[Effect of two Treatments of Sophoridine on TLR4-JNK Signaling Pathway of RAW264.7 Cells Induced by LPS].

Abstract

Objective: To observe the effect of two treatments of sophoridine( pretreating and premixing) on lipopolysaccharide( LPS)-induced on TLR4,the expression of TLR4 and its downstream signaling molecule such as JNK and c-jun in RAW264. 7 cells were,and to detect its mechanism.

Methods: RAW264. 7 cells were randomly divided into five groups,RAW264. 7 cells control group added absence serum DMEM to incubate cells; LPS group, with added 100 μg/L LPS DMEM to incubate cells; Sophoridine control group added 31. 25 mg/L sophoridine DMEM to incubate cells; sophoridine pretreating group,added 31. 25 mg/L sophoridine DMEM to incubate cells 24 h,then threw away sophoridine and added 100 μg/L LPS DMEM to incubate cells; sophoridine premixing group added the final concentration of 31. 25 mg/L sophoridine and 100 μg/L LPS DMEM to incubate cells 1 h. After above treatments, collected cells at 5,30,60,and 120 min,respectively. The mRNA expressions of TLR4,JNK and c-jun were determined by reverse transcription PCR( RT-PCR),the protein expression of c-jun in RAW264. 7 cells was measured by immunocytochemistry and Western blot.

Results: Compared with RAW264. 7 cells control group, there were no statistical difference on each index in sophoridine control group( P >0. 05),the mRNA expressions of TLR4,JNK,c-jun and c-jun protein expression were significantly increased in LPS group( P < 0. 01 or P < 0. 05),but each index increased were different at different time points; the mRNA expressions of TLR4,JNK,c-jun and c-jun protein expression in RAW264. 7 cells were lower in sophoridine pretreating and premixing group than LPS group, but each index decreased were different at different time points.

Conclusion: Sophoridine induces the effect of antiendotoxin through regulating TLR4-JNK signal transduction pathway, which showing that the effect of different treatments of sophoridine may be multi-link adjustment.