Expression Of NLRP3 In Human Glioma And Its Effects On Glioma Cell Biological Characteristics

Background and objectiveGlioma is the most common neurological malignant tumor in adults.The prognosis of glioma patients is extremely poor and the pathogenesis of glioma remains to be elucidated.NLRP3 is reported to be closely related to the development and progression of many kinds of cancer.However,the conclusions about the function and mechanism of NLRP3 in different reports are controversial.The expression and mechanism of NLRP3 in human glioma have not been reported yet In order to explore the role of NLRP3 in glioma,we detected the expression level of NLRP3 in human glioma tissues and its effects on proliferation,apoptosis,migration and invasion of glioma cell,as well as their possible molecular mechanisms.Methods1.Detecting the expression of NLRP3,ASC,caspase-1 and IL-1?in human glioma tissues.The expression level of NLRP3,ASC,caspase-1 and IL-1? in 39 cases of glioma tissues were detected by immunohistochemistry.The data was analyzed by Spearman rank correlation analysis,taking P<0.05 as statistically significant.2.Exploring the effects of NLRP3 on human glioma cells and its possible mechanism.The si-NLRP3 and NLRP3-overexpressing plasmid vector were transfected into SHG44 and A172 glioma cellsrespectively to knock down or overexpressing NLRP3.The efficiencyof the si-NLRP3 and the NLRP3-overexpressing plasmid vector was confirmed by Western Blot and the changes of cell proliferation,apoptosis,migration and invasion abilities of transfected glioma cell lines SHG44 and A172 were detected by colony formation assay,CCK-8 assay,EdU cell proliferation assay,flow cytometry,wound healing assay,transwell cell migration assay and transwell cell invasion assay.Afterward,the protein expression levels of ASC and caspase-1,IL-1?and EMT-related molecular markers including E-cadherin,ZO-1,Claudin-1,N-Cadherin,vimetin and transcription factor Snail were detected by Western Blot to explore the effects of NLRP3 on human glioma cells and its possible mechanism.Results1.The expression levels of NLRP3,ASC,caspase-1 and IL-1? were positively correlated with the pathological grades of glioma.The expression levels of NLRP3,ASC,caspase-1 and IL-1?detected by immunohistochemistrywere up-regulated in the high grade glioma tissues compared with the low grade glioma tissues,and the data was tested by Spearman rank correlation analysis.There was a significantly positive correlation between the protein expression levelsof NLRP3,ASC,caspase-1,IL-1?and the WHO pathological grades of glioma tissues.2.NLRP3 promotescell proliferation,inhibites apoptosis and promotes migration and invasionon glioma cells.(1)The specific si-NLRP3 and NLRP3-expressingplasmid vector were constructed and transfected into glioma cell lines SHG44 and A172 respectively.The efficiency has been confirmed by detecting the level of NLRP3 protein by Western Blot.(2)NLRP3 knock-down promoted cell apoptosis and inhibited the proliferation,migration and invasion of glioma cells.Overexpression of NLRP3 showed opposite effects,inhibiting cell apoptosis and promoting cell proliferation,migration and invasion.(3)After knockdown of NLRP3 in glioma cells,the protein expression levels of ASC,pro-caspase-1 and its activated cleaved subunit P20,pro-IL-1? and its cleaved mature form IL-1? were all down-regulated,along with increasingE-Cadherin,ZO-1,Claudin-1and decreasingvimetin,N-Cadherinand snail,while overexpression of NLRP3 could upregulate ASC,pro-caspase-1 and P20,pro-IL-1? and mature IL-1?,downregulate E-cadherin,ZO-1,Claudin-1 and upregulateN-cadherin,vimetin and snail.Conclusions1.The protein expression levels of NLRP3,ASC,caspase-1 and IL-1? are significantlypositive correlated with the pathological grades of glioma.2.NLRP3 inhibits glioma cell apoptosis and promotes proliferation,migration and invasion.3.The role of NLRP3 in glioma may be related to the changes of NLRP3 inflammasome activity and the effect of NLRP3on EMT in glioma cells.