MHC class II tetramer complexes have successfully been developed to gauge CD4 + T cells directed against viral pathogens [7][8][9]. Up to now, antigen-specific CD4 + Tcell responses directed against bacterial pathogens have only been visualized in synovial fluid in three patients with Lyme arthritis [10]. We report here the identification of HLA-DR4 tetramer-reactive CD4 + T cells in situ (in granuloma tissue) and in blood directed against antigens shared among MTB and MOTT, i.e.…”