Investigation On The Antibacterial Effect And Mechanism Results From Platelets

【Background】The antibiotics and immunosuppressive agents are widely used,which make drug-resistant bacterial strains increase rapidly.Recent years,conditional pathogenic bacteria in hospital have gradually turned into drug-resistant bacteria,causing the cure rate reduced and mortality increased.The most common drug-resistant pathogens to nosocomial infections in hospital are methicillin-resistant coagulase-negative Staphylococcus(MRCNS),vancomycin-resistant Enterococcus faecalis(VRE),multidrug-resistant Pseudomonas aeruginosa(MDR-PA),extended-spectrum β-lactamase(ESBLs)Escherichia coli strains and so on.We have not given enough attention to the infection by conditional pathogenic bacteria.Therefore,once transformed into drug-resistant bacteria,they would spread rapidly into different patients and environments.Therefore,it is urgent for us to find new efficient and safe ways to fight against bacterial infection.Platelets,as important blood cells,not only play the role in the hemostasis,but also take part in immune response.Platelets participate in the antibacterial action by modulating the innate immunity and acquired immunity.Compared with traditional antibiotics,platelets have a broad spectrum of inhibitory spectrum,non-toxic side effects,and may not cause drug-resistance.For the clinical treatment of bacterial infection,platelets may provide a new strategy.However,the platelet inhibition mechanism had been little kown,which is related with the immune system.So we need to invest more energy to carry out the research.In this study,Staphylococcus epidermidis(SE)and Enterococcus faecalis(EF)were used to study the direct antibacterial effect of platelets and explore the antibacterial mechanism.Our further study were focused on that platelet inhibited methicillin-resistant Staphylococcus epidermidis(MRSE)and got the best time of antibacterial effect.The current study might provide new ideas and experimental basis for the clinical treatment of drug-resistant infections.【Methods and results】 1.The antibacterial effect on SE and EF in vitro by plateletsCompared with the negative and plasma antibacterial group,platelet inhibition might also be a bright spot,by plate counting method and growth curve in vitro.The results showed that platelets could start to inhibit the growth of SE after 3 h,and the antibacterial effect was most obvious after 10 h,and the antibacterial effect was maintained from 10 h to 18 h.The effect on EF was led by platelet happened slowly,compared with that of SE.After 12 h,the antibacterial effect is most obvious.In addition,the onset of platelet inhibition happened later than plasma did,but it could be last longer.2.The antibacterial mechanism on SE and EF by plateletsThe changes of bacterial glycometabolism indexes,GLU,LA,LDH and pH,were detected by biochemical methods.Microstructure changes of SE and EF were observed by transmission electron microscopy.The proliferation of bacteria was detected by flow cytometry.The expressions of AZR,G6 PD and RecA-LexA genes were detected by qRT-PCR.We checked metabolism compensation test,whether Glucose-δ-lactone(GDL)and hypoxanthine(IMP)interfered the antibacterial effect by platelet in PPP or not.The results showed that p H value and LDH level of the bacteria had changed.GLU and LA level had no difference.The results of TEM showed,in the platelet group,bacterial DNA agglutination blocked.The low density of electrons for bacteria expansion and proliferation of bacteria were decreased.It was verified that platelets slowed down the division and multiplication of bacteria.Furthermore,the results of qRT-PCR showed that both AZR and G6 PD mRNA level were down-regulated.RecA-LexA genes were up-regulated,confirming platelet-disrupting bacterial DNA.Finally,when the GDL and IMP were added,platelet inhibition on bacterial growth was weakened.It suggested that platelet inhibition might interfere with bacterial metabolism.3.The antibacterial effect on MRSE in vitro by plateletsThe methicillin-resistant Staphylococcus epidermidis(MRSE)was identified by PCR and DNA sequencing.The MRSE curve of platelets in vitro was drawn by plate counting method.The results of PCR and sequencing showed that mecA-resistant gene was identified in this study.Platelets could effectively inhibit the growth of MRSE in vitro.The PLT inhibition curve of MRSE showed that the inhibition time point of PLT was ahead than that of SE and EF.The antibacterial effect of MRSE led by platelet started after 1 h,and achieved stable state from 6 h to10 h,and the antibacterial effect of platelet was gradually better than that of plasma after 12 h.【Conclusion】In summary,the present project confirmed that platelets could directly inhibit the growth of SE,EF and MRSE.In vitro,we studied the antibacterial effects of SE,EF and MRSE by plasma-poor platelet,and initially explored the antibacterial mechanisms on bacterial glucose metabolism and bacterial microstructure changes.The results showed that platelets might interfer the bacterial glucose metabolism and destroy bacterial chromatin to inhibit the proliferation of bacteria.This study laid the foundation for the clinical study of new antibacterial methods.