Jin L, Raymond D P, Crabtree T D, Pelletier S J, Houlgrave C W, Pruett T L, Sawyer R G
Surgical Infectious Disease Laboratory, Department of Surgery, and Department of Internal Medicine, University of Virginia, Charlottesville, VA 22906, USA.
J Immunol. 2000 Nov 1;165(9):5153-60. doi: 10.4049/jimmunol.165.9.5153.
The immunomodulatory role of unmethylated cytosine-guanine sequences (CpG) in bacterial DNA has been well documented. We have previously demonstrated that murine macrophage-like RAW 264.7 cells respond to CpG DNA with an increase in the proinflammatory cytokine, TNF-alpha, in both a dose-dependent and time-dependent manner. In addition, CpG DNA stimulates a significant, though delayed, secretion of the anti-inflammatory cytokine IL-10. Because TNF-alpha and TNFR (TNFRI and II) expression are tightly regulated responses, we hypothesized that CpG containing oligodeoxynucleotide (CpG ODN) would also affect TNFRI and II shedding. Using both murine peritoneal macrophages and RAW 264.7 cells, we demonstrated a significant, time-dependent increase in soluble TNFRI and TNFRII production with CpG ODN stimulation. RAW 264.7 cells treated with CpG ODN had a transient increase in membrane TNFRII expression, but not TNFRI. Both types of TNFR mRNA were also up-regulated by CpG ODN, and addition of the transcriptional inhibitor actinomycin D abrogated the effect of CpG ODN on TNFR mRNA and protein expression. Addition of anti-IL-10 and anti-TNF-alpha Abs did not change these results. The addition of plate-bound anti-TNF receptor Abs to this system increased the amount of bioactive TNF, implying that these receptors are acting as inhibitors of TNF activity. These results suggest that the de novo, non-IL-10- and non-TNF-alpha-dependent transcription, translation, and shedding of TNFRs are additional potential counterinflammatory effects of CpG DNA.
细菌DNA中未甲基化的胞嘧啶-鸟嘌呤序列（CpG）的免疫调节作用已有充分记载。我们之前已证明，鼠巨噬细胞样RAW 264.7细胞对CpG DNA的反应是促炎细胞因子TNF-α呈剂量依赖性和时间依赖性增加。此外，CpG DNA刺激抗炎细胞因子IL-10显著分泌，不过有延迟。由于TNF-α和TNFR（TNFRI和II）的表达是受到严格调控的反应，我们推测含CpG的寡脱氧核苷酸（CpG ODN）也会影响TNFRI和II的脱落。使用鼠腹膜巨噬细胞和RAW 264.7细胞，我们证明了CpG ODN刺激后可溶性TNFRI和TNFRII的产生呈显著的时间依赖性增加。用CpG ODN处理的RAW 264.7细胞，膜TNFRII表达有短暂增加，但TNFRI没有。两种类型的TNFR mRNA也被CpG ODN上调，添加转录抑制剂放线菌素D可消除CpG ODN对TNFR mRNA和蛋白表达的影响。添加抗IL-10和抗TNF-α抗体并没有改变这些结果。向该系统中添加平板结合的抗TNF受体抗体增加了生物活性TNF的量，这意味着这些受体作为TNF活性的抑制剂发挥作用。这些结果表明，TNFR从头开始的、非IL-10和非TNF-α依赖性的转录、翻译和脱落是CpG DNA额外的潜在抗炎作用。
