Effects Of High Mobility Group Box Protein B1 On The Growth And Radiosensitivity Of Esophageal Squamous Cell Carcinoma And Its Mechanism

Purpose:In this study,the effect of HMGB1 on growth,apoptosis,DNA damage repair,radiotherapeutic efficacy,and its underlying mechanism of radiosensitization were investigated in human esophageal squamous cell carcinoma,in vitro and in vivo.Methods:Firstly,we performed survival analysis and clinical stage analysis on the transcript sequencing data of HMGB1 and LC3B which was extracted from TCGA database.Then,the IHC scores of cancer and adjacent normal tissue obtained from ESCC tissue microarray was analyzed.Thirdly,we constructed stable HMGB1 knockdown ESCC cell lines that were then confirmed using qRT-PCR and Western Blot.In vitro,the cell viability was detected by CCK-8 and EdU assays;the regulation and translocation of HMGB1 after irradiation were tested by Western Blot and ELISA;clonogenic survival data was obtained to assess effects of treatment on radiosensitization;Changes of cell apoptosis was analyzed by flow cytometry assay;immunofluorescence assay were used to assess ability of DNA damage repair;the regulation of autophagy and apoptosis by HMGB1 was identified by Western Blot.To determine in vivo radiotherapeutic efficacy,tumor growth delay was analyzed in a ESCC tumor-bearing xenograft mouse model after treatment of HMGB1_shRNA and/or irradiation.Finally,in vivo and vitro,we studied the radiosensitization effect of Silibinin on ESCC through CCK-8,clonogenic survival assay,immunofluorescence assay,Western Blot and other methods.Results:At first,TCGA database analysis showed that HMGB1 were related to tumor progression and prognosis,and the regulation of autophagy by HMGB1 may explain the results.Secondly,report from ESCC tissue microarrays showed that the expression of HMGB1 in esophageal cancer tissue is significantly higher than that in adjacent normal tissues,providing a basis for follow-up studies.Subsequently,we found that downregulation of HMGB1 could effectively inhibit the proliferation of ESCC cells;increase the radiosensitivity of ESCC cells;impair the DNA damage repair ability of ESCC cells followed irradiation,reduce the level of autophagy and increase the apoptosis ratio of ESCC cells followed irradiation.At last,we found Silibinin could increase the radiosensitivity of ESCC cells,and this radiosensitizing effect was associated with the inhibition of HMGB1.Silibinin is a promised radiosensitizor.Conclusion:HMGB1 is expected to be a potential target for ESCC therapy and radiosensitization.