The Role Of NLRP3 Inflammasome Activation Induced By Endoplasmic Reticulum Stress In The Excessive Release Of Inflammatory Factors From Microglia Induced By Lanthanum

Objective:In China,rare earth minerals are abundant,and resource reserves,production,sales and applications are ranked first in the world.At present,China is the only country in the world that can supply all rare earth elements.In recent years,rare earth elements（REEs）are increasingly used.In high-tech industry,agriculture,and medical care technology,it is released into soil and water,and then transferred to plants,which has many negative effects on the environment and human health and safety.Among them,the impact of rare earth elements on the nervous system is more prominent.Children are particularly sensitive to neurotoxicity caused by rare earth elements.Due to the stability of lanthanum（La）itself,and can enter the brain tissue to accumulate and produce biological activity,it is firmly used as a representative element to study the damage effect of REEs on the nervous system.Microglia is an efficient dynamic monitoring system in the central nervous system and an important link in mediating a variety of immune responses.The activation of microglia can quickly induce a variety of inflammatory factors,NLRP3inflammasomes and a variety of inflammations.Diseases（including neuroinflammation）are closely related and can regulate the maturation and release of a variety of inflammatory factors.In this study,La Cl₃ was used to treat mouse microglia to establish a corresponding in vitro model to observe whether NLRP3 inflammasome is a key link in the activation of microglia and excessive release of multiple inflammatory factors induced by La exposure,revealing La exposure Whether by inducing endoplasmic reticulum stress,the activation of NLRP3 inflammasomes leads to excessive release of inflammatory factors by microglia,which provides a key target and theoretical basis for the protection and treatment of La neurotoxicity.Methods:1.Determine the dose and time of La treatment of BV2 cells by LDH method;2.Observe the morphological changes of La-treated BV2 cells under a microscope;3.Detect the expression level of IBA1 in BV2 cells after La treatment by immunofluorescence method;4.Detect the changes of the inflammatory factors IL-1β,IL-18 and TNF-αreleased by BV2 cells after La treatment by ELISA;5.Western blot was used to detect the expression levels of NLRP3 inflammasome-related proteins NLRP3,caspase-1 and ASC;6.After treatment with si NLRP3,ELISA was used to detect the release of inflammatory factors IL-1β,IL-18 and TNF-αfrom La BV2 cells The change;7.Western blot was used to detect the expression of TXNIP/NLRP3 inflammasome pathways related to endoplasmic reticulum stress（GRP78,PERK,P-PERK,EIFα,P-EIFα,XBP1,ATF6,TXNIP）;8.Determined by cck-8 method The dosage of endoplasmic reticulum stress inhibitor 4-PBA;9.After adding the endoplasmic reticulum stress inhibitor 4-PBA,Western blot was used to detect the expression levels of NLRP3 inflammasome-related proteins NLRP3,caspase-1 and ASC.Results:Compared with the control group,the cell survival rate of La Cl₃ BV2 cells decreased,With the increase of the La Cl₃ treatment dose,the survival rate of BV2 cells gradually decreased;compared with the control group,the BV2 cells of the La Cl₃ treatment group had filopodia significant increase,the fluorescence intensity of IBA1 protein also increased;the contents of IL-1β,IL-6 and TNF-αin the BV2 cell culture medium of each La Cl₃ treatment group were significantly increased compared with the control group,and the difference was statistically significant;Compared with the control group,the protein expression level of NLRP3 inflammasome-related proteins（NLRP3,ASC,caspase-1）in the BV2 cells of the La Cl₃ treatment group increased;after interfering with the expression of NLRP3 inflammasome,the contents of TNF-α,IL-1βand IL-6 in the BV2 cell culture medium of the La Cl₃ treatment group were significantly reduced compared with the pure La Cl₃ treatment group;compared with the control group,the protein expression levels of GRP78,ATF6,XBP1,TXNIP in the BV2 cells of the La Cl₃ treatment group increased,and the protein expression levels of P-PERK and P-EIF2αwere up-regulated,but the total protein expression levels of PERK and EIF2αremained unchanged.Compared with the100μM La Cl₃ treatment group,the protein expression levels of GRP78 and NLRP3inflammasome-related proteins（NLRP3,ASC,caspase-1）in the BV2 cells of the 100μM La Cl₃+2m M 4-PBA group were significantly decreased.Conclusions:1.La causes activation of microglia and excessive release of inflammatory factors.2.La causes ERS in microglia,leading to abnormal expression of key proteins in the three pathways of microglia ERS to activate NLRP3 inflammasomes,and activate NLRP3 inflammasomes.3.Interference with NLRP3 has an antagonistic effect on the excessive release of inflammatory factors from microglia caused by La.4.The endoplasmic reticulum stress intervention agent 4-PBA has an antagonistic effect on La-induced activation of ERS and NLRP3 inflammasomes in microglia.5.La causes excessive release of inflammatory factors in microglia,which is related to the activation of NLRP3 inflammasomes induced by endoplasmic reticulum stress.