Figure 3.
Involvement of CCR4+ cells in high background interferon (IFN)‐γ production by peripheral blood mononuclear cells (PBMCs) from human T‐cell leukemia virus type‐1 (HTLV‐1)–associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients. PBMCs from a HAM/TSP patient (#255) were equally divided into four fractions. One fraction was used directly (whole) and the other three fractions were used after depletion of CD4+[CD4(–)], CD8+[CD8(–)], or CCR4+[CCR4(–)] cells. The cell numbers in each fraction were 2.0 × 105, 0.8 × 105, 1.1 × 105, and 1.3 × 105 cells/well, respectively. (a) The fractions were cultured alone (open bars), with glutathione‐S‐transferase (GST) (gray bars), or with a mixture of GST–Tax proteins (black bars) for 4 days. IFN‐γ in the supernatants was measured by enzyme‐linked immunoabsorbent assay (ELISA). (b) The levels of HTLV‐1 p19 antigen in the culture supernatants of the PBMC fractions in the absence of any stimulation were measured by ELISA at 7 days after the initiation of culture. The results represent the mean ± SD of duplicate wells. (c, d) Whole and fractionated PBMCs from two other HAM/TSP patients (#288 and #290) were similarly examined for IFN‐γ production for 4 days against medium (open bars), GST (gray bars), or GST–Tax proteins (black bars) (c), and HTLV‐1 p19 production for 7 days of culture (d). The cell numbers of whole, CCR4(–), and CD8(–) PBMCs in each well were 2.0 × 105, 1.7 × 105, and 1.2 × 105 for patient #288, and 2.1 × 105, 0.9 × 105, and 0.9 × 105 for patient #290, respectively. (e) HTLV‐1 provirus numbers (copies/1000 cells) in whole (open bars), CCR4+ cell‐depleted (closed bars), and CCR4+ (hatched bars) PBMC fractions before culture from subjects #255 (HAM/TSP) and #211 (AC) were measured by real‐time polymerase chain reaction methods. The results represent the mean ± SD of duplicate samples.