Objective To explore the effect of methotrexate packaged by tumor derived microparticles (T-MP MTX) combined with radiotherapy on lung cancer stem cell (CSC) in vitro. Methods T-MP MTX was prepared from non-small cell lung cancer A549 cells. Proliferative changes of A549 cells, bronchial epithelial cells H460 and 16HBE cells treated by T-MP MTX were assayed by MTT method. Cell cycles of A549 cells in blank group and T-MP MYX group were examined by fluorescence activated cell sorting (FACS). The effect of T-MP MTX combined with radiotherapy on CSCs was assessed by tumor sphere formation experiment and animal experiment. The expressions of stemness relative genes (such as β-catenin, Nanog, SOX-2 and KLF4) were measured by Western blot. Results T-MP MTX dose-dependently inhibited the cell growth in A549 cells, but didn't in H460 cells and 16HBE cells. The S cycle ratio of A549 cells in blank group and T-MP MYX group measured by FACS were (15.83±3.14)%and (47.47±6.69)%, respectively. S cycle ratio of T-MP MYX group was notably higher compared with that of blank group (t=7.411, P=0.002). Further study revealed that the number of tumor sphere in blank group, control group, 2 Gy group, 4 Gy group and 6 Gy group was (268.9±22.4), (172.4±18.7), (48.3±5.1), (16.3±3.5) and (5.1±3.1), respectively. The number of tumor sphere in other groups was decreased compared with that in blank group (F=228.291, P=0.000). The numbers of tumor sphere in 2 Gy group, 4 Gy group and 6 Gy groups was also reduced compared with that in control group. Importantly, the number of tumor sphere in these groups were decreased dramatically as the dose of radiotherapy increased (F=95.142, P=0.000). The results of tumor sphere volume were similar with the number of tumor sphere. Western blot experiment showed that T-MP MTX treatment in A549 cells decreased the expression of stemness relative genes (β-catenin, Nanog, SOX-2 and KLF4), and its role was reinforced when radiotherapy was combined. Animal experiment implied that activity of luciferase in T-MP MTX group was decreased compared with that in blank group (P=0.000), and the activity of luciferase in T-MP MTX plus 2 Gy group was reduced significantly (t=6.887, P=0.002). Conclusions T-MP MTX has a potential to sensibilize radiotherapy, and it will synergistically inhibit the proliferation of CSCs when combined with radiotherapy. Moreover, its mechanism may be related with T-MP MTX activating CSCs from hypometabolism state and blocking process of cell cycle.

Objective To observe the effect of Shenmai injection combined concurrent chemoradiotherapy on immune function in patients with locally advanced non-small cell lung cancer.Methods A total of 80 patients with locally advanced non-small cell lung cancer were randomly divided into treatment group and control group.In treatment group,40 patients received simultaneous three-dimensional conformal radiotherapy combined with TP(docetaxel+cisplatin)chemotherapy,while given Shenmai injection in the process of concurrent chemoradiotherapy.In control group,40 patients only received chemoradiotherapy.The changes of T cell subsets were observed between the two groups.Results It showed no significant differences in percentages of CD+3,CD+4,CD+16CD+56and CD+4/CD+8 ratio between two groups pre-treatment(all P＞0.05).There were no significant adverse reactions occurred after received Shenmai injection in treatment group.Conclusion It play an important role in improving immune function of patients with locally advanced non-small cell lung cancer who received Shenmai injection combined concurrent chemoradiotherapy.

Objective:To study the soil physical and chemical properties， microorganisms， and metabolites in different culture environments of Gastrodia elata， so as to provide scientific basis for subsequent cultivation of G. elata in multiple environments. Method:The tubersphere soil of G. elata cultured in different environments was collected for analyzing the soil nutrients， microbial numbers， and metabolite differences using the agrochemical method， plate-count method， and gas chromatography-time-of-flight mass spectrometry （GC-TOF-MS）-based non-targeted metabonomic approach. Result:The analysis of soil physical and chemical properties revealed the highest soil moisture， pH， available potassium， and available phosphorus in the spinney and the highest electrical conductivity， total nitrogen， alkali-hydrolyzable nitrogen， and organic matter in the pinewood. As demonstrated by the quantitative analysis of soil microorganisms， the cultivable microorganisms were bacteria， actinomycetes， and fungi， with the bacterial population and total microbial biomass in the spinney and the number of fungi and actinomycetes in the barren slope detected to be the largest. The ratio of bacteria to fungi （B/F value） in the pinewood was the highest， while that in the barren slope was the lowest. The results of metabonomic research demonstrated that the compositions and quantities of soil metabolites in the spinney （Z group）， pinewood （S group）， and barren slope （HD group） varied. Through comparisons between S and Z groups， between HD and Z groups， as well as between HD and S groups by orthogonal partial least squares discriminant analysis （OPLS-DA）， 18， 35， and 24 differential metabolites were separately screened out， and the Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis yielded 5， 9， and 13 metabolic pathways. There existed a significant causal relationship of the soil physical and chemical properties and microbial numbers with the metabolites. Conclusion:The soil physical and chemical properties， microbial numbers， and metabolite changes differed significantly in different culture environments of G. elata， which were sorted by the suitability in a descending order as follows： spinney > pinewood >barren slope. The soil physical and chemical properties and microbial numbers are the crucial factors driving changes in soil metabolites， suggesting that regulating the soil physical and chemical characteristics and microbial characteristics in the culture environment is an important mechanism for maintaining the G. elata-soil-microbial symbiotic system.