Figure 8.
Figure 8. The measured decrease in TNFα production is not due to alterations in mRNA production but to retention of TNFα within monocytes. (A) Cytokine mRNA levels were assessed by TaqMan RT-PCR. The results are expressed as the relative change in mRNA compared with a calibrator. (B,C) Intracellular cytokine staining by FACS revealed that TNFα, but not IL-6, was being retained within monocytes after culture in the presence of LPS and anti-MyD-1. The results are expressed as the percentage TNFα-positive CD14+ cells, and these data are representative of 3 experiments. (D) Cell lysates were prepared from human PBMCs stimulated with either LPS (100 ng/mL) plus anti-MyD-1 (10 μg/mL) or LPS plus IgG1 (10 μg/mL) for 24 hours at 37°C. The cell lysates were then subjected to Western blot analysis of TACE expression as described in “Materials and methods.” These lysates were prepared from a single donor and were representative of those from 2 other independent donors.

The measured decrease in TNFα production is not due to alterations in mRNA production but to retention of TNFα within monocytes. (A) Cytokine mRNA levels were assessed by TaqMan RT-PCR. The results are expressed as the relative change in mRNA compared with a calibrator. (B,C) Intracellular cytokine staining by FACS revealed that TNFα, but not IL-6, was being retained within monocytes after culture in the presence of LPS and anti-MyD-1. The results are expressed as the percentage TNFα-positive CD14+ cells, and these data are representative of 3 experiments. (D) Cell lysates were prepared from human PBMCs stimulated with either LPS (100 ng/mL) plus anti-MyD-1 (10 μg/mL) or LPS plus IgG1 (10 μg/mL) for 24 hours at 37°C. The cell lysates were then subjected to Western blot analysis of TACE expression as described in “Materials and methods.” These lysates were prepared from a single donor and were representative of those from 2 other independent donors.

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