Monobutyl Phthalate Induced Pyroptosis of Testicular Interstitial Cells Through Caspase-3/GSDME Pathway
Objective To investigate whether monobutyl phthalate(MBP)induced pyroptosis in testicular stro-mal cells(TM-3 cells)and its possible pyroptosis mechanism.Methods After 24 h of exposure to cells in the logarithmic growth phase with concentrations of 0,2.5,5,and 10 mmol·L-1 MBP,the morphological changes of the cells were observed under an optical microscope.The release rate of lactate dehydrogenase(LDH)was mea-sured by microenzyme.Fluorescence microscopy was used to detect the proportion of PI positive cells.Western blot was used to detect the relative expression levels of apoptosis related proteins Caspase-3 and GSDME.TM-3 cells in the logarithmic growth phase were exposed to the Caspase-3 inhibitor,Ac-DEVD-CHO in a concentra-tion gradient(0,2.5,5,10,20 μmol·L-1)for 24 h.Cell viability was measured by CCK-8 method to determine the toxic dose of Ac-DEVD-CHO.The Ac-DEVD-CHO was applied to the cells to verify the mechanism of MBP-induced pyroptosis in TM-3 cells.Results The level of LDH release rate,proportion of PI positive cells and relative expression levels of Caspase-3 and GSDME protein were increased in each infected group com-pared with the control group(P all<0.05).When Ac-DEVD-CHO concentration was 5.0 μmol·L-1,cell via-bility was 99.17%(P>0.05)and the middle dose add inhibitor group increased than the middle dose group(P<0.01).The intervention concentration of the inhibitor was later determined to be 5.0 μmol·L-1.Moreover,the level of LDH release rate,the proportion of PI positive cells,and the relative expression level of Caspase-3 and GSDME proteins in the middle dose add inhibitor group decreased compared with the middle dose group(P all<0.05).Conclusion MBP can induce cell pyroptosis in interstitial cells of testis by Caspase-3/GSDME pathway.
pyroptosisinterstitial cells of testisCaspase-3GSDME
万方数据
维普
