| Acrolein can be exposed to human living environment through endogenous sources,dietary sources and environmental sources,and induce cells to produce chronic inflammatory diseases such as inflammation.For patients who already have inflammation,it will have an adverse effect on their condition.Some studies have shown that acrolein can activate NLRP3 inflammatory bodies to induce cell scorch death,but there is no evidence to indicate whether acrolein can aggravate lipopolysaccharide(LPS)-induced inflammation by activating NLRP3 inflammatory bodies.In addition,it has been reported that the activation of HMGB1/TLR4/MYD88/NF-κB pathway plays an important role in inflammation,but the regulatory role of acrolein in this signal pathway has not been studied.Therefore,this paper first explores the effect of acrolein on LPS-induced inflammation and its mechanism,and then we also explore the protective effect and mechanism of apigenin(apigenin,API)and apigenin derivative(apigenin-7,4’-Odioctanoate,API-C8)on acrolein-induced inflammation induced by LPS.The specific research methods and results are as follows:1.The inflammatory model of human umbilical vein endothelial cell(HUVEC)was stably constructed by LPS,and the effect of acrolein on the inflammatory model was studied.The gene expression changes of classical pro-inflammatory factors(IL-1β,IL-6and TNF-α)were detected by real-time fluorescence quantitative PCR,and the protein expression changes of proinflammatory proteases(i NOS,COX-2)were detected by Western blot.In order to detect whether the inflammation of LPS induced by acrolein is related to oxidative stress,reactive oxygen species(ROS)and glutathione peroxidase(GSH-PX)kits were used to detect the changes of oxidative stress indexes,and reactive oxygen species inhibitor N-acetyl-l-cysteine(NAC)was added to further study the effects of reactive oxygen species on oxidative stress and inflammation.The results showed that acrolein-treated could promote the expression of IL-1 β,IL-6,TNF-α,i NOS and COX-2compare with LPS group,indicating that acrolein could exacerbate LPS-induced HUVEC cell inflammation.It was further found that acrolein increased the content of ROS and decreased the activity of GSH-PX in LPS.NAC alleviates the effect of acrolein on the above oxidative stress indexes,indicating that acrolein aggravates the inflammation of HUVEC induced by LPS by promoting oxidative stress in cells.2.ROS can activate NLRP3 inflammatory bodies and induce inflammation,some studies have shown that the activation of HMGB1/TLR4/MYD88/NF-κB signal pathway plays an important role in inflammation.LPS-induced HUVEC was used as an inflammatory model to study the specific mechanism of acrolein aggravating inflammation.The expression of NLRP3 inflammasome complexes(NLRP3,ASC and cleavage-caspase-1)and the downstream protein signals(IL-18 and IL-1β)were detected by Western blot.The effect of ROS on the activation of NLRP3 inflammasome was determined by adding NAC.Then Western blot was used to detect the expression of HMGB1,TLR4,MYD88 and P-P65.NAC and ST2825,a specific inhibitor of MYD88,were added to explore the role of HMGB1/TLR4/MYD88/NF-κB signal pathway in this experiment.The results showed that the addition of acrolein could significantly promote the expression of NLRP3,ASC and cleavage-caspase-1 protein,and NAC could significantly inhibit the expression of NLRP3 inflammasome promoted by acrolein,indicating that acrolein could aggravate the inflammatory response by promoting the expression of NLRP3 inflammasome.By studying the expression of HMGB1,TLR4,MYD88 and P-P65,it was found that acrolein significantly increased the expression of HMGB1,TLR4,MYD88 and P-P65,while NAC significantly alleviated the expression of HMGB1,TLR4,MYD88 and P-P65,indicating that ROS plays an important role in activating HMGB1/TLR4/MYD88/NF-κB signal pathway,while the addition of ST2825 inhibits the expression of MYD88/NF-κB signal pathway.It is further suggested that acrolein can aggravate LPS-induced HUVEC inflammation by activating NLRP3 inflammasome and HMGB1/TLR4/MYD88/NF-κB signal pathway.3.Apigenin(apigenin,API)has anti-inflammatory and antioxidant activities,but apigenin derivatives(apigenin-7,4’-O-dioctanoate,API-C8)have not been reported.The inflammation model was established by LPS to study the effect and mechanism of API and API-C8 in relieving the inflammation aggravated by acrolein.The experimental method of CCK-8 was used to screen and determine that the best concentration of the two polyphenols was 10 μM.q PCR and Western blot were used to explore whether API and API-C8 could alleviate acrolein-induced LPS inflammation;flow cytometry was used to detect the effects of API and API-C8 on ROS induced by acrolein-induced LPS-induced inflammation;Western blot was used to detect the effects of API and API-C8 on NLRP3 inflammatory bodies and HMGB1/MYD88 signal pathway.The results showed that API and API-C8 could inhibit the expression of IL-1β,IL-6,TNF-α,COX-2 and i NOS,indicating that API and API-C8 could alleviate the inflammation induced by acrolein.At the same time,it was speculated that API-C8 could improve the lipophilicity of derivatives by introducing fatty acids,which could pass through the cell membrane more easily and showed stronger antioxidant and anti-inflammatory ability than API.The changes of ROS content showed that API and API-C8 could alleviate acrolein-induced inflammation by inhibiting the expression of oxidative stress;in addition,API and APIC8 could significantly inhibit the expression of NLRP3 and the activation of HMGB1/MYD88 signal pathway,indicating that API and API-C8 could inhibit the activation of NLRP3 inflammasome and HMGB1/MYD88 signal pathway in HUVECs.In summary,we conclude that acrolein can aggravate LPS-induced HUVEC inflammation by activating NLRP3 inflammasome and HMGB1/MYD88/NF-κB signaling pathways,while API and API-C8 can alleviate inflammation through these pathways.Our research lays the foundation for further food research and safety control. |
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