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AIM:To detect the changes of active status of bypass signaling pathways in EML4-ALK positive lung cancer cell line H3122 treated with alectinib, hepatocyte growth factor (HGF), epidermal growth factor (EGF) and transforming growth factor-α (TGF-α), and to explore the potential mechanisms.METHODS:EML4-ALK positive cell line H3122 was treated with increasing concentrations of alectinib or/and induced by HGF, EGF and TGF-α.The cell viability was measured by CCK-8 assay.The cell apoptosis was analyzed by flow cytometry.The protein levels and phosphorylation status of ALK, c-Met and EGFR, and the downstream molecules AKT, ERK, p-AKT and p-ERK were examined by Western blot.RESULTS:The viability of the H3122 cells was inhibited by alectinib in a dose-dependent manner after administrated for 72 h, and the IC50 value was 0.042 μmol/L.The concentration-growth curves of the H3122 cells shifted to the right after induced by HGF, EGF and TGF-α.After treatment with alectinib at 0.05 μmol/L for 48 h, the apoptotic rate of H3122 cells was (20.12±1.36)%, while the apoptotic rates of the cells in the groups of alectinib combined with HGF, EGF or TGF-α were (7.85±1.03)%, (5.60±0.79)% and (4.58±1.00)%, respectively.Those values were remarkably lower than those in alectinib single treatment group (P<0.05).Alectinib inhibited the protein levels of p-ALK and its downstream signaling pathway molecules, while HGF significantly up-regulated the protein levels of p-Met and its downstream p-AKT and p-ERK.Besides, EGF and TGF-α remarkablely up-regulated the protein levels of p-EGFR and its downstream p-AKT and p-ERK.Combined treatment with crizotinib and 17-DMAG successfully inhibited the viability of the H3122 cells even in the presence of the HGF and EGFR ligands, respectively.CONCLUSION:Bypass signaling pathways are activated by HGF, EGF and TGF-α in EML4-ALK positive lung cancer cell line H3122, which may be linked to alectinib resistance.
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AIM: To investigate the role of HGF/c-Met signaling pathway in crizotinib-induced apoptosis of different lung carcinoma cell lines and to analyze its potential regulatory mechanisms .METHODS: EML4-ALK positive cell line H2228, c-Met proliferation cell line H1993 and control cell line A549 were treated with crizotinib at different doses for different time periods .The viability of the cell lines was measured by MTT assay .The apoptosis was analyzed by flow cytometry with PI staining.The protein levels of MET and phosphorylated MET (p-MET) of HGF/c-Met signaling pathway as well as its down-stream key proteins AKT , ERK, p-AKT and p-ERK in the cell lines before and after crizotinib treatment were examined by Western blot .RESULTS:The growth of H1993, H2228 and A549 cell lines was inhibited after crizoti-nib treatment for 72 h in a dose-dependent manner .Apoptotic rates of H1993 cells and H2228 cells were increased with the crizotinib concentration and exposure time .Down-regulation of p-MET, p-AKT and p-ERK at protein levels in H1993 cells and H2228 cells after exposure to crizotinib for 72 h was confirmed by Western blot .No obvious change of the related-pro-teins of HGF/c-Met signaling pathway was found in A 549 cell line.CONCLUSION: HGF/c-Met signaling pathway may contribute to crizotinib-induced apoptosis of H1993 cells and H2228 cells, which provides the experimental basis for MET-targeting treatment of lung cancer .
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Objective To investigate the effect and safety of bronchial balloon positioning combined with autologous blood bronchial occlusion in the treatment of intractable pneumothorax.Methods 36 cases of hospitalized patients with refractory spontaneous were collected as the research object,based on the bronchial balloon positioned upstream bronchoscopy combined with autologous blood bronchial occlusion therapy.The efficacy and adverse reac-tions were followed up for one year.Results In the 36 cases of spontaneous pneumothorax in patients with refractory, 30 patients were successfully positioning balloon (83.33%),6 cases of patients who were not successfully positio-ning,were continued with conservative treatment of indwelling thoracic drainage.In 30 patients underwent successful positioning,autologous blood bronchial occlusion were used,and a total of 24 cases (80.0%)patients successfully reduced bubbles escape;6 patients relapsed,who were continued to retainthoracic drainage conservative treatment. Conclusion Bronchial balloon positioning combined with autologous blood bronchial occlusion in the treatment of intractable pneumothorax has accurate positioning and high blocking success rate,and also is safe and effective.It is worth promoting in clinical practice.
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AIM:To investigate the mammalian target of rapamycin ( mTOR) signaling pathway as the center playing a role in the crizotinib-induced apoptosis of non-small cell lung cancer (NSCLC) cell line H2228, which represents positive echinoderm microtubule-associated protein-like 4 (EML4)-anaplastic lymphoma kinase (ALK) fusion gene. METHODS:H2228 cells were processed according to different purposes .Fluorescence quantitative PCR is used to ob-serve the gene states .MTT assay is used to detect the cell inhibition rates .The cell apoptosis and cell cycle were analyzed by flow cytometry .The expression and activation levels of the key proteins in the mTOR signaling pathway were determined by Western blotting .RESULTS:Crizotinib promoted the apoptosis of H 2228 cells in a time-and dose-dependent manner . Crizotinib blocked the H2228 cells staying at the G1 phase.In apoptotic H2228 cells processed with crizotinib, the activa-tion level of mTOR was decreased , and the activation levels of the key proteins in upstream and downstream of mTOR path -way were both declined .The expression level of the fusion protein EML 4-ALK variant 3 was not affected , but its active form of p-ALK was significantly suppressed .CONCLUSION:mTOR signaling pathway has a certain relationship with the crizotinib-induced apoptosis of lung cancer cell H 2228, which represents positive EML4-ALK fusion gene.
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AIM: To study the relationship between MIF gene -173 locus polymorphism, helicobacter pylori (Hp) infection, and gastric cancer in high prevalent (Shanxi) and low prevalent (Guangdong) regions in China. METHODS: Genomic DNA was extracted from peripheral blood of 104 healthy controls, 104 gastric patients from Guangdong and 102 healthy volunteers, and 102 gastric cancer patients from Shanxi. Polymorphism of MIF-173 locus was analyzed by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP). RESULTS: In high prevalence region, the number of patients who carrying with MIF -173 C/C is much higher than those of healthy controls (28.8% vs 15.4%, ?~2=5.47, P0.05). CONCLUSION: The C genotype of MIF -173 locus may be associated with the risk of gastric cancer in China.
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AIM: To study the relationship among peroxisome proliferator-activated receptor-gamma2(PPAR ?2) gene Pro12Ala polymorphism,Helicobacter pylori(H.pylori) infection,and gastric cancer in China.METHODS: 104 consecutive patients with gastric cancer and 104 age-and sex-matched controls from Guangdong Province of southern China were examined.PPAR?2 Pro12Ala polymorphism was analyzed by PCR-restriction fragment length polymorphism method(PCR-RFLP).H.pylori status of subjects was determined by enzyme-linked immunosorbent assay(ELISA) for anti-H.pylori IgG.RESULTS: The prevalence of H.pylori infection was significantly higher in gastric cancer patients than that in control(81.7% vs 59.6%,2=12.27,P