Proper macrophage activation in a T cell-deficient F344/NJcl-rnu/rnu rat.

<p>Bone marrow macrophages (BMMs) were prepared from a T cell-deficient F344/NJcl-rnu/rnu rat and a wild type F344/Jcl rat as described in Materials and Methods in detail. Phagocytic activity (A) and response to inflammatory stimulation with LPS (B, C) of BMMs prepared were then evaluated. To examine phagocytic activity, BMMs from both strains were treated with iron-containing nanoparticles, Ferumoxytol, for 24 h and engulfed particles were detected by Berlin blue staining (A). Bar, 20 μm. Response of BMMs from both strains to LPS was assessed by Western blot analysis and RT-PCR analyses targeting NF-κB-regulated pro-inflammatory genes. For Western blotting, BMMs were stimulated with LPS (1 μg/ml) for indicated time period and total cell lysate prepared from these stimulated cells was subjected to Western blotting for IκBα, phosphorylated form of p65 subunit (s536, p-p65) and p65 subunit to evaluate NF-κB activation, using Western blotting for α-tubulin as an internal control. For RT-PCR analyses, total RNA was purified from stimulated cells (LPS 0.1, 1, 10 μg/ml, 60 min) and subjected to RT-PCR analyses for MCP-1 (<i>Ccl2</i>), TNF-α (<i>Tnf</i>) and COX-2 (<i>Ptgs2</i>) after reverse transcription. Data represents mean ± SEM (n = 4). Noted that BMMs from a T cell-deficient rat shows a similar phagocytic activity and properly responses to inflammatory stimulation as in a wild type BMMs.