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Polygonum cuspidatum polyketide synthase 1 (PcPKS1) has the catalytic activity of chalcone synthase (CHS) and benzylidene acetone synthase (BAS), which can catalyze the production of polyketides naringenin chalcone and benzylidene acetone, and then catalyze the synthesis of flavonoids or benzylidene acetone. In this study, three amino acid sites (Thr133, Ser134, Ser33) that may affect the function of PcPKS1 were identified by analyzing the sequences of PcPKS1, the BAS from Rheum palmatum and the CHS from Arabidopsis thaliana, as well as the conformation of the catalytic site of the enzyme. Molecular modification of PcPKS1 was carried out by site-directed mutagenesis, and two mutants were successfully obtained. The in vitro enzymatic reactions were carried out, and the differences in activity were detected by high performance liquid chromatography (HPLC). Finally, mutants T133LS134A and S339V with bifunctional activity were obtained. In addition to bifunctional activities of BAS and CHS, the modified PcPKS1 had much higher BAS activity than that of the wild type PcPKS1 under the conditions of pH 7.0 and pH 9.0, respectively. It provides a theoretical basis for future use of PcPKS1 in genetic engineering to regulate the biosynthesis of flavonoids and raspberry ketones.
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Amino Acid Sequence , Fallopia japonica/metabolism , Polyketide Synthases/chemistry , Acetone , Mutagenesis, Site-Directed , Flavonoids/metabolism , Acyltransferases/metabolismABSTRACT
OBJECTIVE To study main way and target of Euphorbia kansui after stir-frying with vinegar. METHODS Twenty-four SPF grade SD rats were randomly divided into blank group， E. kansui group （850 mg/kg） and vinegar stir-fried E. kansui group （850 mg/kg）， with 8 rats in each group. Blank group was given 0.5% sodium carboxymethyl cellulose solution intragastrically， and E. kansui group and vinegar stir-fried E. kansui group were given relevant test sample for consecutive 20 d. The rats’ urine of 12 hours was collected on the 20th day. The urine samples of rats in each group were determined by UPLC-Q- Exactive-MS. The data was pre-processed by Compound Discoverer 3.0 software， and the metabolite structure was identified by BioCyc， HMDB and other databases. Whether different groups presented their own clustering phenomenon was observed by principal component analysis （PCA） and orthogonal partial least squares-discriminant analysis （OPLS-DA）， etc. Based on the pathway analysis of MetaboAnalyst， the potential targets of detoxification mechanism of E. kansui after stir-frying with vinegar were predicted. RESULTS Twenty significantly differential endogenous metabolites were identified， of which 10 target metabolites， such as N-acetyl-L-aspartate and 3-phosphonooxypyruvic acid， were targets of detoxification mechanism of E. kansui after stir- frying with vinegar. The main metabolic pathways included arginine biosynthesis， alanine， aspartic acid and glutamic acid metabolism， cysteine and methionine metabolism， and arginine and D-ornithine metabolism. The biological significance of all related metabolites in the pathways was analyzed and speculated； after stir-frying with vinegar， E. kansui may alleviate neurotoxicity by reducing the level of N-acetyl-L-aspartic acid； E. kansui had a protective effect on cardio-cerebrovascular system by increasing the level of L-high arginine. CONCLUSIONS After stir-frying with vinegar， E. kansui can significantly improve the adverse factors in terms of nervous system， cardio-cerebrovascular system， immune system and energy metabolism. The most concentrated metabolic pathway related to its detoxification mechanism is arginine biosynthesis.
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Objective:To acquire the impact of COVID-19 pandemic on the operation of tertiary maternal and child health(MCH) hospitals in China, for decision making support of health administrative departments and hospital managers.Methods:The National Maternal and Child Health Institutions Resources and Operations Survey Direct Reporting System was used to collect the resource allocation, workload, treatment quality, work efficiency and asset operation of the tertiary MCH hospitals in China in 2019(pre pandemic) and 2020(during pandemic). Statistical descriptions were made using median.Results:In 2019 and 2020, the number of tertiary MCH hospitals in China was 236 and 258, respectively, and their relevant data were analyzed. In terms of resource allocation, the number of health technicians in 2019 and 2020 was 560 and 548, respectively, and the actual number of available beds was 308 and 305 respectively. In terms of workload, the annual outpatient visits in 2020 were 337 990, a decrease of 23.6%from that in 2019; The total number of emergency visits was 28 997, a decrease of 32.5%; The total number of discharged patients was 13 673, a decrease by 20.5%; A total of 4 723 training sessions on MCH were held for primary institutions, an increase of 1.2 percent. A total of 1 953 724 primary-level health technicians were trained, an increase of 175.2 percent. In terms of work efficiency, the average length of hospital stay of discharged patients decreased from 5.56 days in 2019 to 5.00 days in 2020. Bed utilization rate decreased from 88.90% to 69.15%; Bed turnover decreased from 53.69 to 44.22. In terms of treatment quality, the critical illness mortality rate of inpatients was 0.37% in 2020, 0.11% lower than that in 2019. The in-hospital mortality rate for neonatal patients was 0.04%, a 0.03% drop. In terms of asset operation, the total revenue in 2020 was 248.355 million yuan, an increase of 4.46% compared with 2019, in which the proportion of financial subsidies increased from 11.26% to 15.72%.Conclusions:The in-hospital services and institutional health care services of tertiary MCH hospitals in China were downsized by the COVID-19 pandemic, while the work efficiency was relatively stable, along with acceptable resource allocation, good treatment quality and asset operation.
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Objective@#To acquire the main result quality indicators status of tertiary maternal and child health(MCH) hospitals in China, supporting the decision making.@*Methods@#The National Maternal and Child Health Institutions Resources and Operations Survey Direct Reporting System was used to collect the management operation, inpatient death, re-return, hospital-acquired infection, surgical complications and patient safety of these hospitals in China in 2017. Statistical descriptions were made using average, rate and composition comparison data.@*Results@#The proportion of health technicians of tertiary MCH hospitals in China was 83.66%, the average hospitalization days were 5.96 days, with bed occupancy rate of 90.01%. The success rate of neonatal resuscitation was high. Meanwhile, the mortality rate of hospitalized maternal critical illness, the total hospitalization mortality rate of neonatal patients, the incidence of complications in surgical patients, the incidence of neonatal injury and the incidence of birth injury in vaginal delivery were higher in the institutions that had not participated in the MCH hospital accreditation.@*Conclusions@#The management of tertiary MCH hospitals in China was in good condition, and the relevant policies and projects have achieved remarkable results. Compared with general hospitals, hospital infection and re-return indicators were good. The quality and safety of tertiary MCH hospitals which have not participated in the MCH hospital assessment were poor. It is recommended to carry out MCH hospital accreditation to improve quality and safety.
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Objective:To study the expression of ubiquilin2 (UBQLN2) in non-small cell lung cancer (NSCLC) and its effect on the proliferation, invasion and metastasis ability of NSCLC cells.Methods:Immunohistochemistry was used to detect the expression of UBQLN2 in NSCLC cancer (24 cases) and adjacent normal tissues (24 cases), and to analyze the relationship between the expression of UBQLN2 and lymph node metastasis of NSCLC cancer. The expression of UBQLN2 in human normal bronchopulmonary epithelial cells and NSCLC cells was detected by real-time quantitative polymerase chain reaction (qRT-PCR)and Western blot; the effect of UBQLN2 on the proliferation of NSCLC cells was detected by lentivirus overexpression technology combined with MTS and EDU experiments in vitro; the effect of UBQLN2 on the invasion and metastasis of NSCLC cells was detected by scratch experiments in vitro and transwell experiments; a dual-fluorescence autophagy flow detection system was constructed by GFP-LC3-RFP-mLC3 plasmid packaging virus and Western blot was used to detect the change of autophagy after overexpression of UBQLN2; TCGA online data was uesd to analyze the expression level of UBQLN2 and lung cancer patients relevance of prognosis. Results:The expression of UBQLN2 in normal lung tissues was significantly higher than that in NSCLC tissues ( P<0.01), and the expression in patients with negative lymph node metastasis was significantly higher than that in patients with positive lymph node metastasis ( P<0.01); the expression of UBQLN2 in NSCLC cells was significantly lower than that in normal lung epithelial cells, and the overexpression of UBQLN2 could inhibit the proliferation and invasion and metastasis of tumor cells. The expression of UBQLN2 was positively correlated with the prognosis of NSCLC. Conclusions:The expression of UBQLN2 is significantly lower in lung cancer tissues and cells, and is negatively correlated with the lymph node metastasis of NSCLC; UBQLN2 can inhibit the proliferation and invasion of NSCLC cells; the expression of UBQLN2 is positively correlated with the prognosis of patients.
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Non-small cell lung cancer (NSCLC) is the histological subtype with highest proportion of lung cancer. Since the discovery of NSCLC driver gene mutations, the drug treatment of NSCLC had evolved from conventional chemotherapy to molecular targeted therapy. Epidermal growth factor receptor (EGFR) was one of the most important driver genes of NSCLC. Three generations of tyrosine kinase inhibitors (TKIs) targeting mutant EGFR had been developed and applied to the clinic, and EGFR-tKIs Targeted therapy had significantly improved the progression-free survival(PFS) and overall survival(OS) of patients with NSCLC. However, most NSCLC patients inevitably developed drug resistance within 10-18 months after receiving targeted therapy with EGFR-TKIs. Great progress had been made on the research of EGFR-TKIs resistance mechanism in recent years. This article intended to briefly review the resistance mechanism of EGFR-TKIs targeted therapy in terms of EGFR secondary mutation, signal bypass activation, cell lineage switching and tumor microenvironment, etc.
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Objective:APOBEC3B (A3B) is an important member of the apolipoprotein B mRNA editing enzyme catalytic polypeptide (APOBEC) family. The study aims to investigate the relationship between A3B expression and prognosis as well as resistance to cisplatin in non-small cell lung cancer (NSCLC).Methods:Real-time quantitative polymerase chain reaction (qRT-PCR) was used to analysis the A3B mRNA expression in 40 NSCLC tissues; Kaplan Meier plotter was used to analyse the correlation between A3B expression and clinical prognosis; in addition, the knock-down A3B expression cell line in human lung adenocarcinoma cell A549 was constructed; MTS and plate cloning experiment were performed to observe the changes in cell cisplatin sensitivity, and γ-H2AX immunofluorescence was used to quantitate the DNA damage.Results:Compared with adjacent tissues, A3B was highly expressed in NSCLC tissues (27/40). Kaplan Meier plotter analysis showed that A3B expression was positively correlated with NSCLC overall survival (OS) [adenocarcinoma: HR=0.64(0.47-0.86), P=0.002 6; squamous cell carcinoma: HR=0.77(0.59-1.01), P=0.006]. Cell-based studies showed that the knock-down A3B expression contributed to sensitivity to cisplatin in A549 cells. Conclusions:A3B mediates the sensitivity of lung cancer to cisplatin. This effect may partly explain why NSCLC patients with high A3B expression have a better prognosis.
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Objective:To explore the role of small molecule glycoprotein Serglycin (SRGN) in chemotherapy resistance of non-small cell lung cancer (NSCLC).Methods:In NSCLC H1299 cell line, shRNA technology was used to interfere with the expression of SRGN and establish stable interfering cell line. Western blot and real time fluorescence quantitative polymerase chain reaction (qRT-PCR) were used to verify the knockdown efficiency; MTS was used to detect the knockdown cell line′s drug sensitivity to cDDP and Oxaliplatin; enzyme linked immunosorbent assay (ELISA), Western blot and qRT-PCR were used to explore the effect of transforming growth factor β (TGFβ) on SRGN and vice versa; Western blot was used to detect the effect of SRGN on epithelial-mesenchymal transition (EMT) related molecules, and online data bioinformatics was used to analyze the correlation between SRGN and EMT related molecules expression; in addition, online prognostic analysis software (kmplot) was used to analyze the correlation between SRGN, TGFβ and prognosis of lung cancer patients.Results:Comparing with the control group, the test group, knocking down SRGN can obviously improve the drug sensitivity of NSCLC cell to cDDP ( P=0.032 7) or Oxaliplatin ( P=0.014 2). TGFβ can enhance the experission of SRGN in NSCLC and SRGN also can help TGFβ secreted from cells. SRGN promotes the epithelial mesenchyme transition by modulating Snail1. By analyzing TCGA database, we found that the expression of SRGN was negatively correlated with the expression of CDH1 (coding for Ecadherin protein) ( r=-0.25) and there was a positive correlation with Snai1 expression ( r=0.37). These results suggest that SRGN can promote the change of EMT in lung cancer cells through TGF β 1 and snail 1. The overall survival time of NSCLC patients with low expression of SRGN was much longer than the patients with high expression of SRGN ( P=0.007 7). The overall survival time of NSCLC patient with low expression in both SRGN and TGFβ1 or TGFβ2 was 73months or 42.8 months longer than that with high expression in both SRGN and TGFβ1/2. Conclusions:Intercting with TGFβ1, SRGN promotes EMT of NSCLC cells, which facilitates the chemoresistence of NSCLC. The simultaneous low expression of SRGN and TGFβ1 or TGFβ2 can significantly prolong the overall survival of patients with NSCLC.
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Objective:To establish osimertinib-resistant non-small cell lung cancer (NSCLC) cell line and explore its drug resistance mechanism.Methods:The human NSCLC cell line H1975 was used as the research object, and low-concentration osimertinib was used to continuously select secondary drug-resistant cell lines. Osimertinib drug sensitivity of cells was detected by MTS method. Cell proliferation was detected by live cell workstations. Flow cytometry was used to detect cell cycle and apoptosis. Protein mass spectrometry was used to construct differentially expressed protein profiles between parental and drug-resistant cells and some resistance-related proteins were validated by real time fluorescence quantitative polymerase chain reaction (qRT-PCR) and Western blot.Results:Secondary drug-resistant H1975/OSI cell line were successfully established. Compared with the parental cells, the resistance index of H1975/OSI cells increased by 27.25 times ( P<0.01), the cell proliferation ability decreased but the apoptosis resistance increased ( P=0.01), and no new drug-resistance related gene mutation in H1975/OSI cells. Meanwhile, the differential protein expression profiles of H1975 and H1975/OSI cells were built, and 307 upregulated proteins and 295 down-regulated proteins were found in resistant cells. When fibroblast specific protein-1 (FSP1) gene with expression up-regulation was diturbed in H1975/OSI cells, the cell IC50 value of osimertinib decreased 3.51 times ( P=0.02) , and when FSP1 was overexpressed in the H1975 cells, the IC50 value of osimertinib increased by 3.75 times ( P<0.01). Conclusions:We successfully established human NSCLC osimitinib-resistant cell line H1975/OSI. Protein differential expression profiles between H1975 and H1975/OSI was constructed successfully. It was found that FSP1 was involved in mediating the resistance of H1975/OSI to osimertinib.
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Objective:To acquire the main result quality indicators status of tertiary maternal and child health(MCH) hospitals in China, supporting the decision making.Methods:The National Maternal and Child Health Institutions Resources and Operations Survey Direct Reporting System was used to collect the management operation, inpatient death, re-return, hospital-acquired infection, surgical complications and patient safety of these hospitals in China in 2017. Statistical descriptions were made using average, rate and composition comparison data.Results:The proportion of health technicians of tertiary MCH hospitals in China was 83.66%, the average hospitalization days were 5.96 days, with bed occupancy rate of 90.01%. The success rate of neonatal resuscitation was high. Meanwhile, the mortality rate of hospitalized maternal critical illness, the total hospitalization mortality rate of neonatal patients, the incidence of complications in surgical patients, the incidence of neonatal injury and the incidence of birth injury in vaginal delivery were higher in the institutions that had not participated in the MCH hospital accreditation.Conclusions:The management of tertiary MCH hospitals in China was in good condition, and the relevant policies and projects have achieved remarkable results. Compared with general hospitals, hospital infection and re-return indicators were good. The quality and safety of tertiary MCH hospitals which have not participated in the MCH hospital assessment were poor. It is recommended to carry out MCH hospital accreditation to improve quality and safety.
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Objective APOBEC3B (A3B) is an important member of the apolipoprotein B mRNA editing enzyme catalytic polypeptide (APOBEC) family.This study aimed to investigate its important role in the metastasis of small cell lung cancer (NSCLC).Methods The statistical relationship between A3 B and clinicopathological data was analyzed in 249 cases of NSCLC.Sanger sequencing was used to detect mutations in exon 5,6,7 and 8 of P53 in 74 cases of lung cancer.A3B overexpression cell line was constructed in human lung adenocarcinoma cells HCC827 to observe the change of cell migration and metastasis capacity.Results A3B was highly expressed in NSCLC tissues compared with normal lung tissues.The expression of A3B was closely related to the lymph node metastasis of NSCLC and the mutation rate of p53 was positively correlated with the expression of A3B.In vitro experiment,it showed enhanced migration and increased metastatic potential in cells after overexpression of A3B.Conclusions A3B-mediated mutations in P53 may play a key role in the metastasis of NSCLC.
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Objective To study the role of leucine rich repeat neuronal 3 (LRRN3) in the prolif-eration of non-small cell lung cancer and its possible mechanism of expression regulation. Methods The expression of LRRN3 in non-small cell lung cancer was detected by real-time quantitative polymerase chain reaction ( qPCR) , immunohistochemistry and bioinformatics retrieval;A lung cancer cell line A549-LRRN3 with stable over-expression of LRRN3 was established by lentivirus over-expression technology;The effect of LRRN3 on the proliferation of non-small cell lung cancer was detected by methyl thiazolyl tetrazolium ( MTT) assay;Bioinformatics search for changes in methylation of LRRN3 promoter region and treatment of lung cancer cells by methyltransferase inhibitors to detect the effect of methylation on the regulation of LR-RN3 expression; Finally, bioinformatics search analyzes the correlation between LRRN3 and lung cancer prognosis. Results The mRNA expression of LRRN3 in clinical tissues of non-small cell lung cancer (n=12) was significantly lower than that of adjacent normal tissues (n=12) (P=0. 0014). The results of immunohistochemistry showed that the protein expression level of LRRN3 in non-small cell lung adenocar-cinoma was lower than that in normal tissues (P=0. 001), and the expression in non-small cell lung squa-mous cell carcinoma was also lower than that in normal tissues (P=0. 003). Overexpression of LRRN3 in-hibited the proliferation of tumor cells (P<0. 01), and the hypermethylation of LRRN3 in the promoter re-gion inhibited its transcriptional expression. LRRN3 was positively correlated with the survival prognosis of lung adenocarcinoma (P=5. 2e-09;HR=0. 48). Conclusions Hypermethylation in the promoter region of LRRN3 inhibits its transcriptional expression, thereby promoting the proliferation of lung cancer cells.
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Objective:To establish the HPLC specific chromatogram and provide comprehensive evaluation of Xanthii fructus from different regions.Methods:The HPLC analysis was performed on an Alltima C18 column (250 mm ×4.6 mm,5μm) with acetonitrile-0.1％phosphoric acid solution as the mobile phase with gradient elution at a flow rate of 0.8 ml· min-1 .The detection wavelength was 278 nm.The column temperature was 25℃.The software"Similarity Evaluation System for Chromatographic Fingerprint of TCMs"was employed to carry out the similarity analysis of the samples from Henan , Jilin, Anhui and the other regions .Results:The specific chromatogram was preliminarily constructed and 5 common peaks with chlorogenic acid as the reference were identified .Conclusion:The method is scientific basis of the quality assessment of Xanthii fructus with convenient and reliable properties .
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Objective:To explore the influential factors for hospitalization costs regarding the final phase of malignant tumor patients in Shanghai,and to explore the relevant policy for reasonable control of hospitalization costs.Methods:A total of 10 065 patients with malignant tumors were enrolled in this study.The multiple linear regression analysis was used to seek the determinants for hospitalization cost of malignant tumor patients during the final phase.Results:The median length of hospital stay was 43 days for the patients,with an average age of (70.73±12.87) years.Among them 61.66％ of hospitalized patients were male and the median hospitalization cost of malignancy was 55 447.84 yuan.Hospitalization cost showed the linear regression relationship with type of health care,hospital level,hospital types,tumor types,length of hospital stay,surgery,age,gender,and time from hospital admission to death.Conclusion:Proximity to death in malignant tumor patients is an important factor for the hospitalization cost.Medical resources should be allocated rationally,and the comprehensive measures should be taken to control the cost reasonably.
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Objective@#To analyze the therapeutic effect on HBeAg-negative chronic hepatitis B patients treated with Peg-IFNα-2a combined with NAs to obtain the influencing factors for predicting HBsAg clearance.@*Methods@#A retrospective study was conducted to investigate the effect of pegylated interferon alpha-2a combined with nucleoside analogues (lamivudine/adefovir dipivoxil) on HBeAg-negative chronic hepatitis B. The treatment course was 96 weeks. Patients were followed up 120 weeks after the treatment. HBsAg clearance at 120 weeks was taken as the objective of the study. Logistic regression and receiver operating characteristic curve analysis screened the related factors affecting HBsAg clearance. χ 2 test was used to compare count data.@*Results@#111 patients were treated with pegylated interferon alpha-2a combined with nucleoside analogues, and 107 patients completed the scheduled course of treatment and follow-up. HBsAg clearance rate at120 week was 29.0% (31/107). The influencing factors for analysis were: (1) gender had no effect on HBsAg clearance rate; age and baseline levels of HBV DNA and alanine aminotransferase had no significant effect on HBsAg clearance; low baseline level of HBsAg (< 3.023 lgIU/ml) was beneficial to HBsAg clearance. The area under the working characteristic curve of the subjects was 0.746, the positive predictive value was 44.4%, and the negative predictive value was 86.8%. (2) HBsAg quantification or decline in 24 weeks and 48 weeks of treatment had a good predictive effect on HBsAg clearance, and the 48 weeks predicted value was higher than 24 weeks. When the HBsAg quantification was≤2.070 lgIU/ml at 48 weeks, the area under the receiver operating characteristic curve was 0.931, the positive predictive value was 52.8%, and the negative predictive value was 94.4%. When HBsAg decreased from baseline to≥0.991 lgIU/ml, the area under the receiver operating characteristic curve was 0.888, the positive predictive value was 50.8%, and the negative predictive value was 97.9%. (3) The analysis of HBsAg subgroup levels at 48 weeks suggested that the "interval analysis" can forecast HBsAg clearance more exactly than "nodal analysis" .The final HBsAg clearance rate of 100 IU/ml < HBsAg≤1 000 IU/ml, 10 IU/ml < HBsAg≤100 IU/ml and HBsAg≤10 IU/ml groups reached 6.7%, 31.8% and 67.7%, respectively. (4) The ALT abnormal group in the course of treatment obtained a higher HBsAg clearance rate (48.0%, 12/25).@*Conclusion@#96-weeks long-term treatment with pegylated interferon-alpha -alpha-2a combined with nucleoside analogues for HBeAg-negative chronic hepatitis B has a good predictive value for HBsAg clearance at baseline and during treatment. The "interval level" of HBsAg at 48-weeks is more accurate in predicting HBsAg clearance, suggesting that HBeAg-negative chronic hepatitis B patients with low HBsAg levels at 48-weeks are the advantageous populations with HBsAg clearance. These patients are worthy of prolonged treatment to pursue "clinical cure".
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Objective To explore the effect of tumor-associated macrophage (TAM) on invasion and migration of breast cancer cells T47D.Methods Briefly,phorbol 12-myristate 13-acetate (PMA) was employed to treat mononuclear cells THP-1.And then interleukin 4 (IL-4) was applied to stimulate mentioned cells to establish activated TAM.Furthermore,the supernatant of M0 and M2 was used to culture T47D cells,respectively.Then,the wound healing experiment and transwell assay were carried out to investigate cells invasion and migration.Finally,epithelial-mesenchymal transition (EMT) biomarkers in T47D cells treated with M0 or M2 were determined by quantitative real time polymerase chain reaction (qRT-PCR) and western blot assays.Results Activated TAM model was successfully established by PMA following with IL-4.Comparing with M0,M2 evidently accelerated invasion and migration in T47D cells.Meanwhile,M2 upregulated N-cadherin,vimentin and β-catenin,downregulated E-cadherin at mRNA and protein expression levels.Conclusions M2 promotes invasion and migration of T47D cells via EMT.
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The interaction between tumor cells and tumor microenvironment is a necessary condition for tumor metastasis.Tumor-associated macrophages (TAM) are macrophages that infiltrate tumor tissues and are the most abundant immune cells in the tumor microenvironment.Tumor-associated macrophages play a key role in tumor invasion and metastasis,and has been suggested an important target for anti-tumor metastasis treatment.This review focuses on the molecular mechanisms related to tumor-associated macrophage polarization and its role in tumor invasion and metastasis.
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Objective To evaluate the feasibility and safety profile of pegylated-interferonα-2a (Peg IFNα-2a) combined with adefovir dipivoxil (ADV) in inactive hepatitis B surface antigen (HBsAg) carriers (IHC).Methods This was a single center, prospective and open-label study.IHC were divided into therapeutic group (T, 112 subjects) and control group (C, 72 subjects) according to personal willingness.Patients with hepatitis B virus (HBV) DNA<20 IU/mL were treated with Peg IFNα-2a monotherapy, and those with HBV DNA ≥20-<2 000 IU/mL were treated with Peg IFNα-2a combined with ADV.Total therapy duration was 96 weeks.For patients who achieved HBsAg seroconversion and continued consolidation treatment for 24 weeks, the treatment duration could be less than 96 weeks.t test was used for continuous variable comparison between the two groups, while chi-square test or Fisher′s exact probability method was used for counting data analysis.The related factors affecting HBsAg clearance was analyzed by univariate or multivariate logistic regression analysis.Results A total of 194 patients were enrolled with 112 in therapeutic group and 72 in control group.The HBsAg clearance rate and seroconversion rate at week 48 in therapeutic group were 30.8% (32/104) and 26.0% (27/104), respectively.The rates at week 96 increased to 45.2% (47/104) and 38.5% (40/104), respectively.The HBsAg clearance rates at weeks 48 and 96 in control group were both 1.5% (1/68).HBsAg seroconversion was not achieved in control group.The HBsAg clearance rate in treatment group was significantly higher than that in control group (χ2=39.066, P<0.01).The quantitative HBsAg levels at baseline (OR=2.313, 95%CI: 1.258-4.251, P=0.007), week 12 (OR=3.159, 95%CI: 1.826-5.466, P<0.01) and week 24 (OR=3.347, 95%CI: 2.050-5.465, P<0.01), the decline of HBsAg at week 12 (OR=5.343, 95%CI: 2.085-13.689, P<0.01), and week 24 (OR=4.855, 95%CI: 2.380-9.902, P<0.01), and alanine transaminase (ALT) elevation at week 12 (OR=3.520, 95%CI: 1.369-9.052, P=0.009) were independent predictors for HBsAg clearance.Conclusions Peg IFNα-2a-based treatment for IHC could achieve higher HBsAg clearance rate and seroconversion rate, and has a safety profile.Decline of HBsAg at week 12 and week 24 with ALT elevation at week 12 could predict a higher HBsAg clearance rate.
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Objective The method for detecting expression of human CDK14 gene with Real-time quantitative PCR was developed.Methods To establish a method for detecting expression of human CDK14 gene with Real-time quantitative PCR by designing and synthesis of the primers of CDK14 target gene andβ-Actin reference gene and extracting total RNA from different lung cancer cell lines.Then the specificity,detection range and repeatability of this method were evaluated.At last,the expression level of CDK14 gene in different cell lines,which were with or without siRNA interference,were carried out by using this method.Results The method for detecting expression of human CDK14 gene with Real-time quantitative PCR,which had good specificity,good repeatability (CV=7.3 ％) and wide detection range (Ct value range of CDK14 and β-Actin amplification curve were 22.47～32.96 and 15.14～ 27.55 respectively,r2 =0.9844),was developed and it was verified by electrophoresis analysis,melting curve,PCR product sequencing.And CDK14 gene expression level,which was detected by this method,increased in HCC827 D5,H1650 and number 1 siRNA segment was effective interference segment.Conclusion The method for detecting expression of human CDK14 gene with Real-time quantitauve PCR was established successfully.
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Objective To explore the role of forkhead box protein O3a (FOXO3a) in inhibiting the proliferation and metastasis of breast cancer cells by downregulation of vascular endothelial growth factorA (VEGF-A).Methods Cell proliferation bioassay and plate clone formation assay were used to compare the changes of proliferation after siRNA interference.Wound healing and Transwell were used to compare the changes of invasion metastasis after interference.Meanwhile,the changes of VEGF-A expression in the cells before and after interference were compared by real time polymerase chain reaction (RT-PCR) and Western blot.Furthermore,the expressions of mRNA FOXO3a and VEGF-A were detected in 20 cases of breast cancer patients in breast cancer tissues and adjacent normal breast tissues.Results The MCF-7 and MDA-MB-231 cells were increased in cell proliferation and invasion after interference.Further studies found that mRNA and protein expression of VEGF-A were up-regulated in MCF-7 and MDA-MB-231cells after interference.In vivo the expression of FOXO3a was lower in 15 cases of cancer compared to normal tissues,and the expression of VEGF-A was high in 15 cases of cancer (75％).FOXO3a and VEGF-A expression was highly negatively correlated.Conclusions This study showed that FOXO3a could inhibit the proliferation and invasion of breast cancer cells,which might be regulated by VEGF-A.It provides an important theoretical evidence for targeted inhibition of breast cancer metastasis.