| Background and Objective:Sepsis is a systemic inflammatory response syndrome caused by infection. It is oneof serious complications in trauma, burns, shock, infection and other severelyclinical situation. Furthermore, it is important response which can induce septicshock and multiple organ dysfunction syndrome (MODS). The immune system canidentify invading microbes and induce fast immune response when bacterial infected.The recognition system consists of a series of receptors on surface of immune cells,named Toll-like receptors, which plays a vital role in signal transduction pathwaysof inflammation. TLRs can sense different PAMPs (pathogen associated molecularpatterns). And for respond to invading microorganisms, it can cause a series ofinflammatory response through MyD88 (myeloid differentiation factor 88), TRIF(TIR-domain containing adapter molecule inducing IFN-beta) and other downstreamjunction protein. Human Toll-like receptor 2 (TLR2) transmits intracellular signal inresponse to BLP (Bacterial lipoprotein), which activates inflammatory response toeliminate the invading pathogens and coordinate systemic defense. However, if theinflammation over-reacts, it will cause excessive organs damage, leading to MODS. Our Previous research indicated that, pretreatment of THP-1 cells with a low-dose BLPinduced BLP tolerance through suppressed TNF-αoverexpression. Besides, theblocking TLR2 antibody decreased the expression of NF-κB and release of TNF-αand IL-6 which was induced by the attacking of BLP to THP-1 cell significantly.They probably provide protective effects on the body. However, it is not onlydifficult to induce tolerance by given low-dose pathogens in advance, but also thehigh expense and not well-developed technology in antibodies preparation, so it willunable to extend and make using in the field of clinic. Paeoniflorin has been reportedto have many pharmacological effects including anti-inflammatory andimmunomodulation. In the present study, we investigated the effects of paeoniflorinon NF-κB and TLR2 signal transduction pathways as well as downstream cytokinesin BLP-induced human THP-1 cells.Methods:1. Human THP-1 cells were stimulated with BLP 1000ng/ml for 1,2,4,6,8,12,24,36,48h. Paeoniflorin at various concentrations (from 10-8 to 10-4 M)incubated with BLP 1000ng/ml stimulated human THP-1 cells for 4hr and 48hr.TNF-αand IL-6 were detected using available ELISA kit.2. Paeoniflorin (10-4M) incubated with BLP (100ng/ml) stimulated THP-1 cells for20 mins. Western blot was used to test the expression of NF-κB in nuclear andTLR2, MyD88 in cytoplasm.3. The experiment was divided into 4 grougs: The black control: neitherpaeoniflorin nor BLP were added into THP-1 cells; THP-1 cells were treated for24h with paeoniflorin 10-4M (the Pae group), with BLP 1000ng/ml (the BLPgroup), or with BLP 1000ng/ml + Pae 10-4M (the Pae+BLP group). The cellswere harvested for quantification of apoptosis by flow cytometry measured as the percentage of Annexin V-FITC positive cells.Results:1. The production of TNF-αand IL-6 in human THP-1 cells stimulated with BLPwere promoted. TNF-αwas in peak at 4hr, IL-6 were significantly increasedafter 24hr, and reached peak at 48hr. Paeoniflorin dose-dependently inhibited thelevels of TNF-αand IL-6 that were increased by BLP stimulation.2. BLP induced the activation of NF-κB in THP-1 cells. This activation waspartially inhibited by pretreatment with paeoniflorin (1×10-4M) (P<0.05). Therewas no significant difference among each group in the expression of TLR2 andMyD88 (P> 0.05).3. BLP could promote the apoptosis of THP-1 cells, 6.1% for control versus.20.9% (P<0.05). However, paeoniflorin, at concentration of 10-4M and theperiod of 24h, did not induce apoptosis in THP-1 cells, 6.1% for control versus.8.6% (P>0.05).Conclusion:1. BLP up-regulates the production of TNF-αand IL-6.2. Paeoniflorin has the capacity to inactivate inflammatory response.3. The anti-inflammatory mechanism of paeoniflorin may inhibit activation of theNF-κB pathway.
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