Role of TIPE2 in pyroptosis in macrophage of mice: evaluation using siRNA technique / 中华麻醉学杂志

ABSTRACT

Objective To evaluate the role of tumor necrosis factor α-induced protein 8-like-2 ( TIPE2) in pyroptosis in macrophage of mice using small interfering RNA ( siRNA) technique. Methods J774A. 1 macrophages of mice were divided into 4 groups ( n=6 each) using a random number table method: non-specific siRNA (Scr-siRNA) group (S group), Scr-siRNA plus LPS∕ATP group (S+LPS∕ATP group ) , TIPE2-siRNA group ( T group ) and TIPE2-siRNA plus LPS∕ATP group ( T+LPS∕ATP group) . The corresponding siRNA and Lipofectamine2000 transfection reagents were added to each group, and transfection was performed for 24-48 h, and in addition LPS 1. 0 μg∕ml was then added, cells were incubated for 5 h, then ATP 5. 0 mmol∕L was added, and cells were incubated for 1 h in S+LPS∕ATP and T+LPS∕ATP groups. Cells were collected to detect the expression of TIPE2, NOD-like receptor familypyrin domain containing 3 (NLRP3), caspase-1, apoptosis-associated speck-like protein containing a caspase activation and recruitment domain ( ASC) and Gasdermin D ( GSDMD) ( by Western blot) . The superna-tant was collected for determination of lactic dehydrogenase ( LDH) activity and concentrations of interleu-kin-1beta (IL-1β) and IL-18 (by enzyme-linked immunosorbent assay). Results Compared with group S, the expression of TIPE2 was significantly down-regulated, the expression of NLRP3, caspase-1, ASC and GSDMD was up-regulated, and the LDH activity and concentrations of IL-1βand IL-18 in supernatant were increased in group S+LPS∕ATP ( P<0. 05) . Compared with group T, the expression of TIPE2 was sig-nificantly down-regulated, the expression of NLRP3, caspase-1, ASC and GSDMD was up-regulated, and the LDH activity and concentrations of IL-1βand IL-18 in supernatant were increased in group T+LPS∕ATP (P<0. 05). Compared with group S+LPS∕ATP, the expression of TIPE2 was significantly down-regulated, the expression of NLRP3, caspase-1, ASC and GSDMD was up-regulated, and the LDH activity and con-centrations of IL-1βand IL-18 in supernatant were increased in group T+LPS∕ATP ( P<0. 05) . Conclusion Application of siRNA technique once again confirms that TIPE2 can inhibit pyroptosis in macrophages of mice.