The presence of activated CD4⁺ T cells results in enhanced DC activation and heightened inflammation within the tumor microenvironment. The schema outlines the timeline of experimental procedures. Mice with subcutaneous A20HA tumors were divided into 4 groups when tumor size reached ∼ 170 mm²: no treatment, CD4⁺ T-cell transfer only, Cy only, and Cy plus CD4⁺ T-cell transfer. Seven days after T-cell transfer, tumor masses were excised and analyzed. (A) The immune status of tumor-infiltrating donor CD4⁺ T cells in mice with or without chemotherapy. Cells were stained as described in Figure 1. The frequency and cell division of the transferred CD4⁺ T cells, CD40L profile relative to cell division, and cytokine profile in divided cells are shown in representative plots. Numbers represent the percentages of cells in the given gate or quadrant. Bar graphs summarize results from 3 independent experiments with 3 mice per group in each experiment (mean ± SD). (B) Enhanced activation of intratumoral CD11c⁺ cells in mice receiving CD4⁺ T-cell transfer after Cy. Representative histograms show expression profiles of CD40 and B7.1 in gated CD11c⁺ cells in tumors resected from mice treated with Cy or Cy plus CD4⁺ T-cell transfer. Bar graphs summarize the median fluorescence intensity (MFI) of CD40 and B7.1 shown as mean ± SD (3 samples per group). (C) Cytokine/chemokine profile in tumor mass. Total RNA was extracted from resected tumor tissues and subjected to quantitative real-time PCR for the indicated genes. Each sample was measured in triplicate for each gene. Data represent the relative amount of target mRNA normalized to β-actin (mean ± SD). Data shown are results from 1 of 3 independent experiments with similar results (*P < .05, **P < .005, ***P < .001).