The Study Of CK2/NF-κB Pathway In The Model Of Inflammatory Cytokines-induced Apoptosis Of Islet Beta Cells

Background:Inflammatory cytokines-induced apoptosis of islet beta cells is an important mechanism for the development of diabetes mellitus.The transcription factor NF-κB plays a critical role in beta cells apoptosis,and the mechanisms are involved that many inflammatory factors,metabolic stress and other factors activate the NF-κB signaling pathway,then it induces the translocation of NF-κBp65,thus results in the apoptosis of islet beta cells.Recently,many studies found that CK2 is closely related to cell apoptosis.But whether the mechanism is related with the NF-κB signaling pathway in islet beta cells is not understand well.Objectives:In this study,inflammatory cytokines(TNF-α + IFN-γ + IL-1β)were used to establish the inflammatory model in islet beta cells.The apoptosis of beta cells,the protein levels of NF-κB signaling pathway,and the function of beta cells were observed.Meanwhile,after down-regulating the expression of CK2 protein,the effects of CK2 on the beta cells apoptosis and function,the activation of NF-κB signaling pathway were detected.Methods:The MIN6 cell lines were treated with different concentrations of inflammatory cytokines for 24 h,48 h and 72 h respectively.The cell viability was detected by CCK.The apoptosis of beta cells was detected by flow cytometry and Hoechst/PI staining.The translocation of NF-κBp65 was detected by immunofluorescence.Western blot was used to detect the protein levels of NF-κB signaling pathway,bax and bcl-2,PDX-1 and Maf A.Insulin levels in GSIS were detected by ELISA.Results:The results indicated that the inflammatory cytokines significantly reduced the beta cell viability,which was concentration-dependent and time-dependent.The protein level of bax was increased,while the protein level of bcl-2 was decreased.In the model of inflammatory cytokines-induced apoptosis of islet beta cells,the p65 was increased in the nucleus.Compared with the control group,the protein levels of IκB and p65 decreased,the protein level of phosphorylated p65 increased,it indicated that IκB was degraded which promoted the translocation of p65 and the activation of the NF-κB signaling pathway.In the model of inflammatory cytokines-induced apoptosis of islet beta cells,the protein levels of PDX-1 and Maf A were significantly decreased.We observed the effects of cytokines on GSIS in beta cells.The results showed that the insulin levels were not significantly different from the control group in the 3.3mmol/L glucose.However,when the beta cells were exposed to the 16.7mmol/L glucose,insulin levels were significantly decreased.It suggested that cytokines injured the function of beta cells.The expression of CK2 protein was down-regulated by si RNA,the apoptosis of beta cells was decreased significantly.The protein level of bax was decreased while the protein level of bcl-2 was increased.The protein level of IκB was higher than the cytokine group.The p65 was increased in the cytoplasm and decreased in the nucleus,which confirmed that the down-regulation of CK2 inhibited the activation of NF-κB signaling pathway.The protein levels of PDX-1 and Maf A were increased,and the decreased GSIS which caused by cytokines was alleviated.These results suggested that the inhibition of CK2 improved the function of beta cells.Conclusions:The inflammatory cytokines induced the apoptosis of islet beta cells.The activation of NF-κB signaling pathway is pivotal in islet beta cell apoptosis.The down-regulation of CK2 improve the apoptosis of islet beta cells by inhibiting the NF-κB signaling pathway activation.