Study On Radiosensitization Effect Of Shengmai Jianghuang Powder On Human Nasopharyngeal Carcinoma Cell And Its Underlying Mechanism

Research purposes:Through the effect of Shengmai Jianghuang powder on human nasopharyngeal carcinoma CNE-2 nude mice transplanted tumor and human nasopharyngeal carcinoma cell CNE-2,it was determined that Shengmai Jianghuang powder has a sensitizing effect on radiotherapy of nasopharyngeal carcinoma.Through the differences in the expression of CDK6 and CCND1 genes in the tumor xenografts and different subgroups of nude mice in different groups,the related molecular mechanisms of Shengmai Turmeric in sensitization of nasopharyngeal carcinoma were preliminarily explored.Methods:1.Determination of main components in Shengmai Jianghuang powder by high performance liquid chromatography2.Establishment of human nasopharyngeal carcinoma CNE-2 xenograft model in nude miceThe model of CNE-2 xenografts in nude mice was established by cell suspension method.The nude mice were randomly divided into 7 groups:blank control group(saline),irradiation group,low-dose Shengmai Jianghuang powder group(4.29g/kg),high-dose Shengmai Jianghuang powder(12.87g/kg),low-dose Shengmai Jianghuang powder combined irradiation group(4.29g/kg),high-dose Shengmai Jianghuang powder combined irradiation group Irradiation group(12.8g/kg),cisplatin combined irradiation group(0.001g/kg),6 in each group.Detect and record tumor growth.3.RT-qPCR detection of CDK6 and CCND1 related factors in tumor tissues4.Immunoblotting and immunohistochemistry to detect the expression of CDK6 protein in tumor tissues of each group5.MTT assay detects the inhibitory effect of Shengmai Turmeric Powder on the proliferation of human nasopharyngeal carcinoma CNE-2 cells6.Cloning assay to detect the radiosensitizing effect of Shengmai Jianghuang Powder on human nasopharyngeal carcinoma CNE-2 cells7.RT-qPCR detected the expression of CDK6,CDK4 and CCND1 in each group of cellsThe CNE-2 cells were divided into three groups:blank group,irradiation group,and Shengmai Jianghuang powder combined irradiation group.RT-qPCR was used to detect the expression of CDK6,CDK4 and CCND1 in each group of cells.Results:1.According to the HPLC chart,the aqueous extracts of Shengmai Jianghuang powder Formulations contained ginsenosides Rb2 and Rd,respectively,Schizandrol A,Methylophiopogonanone,and Deoxyschizandrin.The herbal extract of Shengmai Turmeric Powder contains schisandrin A,Deoxyschizandrin,and curcumin.2.Human nasopharyngeal carcinoma CNE-2 nude mice models,Compared with blank group,the growth of tumor in each experimental group was inhibited to a certain extent.Compared with simple irradiation group,high-dose Shengmai Jianghuang powder combined irradiation group and cisplatin combined irradiation group had high tumor volume inhibition rate,tumor growth Was significantly inhibited,indicating that Shengmai Jianghuang powder nasopharyngeal carcinoma CNE-2 xenografts with radiosensitization.3.By immunoblotting,immunohistochemistry and in vitro and in vivo RT-qpcr experiments,the results showed that compared with the blank group and the irradiated group,the expression of CDK6 was increased in the Shengmai Jianghuang powder combined irradiation group(P<0.05).In vitro RT-qPCR results showed that CCND1 expression was up-regulated in Shengmai Jianghuang powder combined irradiation group compared with irradiated group(P<0.05).Compared with blank group and irradiation group,the expression of CDK4 inShengmai Jianghuang powder combined irradiation group did not change significantly(P>0.05).The above results showed that Shengmai Jianghuang powder can up-regulate the expression of CDK6 and CCND1 after irradiation,and the effect on CDK4 expression is not obvious.4.The results of MTT assay showed that different concentrations of Shengmai Jianghuang powder scattered CNE-2 cells in a concentration-dependent manner at 24h and 48h.The median inhibitory concentrations(IC50)at 24h and 48h were 10.04 mg/mL and 7.92 mg/mL,respectively.5.Clone formation experiments showed that Shengmai Jianghuang powder decreased the colony formation rate of irradiated CNE-2 cells.It was shown that the intervention of Shengmai Jianghuang powder can significantly improve the radiosensitivity of nasopharyngeal carcinoma cells.Conclusion:Through human nasopharyngeal carcinoma CNE-2 xenograft tumor model and colony formation experiments,it was confirmed that Shengmai Jianghuang powder scattered radiosensitization of nasopharyngeal carcinoma in vitro and in vivo,and its effect may be related to CDK6 and CCND1 genes.