Protective Effect Of Human Amnion Epithelial Cells On LPS-induced Mouse Acute Lung Injury And Relationship With NF-?Bp65

Background:Acute lung injury(ALI)is a common clinical syndrome,its characteristic is stubborn hypoxemia,progressive dyspnea and non cardiac pulmonary edema,etc.A variety of harmful factors can cause the occurrence of ALI.Now it is reported that excessive inflammatory reaction and anti-inflammatory imbalance play an important role in ALI.The specific pathogenesis has not been fully elucidated.Although the support of clinical treatment has been carried out,there is still a high mortality.Nuclear factor kappa B(NF-?B)is a set of eukaryotic cells transcription factors.It participates in many activities of different kinds of cells and possesses essential biological functions,especially adjusting the body's immune response and inflammatory response.The expression of many inflammatory cytokines are related with NF-?B closely.So it can be said that NF-?B is at a critical link in the development of inflammation.Therefore,it may be possible to inhibit NF-?B to resist inflammation and reduce lung injury.Human amnion epithelial cells(h AECs)are a layer of simple cubic or columnar cells on amniotic membrane and directly contact with the amniotic fluid.h AECs possess characteristics similar to Embryonic stem cells and have stage-specific embryonic antigen-4(SSEA-4),SSEA-3,NANOG and octamer-binding transcription factor-4(OCT-4).hAECs have the ability to differentiate to three layer organization cells.In addition,hAECs never or rarely express human leukocyte antigen ? and ?,but express the immune inhibitory human leucocyte antigen G.Thus,h AECs can resist immune response and show great potential in terms of disease treatment.By establishing ALI model with endotracheal LPS instillation and treating with hAECs,we investigate the effect of h AECs on ALI induced by LPS in mice and its underlying mechanism.Objective:(1)To verify whether the human amnion epithelial cells(h AECs)have therapeutic effect on acute lung injury induced by LPS.(2)To verify whether the human amnion epithelial cells(h AECs)have the influence on NF-?Bp65 that h AECs can reduce its expression,so as to reduce the inflammation of the lung injury.Methods:40 male C57BL/6 mice were randomly divided into normal Control group(Control),hAECs Control group(h AECs),ALI model group(LPS)and h AECs treatment group(LPS+h AECs)(n=10).In ALI model group,mice were subjected to LPS(5 mg/kg)by oral insertion,and 1h later,the PBS(50 ?L)were administered by oral insertion.In hAECs treatment group,mice were subjected to LPS(5 mg/kg)by oral insertion,and 1h later,the h AECs(1×107/ml,50 ?L)were administered by oral insertion.The h AECs Control group and normal Control group were injected with the same amount of h AECs or PBS by oral insertion.24 h after LPS administration,the mice were sacrificed to collect blood from heart and lung tissues.Part of right lung was used to calculate the wet/dry weight ratios.Part of right lung was used to detect the myeloperoxidase activity.Others were used to extract cytoplasmic and nuclear proteins.Pulmonary histological changes in the left lung were evaluated by hematoxylin-eosin stain.Concentrations of interleukin(IL)-10,IL-6,IL-1? and tumor necrosis factor(TNF)-? in sera were measured by enzymelinked immunosorbent assay.The nuclear transcription factor-kappa Bp65 activation in lung tissues was detected by Western blot.Results:Compared with the Control group,the lung wet/dry weight ratios,myeloperoxidase activity and the serum concentrations of tumor necrosis factor(TNF)-?,interleukin(IL)-1? and interleukin(IL)-6 in the LPS-treated group were increased,while the level of interleukin(IL)-10 was decreased.Under light microscopy,the lung tissues from LPS-treated group showed haemorrhagia in stroma,serious pulmonary edema,alveolar wall thickness,alveolus collapse and obvious inflammatory cells infiltration,suggesting that it is successful to establish the ALI model mice.h AECs administration significantly decreased LPS-induced evident lung histopathological changes,lung wet/dry weight ratios,myeloperoxidase activity and the levels of IL-6?IL-1? and TNF-? in serum,and inhibited the activation of NF-?Bp65 in lung tissues.Compared with the Control group,LPS administration also decreased the amount of NF-?Bp65 protein in plasma(P<0.05)and increased its amount in nucleus(P<0.05)while this could be reversed by hAECs administration.The result suggested that h AECs partly inhibited the transfer of NF-?Bp65 to nuclear.Conclusions:(1)h AECs can significantly reduce the LPS induced acute lung injury in mice.(2)The protective effect of hAECs on alleviating acute lung injury may be related to that h AECs inhibited the activity of NF-?Bp65 in the lung tissue,reducing the production of inflammation factor.