WANG Dexuan,YE Xiaohua,YU Lingfang, et al. The protective effect of WNK3 kinase on apoptosis in HEK293 cell induced by interleukin-1β [J]. JOURNAL OF WEZHOU MEDICAL UNIVERSITY, 2016, 46(1): 1-6.
Abstract：Objective: To observe the effect of high expression of WNK3 on apoptosis in HEK293 cells induced by interleukin-1β. Methods: The HEK293 cells were divided into 3 groups (Vector+NS group,Vector+IL-1β group and WNK3+IL-1β group), then tansfection was performed. Liposome 2000 was added to the 1st and 2nd group with PCMV-HA-Vector as the blank and negative control respectively; WNK3 was added to the 3rd group. After treated with 10 ng/mL IL-1β (the 2nd and 3rd group) or the same amount of normal saline (the 1st group), the CCK8 analysis was performed for the detection of cell activity after incubation for 0 h, 12 h,24 h, 36 h and 48 h at 37 ℃ respectively. Cell proteins were collected after 0 h, 18 h and 36 h incubation respectively for detection of Caspase-3, Cleaved Caspase-3, Caspase-9, Cleaved Caspase-9. In addition, cell protein were collected 0 min, 30 min and 60 min after the treatment of IL-1β for JNK detection. Results: No significant difference of cell activity was observed in the Vector+NS group within 48 h. In the Vector+IL-1β group, cell activity began to decline soon after administration. Significant difference could be found after 24 h and the cell activity reached the lowest at 48 h (P<0.01). Cell activity of WNK3+IL-1β group declined in a moderate manner and the significant difference appeared at 36 h and 48 h when comparing with that at 0 h in the same group (bothP<0.05). However, in comparison with the Vector+IL-1β group at the same time point, cell activity of WNK3+IL-1β group was still a little higher and the significant difference could be observed at 48 h (P<0.01). The expression of Caspase-9 and Caspase-3 began to decline 18 h after the administration with an increasing peak of cleaved Caspase-9 which decreased slightly at 36 h. An obvious cleaved Caspase-3 could also be observed 36 h after the administration. In the WNK3+IL-1β group, the decline of Caspase-3 was more moderate when comparing with that in the Vector+IL-1β group. A small quantity of cleaved Caspase-3 could be detected and showed an statistical significance in comparison with that of the same group at 0 h (P<0.05) with the WNK3; the expression of p-JNK declined but t-JNK did not change. Conclusion: IL-1β may decrease activity of HEK293 cells while transfection of WNK3 may partly reverse this affection of IL-1β. The over expression of WNK3 may inhibit the activation of JNK pathways induced by IL-1β, decrease the activation of Caspase pathway to reduce apoptosis, and finally protect the kidney cells under adverse conditions.
