AKR1B10 Confers Resistance To Radiotherapy Via FFA/TLR4/NF-?B Signaling Pathway In Nasopharyngeal Carcinom

Background:Nasopharyngeal carcinoma(NPC)is a common malignant tumor of the head and neck.Radiotherapy is the main treatment for NPC.However,in some cases,radiotherapy resulted to radiotherapy resistance,It it led to tumor recurrence and metastasis,and reduced patient quality of life.The molecular mechanism of radioresistance in NPC is needed to be elucidated.Objective:To study the relationship between the expression of AKR1B10 and the treatment effect of NPC patients,the survival,apoptosis and DNA damage repair of NPC cells,and to explore the effect and mechanism of AKR1B10 expression on radioresistance of NPC.Methods:The paraffin tissues of 58 NPC patients ever received radiotherapy,including 30 patients with radiosensitivity and 28 patients with radioresistance.Immunohistochemical experiments were used to detect the expression of AKR1B10 and the correlation beteeen the expression of AKR1B10 and clinical characteristics was analised.AKR1B10 overexpressed and knocked down cell lines were constructed by lentivirus system.The effects of AKR1B10 on cell survival and apoptosis were detected by clonogenic assays and Annexin V-APC / 7-AAD double staining experiments mediated flow cytometry.The expression of apoptotic proteins Caspase3 and PARP were detected by Western blot.Cellular immunofluorescence assay and neutral comet assay were used to detecte the degree of DNA double-strand breaks(DSB),and the cell cycle protein CHK1/2 expression was checked by Western blot.The changes of I?B?,NF-?B p65 and the expression of NF-?B p65 were also detected in overexpression cells and control cells.The quantity of FFA in overexpression cells,control cells and knocked down cells were detected by ELISA.Nuclear and plasma proteins were extracted,and the expression of MyD88,and NF-?B p65 were detected by Western blot.The levels of TLR4,MyD88,NF-?B p65 and p-NF-?B were detected in AKR1B10 expressed cells after treated by FFA alone or combined with TLR4 inhibitor TAK-242.Furthermore,the Xenugraft tumor of nude mice was used to test the radioresponse mediated by AKR1B10.Results:Compared to that in radiosensitivity patients,the expression of AKR1B10 was relatively higher in patients with radioresistance,and the AKR1B10 expression was correlated to tumor invasion,clinical grade and tumor differentiation.AKR1B10 overexpression increased cell survival,decreased cell apoptosis,and reduced DNA damage,accelerated DNA damage repair,and affected the cell cycle related protein CHK1 expression.The results showed that the expression of p-I?B? and p-NF-?B p65 in AKR1B10 overexpressed cells was increased,and decreased after AKR1B10 knocked down.Furthermore,the FFA amount in AKR1B10 overexpressed cells was higher than that in control cells,and the FFA amount was reduced after AKR1B10 was knocked down.Meanwhile,the level of NF-?B p65 protein was higher in the nucleus of AKR1B10 overexpression cell lines than that of control cells.Compared without FFA addition,after treated by FFA,higher expression of TLR4,MyD88,and pNF-?B p65 was detected in AKR1B10 overexpression cells.After TLR4 inhibitor TAK-242 treatment,the expression levels of these proteins decreased,and the levels of MyD88 and p-NF-?B p65 showed a certain increase after FFA stimulation.The tumor Xenograft experiments showed that the tumor growth of the AKR1B10 overexpression cells and radiotherapy groups were much faster than that of control cells and radiotherapy groups,and the ?H2AX expression in the AKR1B10 overexpressed cells and radiotherapy groups higher than that of control cells and radiotherapy as well.These data showed that AKR1B10 promoted NPC radioresistance in vivo and in vitro.Conclusion: 1.Compared to that in radiosensitive patients,the expression of AKR1B10 was higher in radioresistance patients of NPC.2.AKR1B10 increased NPC cell survival,reduced cell apoptosis,promoted DNA repair after radiotherapy,and participated in DNA damage repair by regulating cyclin protein CHK1.3.AKR1B10 participated in radioresistance of NPC by promoting FFA synthesis and activating TLR4 / NF-?B signaling pathway.