| Objective To investigate the inhibitory effect of Xiangcao Fuzheng Mixture(XCFZ)on lung cancer and its effects on autophagy and Notch signal expression.Methods 1.Eight of the 70 C57BL/6 mice were randomly selected as the normal control group,and the remaining mice were co-modeled.The mice that met the modeling requirements were selected and randomly divided into six groups,with eight mice in each group,namely,group M,LD,MD,HD,DDP and MD+DDP groups.The rats in the LD group were intragastrically administered with XCFZ 9.23 g·kg-1·d-1,the ones in the MD group were intragastrically administered with XCFZ 18.46 g·kg-1·d-1,the ones in the HD group were intragastrically administered with XCFZ 36.92 g·kg-1·d-1,the ones in the DDP group were intraperitoneally administered with cisplatin 3.21 mg·kg-1·(2d)-1,and the ones in the MD+DDP group were intraperitoneally administered with cisplatin 3.21 mg·kg-1·(2d)-1.XCFZ 18.46 g·kg-1·d-1 was intragastrically administered at the same time,while normal control group and group M were intragastrically administered ddH20 0.2 mL d-1.Dosing was timed for 14 days.The mouse body weight,the tumor weight and the tumor-removed body weight were weighed,and the tumor inhibition rate was calculated.The pathological changes of the tumor tissue were observed with a conventional paraffin section,and autophagosomes in the tumor tissue were observed with a transmission electron microscope.Western Blot was used to detect the expression levels of autophagy-related proteins Beclinl,ATG5,LC3,Bax,BCL-2,JAK2,Phospho-JAK2,STAT3,Phosho-STAT3(Tyr705)and Phosho-Stat 3(Ser727)in the tumor tissue.RT-qPCR was used to detect the mRNA expression levels of Beclinl,ATG5,LC3b,Bax,BCL-2,JAK2,Stat3,Notch1 and Hes1 in the tumor tissue.2.Ultra-performance liquid chromatography-tandem mass spectrometry is used to analyze the chemical composition of XCFZ.Results(1)Compared with group M,the tumor weight was decreased in each administration group(P<0.01).The tumor inhibition rate in group MD+DDP was the highest,followed by group S.Group Z had the highest inhibition rate in group LD,group MD and group HD.Compared with the group M,the body weight of the group MD was increased after tumor removal(P<0.05).(3)Compared with group M,the cell destruction and apoptosis of tumor tissues in each administration group were significantly increased.(4)Autophagosomes were found in tumors of all the administration groups.(5)Western Blot results showed that compared with group M,the expression levels of Beclin1,ATG5,LC3Ⅱ protein and Bax/BCL-2 protein ratio in each administration group were increased(P<0.05,P<0.01),and the protein expressions of Phospho-JAK2,Phospho-STAT3 Tyr705 and Phospho-STAT3 Ser727 were decreased(P<0.05,P<0.01).Compared with the group Z,the expression levels of LC3Ⅱ in the group LD and group HD were decreased(P<0.05).The expression levels of Phospho-JAK2,Phospho-STAT3 Tyr705 and Phospho-STAT3 Ser727 were increased(P<0.05,P<0.01).The protein expressions of Beclinl and ATG5 in the group LD were decreased(P<0.41).The ratio of Bax/BCL-2 was decreased in the group HD(P<0.01).Compared with the MD+DDP group,the expression levels of Beclin1 protein in the MD group and the DDP group were decreased(P<0.05,P<0.01),the ratio of Bax to BCL-2 in the DDP group was decreased(P<0.05,P<0.01),and the expression levels of Phospho-JAK2 and Phospho-STAT3 Tyr705 in the MD group were increased(P<0.05,P<0.01).(6)RT-qPCR results showed that compared with group M,the mRNA expression levels of LC3b and Beclinl and the ratio of Bax/BCL-2 mRNA in each administration group were increased(P<0.05,P<0.01),and the mRNA expression levels of JAK2,STAT3,Notch3 and Hes1 in each administration group were decreased(P<0.05,P<0.01).ATG5 mRNA expression was increased in DM,HD,DDP and MD+DDP groups(P<0.01),while Notchl mRNA expression was decreased in LD,MD,DDP and MD+DDP groups(P<0.05,P<0.01).Jaggedl mRNA expression was also decreased in MD,DDP and MD+DDP groups(P<0.05).Compared with the group MD,the mRNA expression levels of Beclinl,ATG5 and Notch3 as well as the ratio of Bax to BCL-2 were decreased in the groups LD and HD(P<0.01),while the mRNA expression levels of STAT3,JAK2 and Hes1 in the groups LD and HD were increased(P<0.05,P<0.01).The mRNA expression level of LC3B was decreased in the group LD(P<0.01),and the mRNA expression level of Notch1 mRNA the group HD was increased(P<0.05,P<0.01).Compared with the group DDP,the group MD had higher Beclinl mRNA expression(P<0.01),and lower JAK2 and Notch3 mRNA expressions(P<0.01).Compared with the MD+DDP group,the mRNA expression of Beclinl mRNA the DDP group was decreased(P<0.05),while the mRNA expressions of JAK2,Notch1,Notch3 and Hes1 in the MD group were decreased(P<0.01),and Notch1 mRNA expression in the DDP group was increased(P<0.05).(7)A total of 64 compounds were identified by XCFZ,including volatile oils,glycosides,flavonoids,terpenes and miscellaneous compounds.Conclusion 1.XCFZ has inhibitory effect on tumor growth of lung cancer mice,and the medium dose effect is the best.2.XCFZ medium dose can increase the body weight of mice with lung cancer after tumor removal.3.XCFZ may inhibit lung cancer by inhibiting the JAK2/STAT3 pathway,promoting autophagy,and inhibiting Notch signaling pathway,which are three interrelated pathways,to induce apoptosis of lung cancer cells.4.XCFZ contains a variety of chemical components,mainly including volatile oils,glycosides,flavonoids,terpenes and miscellaneous compounds. |
