Objective To investigate the effects of alantolactone on cell proliferation,cell-cycle and cell cycle-related proteins in human chronic myelogenous leukemia drug-resistant cell line K562/ADR.Methods K562/ADR cells were treated with 0,1.0,2.0,4.0,6.0,8.0,and 10.0 μmol/L of alantolactone for 12,24 and 48 h,with its cell viability analyzed by MTT assay.Flow cytometry was used to examine the effect of alantolactone on the cell-cycle of K562/ADR cells.The cell cycle-related proteins were analyzed by using Western blot after treatment with alantolactone.Results The results of MTT showed that alantolactone effectively inhibited the proliferation of K562/ADR cells in dose and time-dependent way,and the IC50 value of alantolactone in K562/ADR cells was about 5 μmol/L.Flow cytometric analysis displayed that alantolactone could arrest cell cycle at G2/M phase.The percentage of accumulated cells in the G2/M phase was increased from (15.8±1.7) ％ in the control group to (21.0±2.4) ％,(26.4±2.7) ％,and (30.1±3.9) ％ in cells treated with 2.5,5.0,and 7.5 μmol/L of alantolactone for 24 h,respectively (P ＜ 0.05).Alantolactone significantly decreased the expression of CDK1 and CyclinB1 and increased the expression of cyclin-dependent kinase inhibitor p21.Meanwhile,the treatment of K562/ADR with alantolactone led to a dose-dependent decrease in bcr-abl protein levels.Conclusion Alantolactone can significantly inhibit the proliferation and cell-cycle arrest in G2/M phase of K562/ADR cells,in which mechanism may be associated with the regulation of cell cycle-related proteins and downregulation of bcr-abl protein.

Objective:To study the expression of Toll like receptor 3 (TLR3) in human adenocarcinoma of the lung cells induced by respiratory syncytial virus (RSV) and its significance in the diagnosis of pneumonia in children.Methods:A549 cells were divided into RSV infection group [added 1 μg/ml Lipopolysaccharide (TLR3 agonist) transfected RSV virus after 150 μl intervention], Lipopolysaccharide stimulation group (added 1 μg/ml Lipopolysaccharide 150 μl intervention) and normal control group (normal culture). The mRNA expressions of tumor necrosis factor-α, interleukin 8, TLR3 protein and TLR3 in A549 Cells of different groups were compared. We prospectively selected 80 children with RSV infectious pneumonia admitted to Baoding Second Central Hospital from August 2019 to October 2021 as the RSV pneumonia group, and sixty children with common pneumonia were taken as the common pneumonia group, and 60 healthy children in our hospital were taken as the control group. The mRNA expression of serum TLR3 in different groups was compared, and the diagnostic efficacy of serum TLR3 in RSV pneumonia was evaluated by receiver operating characteristic.Results:There was a statistically significant difference in the expression of TLR3 protein among different groups of A549 cells ( P<0.001). The expression differences of TLR3 mRNA in different groups of A549 cells at different time points were statistically significant(all P<0.001). There was significant difference in the expression of tumor necrosis factor-α and interleukin 8 of A549 cells at different time points in different groups (all P<0.05). There was a statistically significant difference in the expression of serum TLR3 mRNA among the three groups of subjects ( F=155.237, P<0.001). The critical value for TLR3 gene diagnosis was 66.87, with corresponding sensitivity of 73.75%, specificity of 70.83%, and the area under curve (AUC) of 0.803(95% CI: 0.753-0.855). Conclusions:Respiratory syncytial virus induces human lung cancer cells and promotes disease progression through TLR3 expression; Serum TLR3 can be used for the diagnosis of RSV pneumonia.

OBJECTIVETo explore the inhibitory effect of alantolactone on the proliferation of adriamycin-resistant human chronic myelogenous leukemia cell line K562/ADR cells and its mechanism.

METHODSK562/ADR cells were treated with various concentrations of alantolactone (0, 1, 2, 4, 6, 8, and 10 μmol/L) for different time points. Cell viability was analyzed with MTT assay. The effect of alantolactone on the apoptosis of K562/ADR cells was measured by flow cytometry. The expression of apoptosis-related proteins after treatment with alantolactone was analyzed using Western blot.

RESULTSAlantolactone could effectively inhibit the proliferation of K562/ADR cells in dose- and time- dependent manner, the IC50 value of alantolactone treatment of K562/ADR cells for 24 h was 4.7 μmol/L (P<0.05). Flow cytometric analysis displayed that the apoptotic rates were 1.35%, 16.91%, 29.61% and 46.26%, respectively, after treatment with alantolactone at 0, 2.5, 5 and 7.5 μmol/L. Meanwhile, the expression of Bcl-2 and BCR-ABL proteins were significantly decreased and that of Bax, cytochrome C, cleaved-caspase-9, cleaved-caspase-3 and cleaved-PARP increased by alantolactone treatment.

CONCLUSIONAlantolactone had obvious inhibitory effect on the proliferation of K562/ADR cells through the caspase dependent mitochondrial(or intrinsic)apoptotic pathway.

Apoptosis ; Caspase 3 ; metabolism ; Caspase 9 ; metabolism ; Cell Proliferation ; drug effects ; Fusion Proteins, bcr-abl ; metabolism ; Humans ; K562 Cells ; Lactones ; pharmacology ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Sesquiterpenes, Eudesmane ; pharmacology ; bcl-2-Associated X Protein ; metabolism

Objective: To study the diagnostic value of a multi-gene molecular testing in cytologically indeterminate thyroid nodules. Methods: From February 2018 to September 2018, patients with thyroid nodules who underwent fine needle aspiration(FNA) at Peking University Cancer Hospital were enrolled. Three hundred and sixty patients were included, consisting of 86 men and 274 women, with a mean age of 45.8 years (between 13 and 89 years old). Among 391 nodules, 141 were cytologically inderminate and 75 were resected. FNA samples underwent prospective testing using a next-generation sequencing (NGS) assay, which included 16 genes for point mutations and 26 types of gene fusions. The testing results of indeterminate nodules were compared with surgical outcomes, to determine the diagnostic performance. The results were compared with the BRAF V600E single gene mutation analysis by χ² test. Results: The multi-gene testing showed a sensitivity of 73.2%, specificity of 96.8%, positive predictive value of 96.8%, and negative predictive value of 73.2%. The diagnostic accuracy of multi-gene testing was significantly higher than the BRAF V600E mutation test (83.3% vs 73.6%, χ²=31.588, P<0.01). Conclusion Multi-gene testing in FNA samples is an effective method to diagnose cytologically indeterminate thyroid nodules, which has a higher accuracy than BRAF V600E mutation detection.