| Backgroud:Over the past decade,the incidence of adverse pregnancy outcomes and infertility caused by genital tract infections is on the rise,and the reproductive health of the society is gradually deteriorating.Chlamydia trachomatis,the most common causative agent in the world,can survive in the epithelial cells and evade the host immune response,often leading to persistent,long-term subclinical infections.Related studies have confirmed that in the female reproductive tract there are a variety of immune cells,the neutrophil-based effector cells migrate to the site of infection caused by the natural immune response play an important role in the invasion of chlamydia;Toll-like receptor-mediated Signaling is a key pathway for the induction of innate immunity and cellular inflammatory responses in chlamydia-infected individuals.Excess cytokines lead to local inflammation and tissue remodeling.Given the fact that there are no vaccines available yet,many antibiotics or combined surgeries are used clinically to treat CT infections.However,the use of antibiotics does not eliminate inflammatory damage to reproductive tract tissue by CT infection.Therefore,to find effective drugs to regulate the cellular immune function and thus control the reproductive injury mediated by chlamydia has important clinical significance and social value.Dexamethasone is an artificially synthesized glucocorticoid that has become more and more clinically useful due to its anti-inflammatory and antifibrotic effects,especially in rheumatoid arthritis and aggressive meningococcal disease of the treatment has a significant effect,and in recent years many clinical literature mentioned it can be used for the treatment of genital tract infections.However,there is no specific experimental study about the effect of dexamethasone in the treatment of genital tract infection.The effects of chlamydial infection on the genital tract have been observed in previous studies in our group.In this dissertation,we focused on infecting rat neutrophils with appropriate concentration of chlamydia to make a model of cell infection,and observing the changes of neutrophil function after dexamethasone administration,so as to study the therapeutic value of dexamethasone in genital tract inflammation of chlamydia;Add TLR2/4 agonist,inhibitor changes after the relevant indicators to study the mechanism of dexamethasone drugs in the regulation of immune response.Objective:To investigate the appropriate concentration of chlamydia infection in making chlamydial infection model and study the effect and mechanism of dexamethasone on neutrophil functionin after chlamydial infection.Materials and methods:The neutrophils of rats were isolated and cultured in six-well plates.After the cells adhered,the cells were infected with different concentrations of chlamydia for a period of time.According to the changes of ROS and MPO levels,the infection model was established(the infection dose of chlamydia was mio = 2,and the infection time was 6h).The model of infection was induced by different doses of dexamethasone(lOng/ml,10?g/ml,1mg/ml)for different times(6h/12h/24h).MPO and NTEs were measured to observe the effect of dexamethasone on neutrophil activity,and then established the model of Dexamethasone drug(10ug/ml and action time 12h).TLR2/4 agonist and inhibitor were added into the model,and then analyzed the molecular mechanism of dexamethasone on neutrophil activity according to the changes of TLR2/4 gene expression and TLR2/4 levels.Results:(1)The amount of ROS production in neutrophils infected with Chlamydia was overall increased and time-dependent,which showed that the amount of ROS gradually decreased with the passage of time,while the concentration dependence was not obvious at the same time point;The MPO secreted by granulocytes increased in a time-dependent manner,but at the same time point,there was no significant concentration-dependent.(2)Compared with NC group,the phagocytosis rate of PMN in DXMS group increased,and all of them were obvious at 6h after drug treatment;NETs formed more clearly after administration,and NETs formed in medium dose group;The number of chlamydia in the culture medium was significantly reduced(P<0.01),but there was no significant difference between the groups.Compared with NC,the level of ROS in the treated group was significantly decreased(P<0.01)and in a concentration-dependent manner.Under the same drug concentration,ROS levels were lowest in the low,medium and high concentration groups at 12 h after treatment;Group MPO decreased significantly(P<0.01),but ELA2 levels in high concentration group at 24 hours significantly decreased(P<0.01).(3)Compared with DM group,there were more filamentous structures in TLR2/4 agonist group,especially in DM + J4(LPS)group.The TLR2/4 agonist Chlamydial load in the inhibitor group decreased slightly;MPO in the TLR2/4 agonist group was significantly lower(P<0.05).(4)Compared with NC group,the gene TLR2/4 of DXMS group was down-regulated within 12h;TLR2 was the highest at 6h;TLR4 was more produced at 6h and 12h.TLR2 gene expression was down-regulated in TLR2 agonist and inhibitor groups compared with DM group(P<0.01).TLR4 gene expression was up-regulated in TLR4 agonist group and the expression level was down-regulated in a time-dependent manner.Compared with the DM group,the TLR2 levels in DM + J2(agonist PGN)group and DM + Y2(inhibitor anti-TLR2)group were significantly decreased;TLR4 levels in all agonist and inhibitor groups DM group Flat.Conclusion:Dexamethasone drug administration of chlamydia-infected neutrophils can reduce the number of chlamydia to a certain extent by increasing the phagocytic capacity of neutrophils and increasing the formation of NETs;by inhibiting the levels of ELA2,MPO and ROS may inhibit the over-activation of neutrophils,and thus may be able to combat the inflammatory response of the body;dexamethasone drugs regulate the activity of chlamydial neutrophil infection mechanism may have TLR4-mediated signaling pathway involved,but with TLR2 Mediated signaling pathways may not overlap. |
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