Figure 1.

IL-2/α-CD40 induces NOS expression via tumor-associated macrophages. (A) WT and GKO mice were treated between days 11 and 21 as indicated in the Materials and methods. On day 22, mice were euthanized, and the primary tumor was dissected and homogenized in RIPA lysis buffer. The expression of NOS2 was determined by Western blot analysis. Hypoxanthine phosphoribosyltransferase (HPRT) was used as a loading control. (B and C) Tumors from treated WT mice were dissected on day 22, fixed in formalin, and analyzed by two-color immunofluorescent microscopy (200× magnification). NOS2 immunoreactivity was visualized using Alexa Fluor 488 (green). (B and C) F4/80 (B) and GR1 (C) immunoreactivity was visualized using Alexa Fluor 594 (red). In some regions, colocalization of the two colors results in yellow staining. The results are representative of three separate experiments. Bars, 100 µm.