槐定碱与TLR4/MD—2阻断剂对LPS诱导的RAW264.7巨噬细胞TLR4—NF—κB—TNF—α通路的影响(4)
[Abstract]Objective： To study the effect of sophoridine and TLR4/MD-2 blocking agent on pathway of LPS-induced RAW264.7 macrophage TLR4-NF-κB-TNF-α in and its pharmacological mechanism of antiendotoxin. Method： RAW264.7 macrophages were cultured and divided into 5 groups， namely the blank control group， the LPS model group， the sophoridine+LPS group， the TLR4/MD-2 blocking agent + LPS group and the anti-TLR4/MD-2+sophoridine+LPS group. Cells and cell culture fluids were collected at 120 min after the each group was processed. The expression of TLR4 protein was measured by western blot， the distribution and expression of NF-κB protein were measured by immunocytochemistry， and the expression of NF-κB and TNF-α mRNA were measured by western blot and reverse transcriptase polymerase chain reaction (RT-PCR). The content of TNF-α in the cell supernatant was detected by using radioimmunoassay. Result： Compared with the LPS group， the expression of TLR4 protein， NF-κB mRNA， the rate of NF-κB entry the nucleus， TNF-α mRNA and TNF-α content in the cell supernatant were significantly decreased in the sophoridine+LPS group (P<0.01). The rate of NF-κB entry the nucleus and TNF-α in the TLR4/MD-2 blocking agent+LPS group and the TLR4/MD-2 blocking agent+sophoridine+LPS group were notablly lower than that of the LPS model group (P<0.01)， close to that of the blank control group. However， there was no statistical significance between the two groups. Conclusion： TLR4/MD-2 may be one of sophoridine′s targets. Sophordine′s inhibitory effect on pathway activity of TLR4-NF-κB-TNF-α may be one of its antiendotoxin mechanisms.
[Key words]sophoridine； TLR4/MD-2 blocking agent； LPS； RAW264.7 macrophage； TLR4
doi：10.4268/cjcmm20122022
[责任编辑张宁宁], 百拇医药(王艳荣,杨峰,刘静,周娅)
[Key words]sophoridine； TLR4/MD-2 blocking agent； LPS； RAW264.7 macrophage； TLR4
doi：10.4268/cjcmm20122022
[责任编辑张宁宁], 百拇医药(王艳荣,杨峰,刘静,周娅)