Objective To investigate the relationship between growth inhibiting effect of arsenic trioxide(As_2O_3) and phosphorylation of P27kip threonine residue 187(P27T187) in human hepatocellular carcinoma(HCC) cell line SMMC-7721.Methods SMMC-7721 were treated for 72 h with 2 ?mol/L As_2O_3.The cell growth inhibition was detected by cell counting and the cell cycle was detected by flow cytometry(FCM).The expression and localization of P27,T187 phosphorylated P27(p-P27T187) were detected by Subcellular Fractionation,Western blot and immunoflurescence.Results As_2O_3 significantly inhibited the proliferation of SMMC-7721 cell and cell cycle was arrested in G2/M.A significant decrease in p-P27T187 expression and a reciprocal increase in P27 expression were found in 2 ?mol/L As_2O_3-treated SMMC-7721 cell.Meanwhile,As_2O_3 decreased the protein levels of Cdk2 and cyclinE.The location of P27 was transferred from cytoplasm to nuclei and the expression of p-P27T187 was decreased in nuclei.Conclusion As_2O_3 inhibits the phosphorylation of P27T187,thereby promoting P27 accumu-lation in SMMC-7721 cell nuclei,inducing cel1 cycle arrest and growth inhibition.

Objective To investigate the expression of C-JUN activation domain binding protein 1(JAB1)and its relationship with expression of P27 protein in human hepatocellular carcinoma(HCC),and to determine whether JAB1 is associated with clinicopathological parameters and prognosis of HCC.Methods Immunohistochemical analysis was performed to investigate the expression of JAB1 and P27 in 76 cases of HCC and adjacent nontumorous tissues.Fresh tumor tissues and their adjacent nontumorous tissues from 8 cases of HCC were collected for Western blot and immunoprecipitation assays.Results The expression of JAB1 in HCC was significantly higher than that in adjacent nontumorous tissues.In contrast,P27 level was higher in nontumorous liver tissues than that in HCC.JAB1 overexpression was correlated with histological differentiation,serum alpha-fetoprotein(AFP)level and metastasis(P

Objective To investigate the expression of DNA methyltransferase 3b (DNMT3B) in hepatocellular carcinoma (HCC) and its effect and mechanism on the proliferation,invasion and migration of HCC cells.Methods The expression of DNMT3B gene was detected by qRT-PCR in 46 cases of HCC tissues and corresponding adjacent tissues;the results and clinical pathological parameters were analyzed.SiRNA targeting DNMT3B was transfected into MHCC97-H cells by RNA interference (RNAi) technique.The mRNA and protein expression levels of related genes were detected by qRT-PCR and Western blot.The cell proliferation was measured by MTT assay,and the invasion and migration abilities were measured by Transwell assay.Results In 46 HCC patients,the expression of DNMT3B (73.91％) was significantly higher in HCC than in adjacent normal tissue.The high expression of DNMT3B gene was associated with histological type and tumor size of HCC (all P＜0.05).Inhibition of DNMT3B gene expression decreased proliferation,invasion and migration of MHCC97-H cells.Interference with DNMT3B gene increased the expressions of tumor suppressor genes RASSFA1,APC and MTSS1 at mRNA and protein levels.Conclusion DNMT3B is associated with the progression of HCC.It may inhibit the proliferation,invasion and migration of HCC cells by regulating the methylation of downstream tumor suppressor gene.

ABSTRACT:Objective To observe the influence of saikosaponin-d (SSd)on the proliferation and the function of autophagy of human hepatocellular carcinoma (HCC)cell line SMMC-7721 to explore the possible mechanisms. Methods SMMC-7721 was cultured invitro and then treated with SSd of various concentrations (5.0,7.5,10.0, 12.5,15.0 and 17.5 mg/L)for 24,48 and 72 h.We used MTT to detect cell proliferation,selected the optimal concentration and time,and detected the expressions of BECN1 at mRNA and protein levels by PCR and Western blot.Results The inhibition rate of the proliferation of SMMC-7721 cell line increased with the increase of the concentration of SSd,and the highest inhibition rate (60%)appeared when the concentration reached 12.5 mg/L. The expression of BECN1 in the group with SSd was obviously higher than that in the control group (P<0.05). 3-MA decreased not only the expressions of BECN1 at mRNA and protein levels but also the expression of BECN1 when used in conjunction with SSd.Conclusion The inhibiting function of SSd on SMMC-7721 presents a dependency between drug concentration and function time,basically in line with the drug dose-effect relationship. SSd induces the occurrence of autophagic cell death through up-regulating the expression of BECN1 ,thus inhibiting the proliferation of SMMC-7 7 2 1 .

Objective:To explore the expression of polypyrimidine tract-binding protein 1 (PTBP1) in gastric cancer (GC) tissues and GC cell lines, and the role of PTBP1 in the proliferation and metastasis of GC cells.Methods:From January to June in 2019 at The First Affiliated Hospital of Xi′an Jiaotong University, the cancer tissues and corresponding para-cancer tissues of GC patients underwent surgical resection were collected. The Kaplan-Meier Plotter database was used to analyze the survival of GC patients. The expression of PTBP1 was down-regulated by transfecting small interfering RNA (siRNA) in human GC cell lines SGC7901 and AGS with relatively high expression of PTBP1. The cells were divided into blank control group, negative control group, and PTBP1 knockdown group. The expression of PTBP1 at mRNA and protein level were detected by real-time fluorescence quantification polymerase chain reaction (RT-qPCR) and Western blotting. At 24, 48, 72 and 96-hour after transfection, the effect of PTBP1 on the proliferation of GC cells was observed by 3-(4, 5 dimethylthiazol)-2, 5 diphenyltetrazolium bromide (MTT) experiment. The changes of invasion and migration of GC cells after down-regulation of PTBP1 were detected by transwell assay. The expression changes of epithelial-mesenchymal transition (EMT) markers E-cadherin, N-cadherin and vimentin after down-regulation of PTBP1 in GC cells were determined by Western blotting. Indenpendent samples t test, analysis of variance and rank sum test were used for statistical analysis. Results:The Kaplan-Meier Plotter prognostic analysis showed that the overall survival of GC patients with high PTBP1 expression was shorter than that of GC patients with low PTBP1 expression (9.2 months, 6.2 months to 17.2 months vs. 19.0 months, 14.5 months to 28.4 months), and the difference was statistically significant ( Z=5.31, P<0.05). The results of RT-qPCR showed that in GC cell lines SGC7901 and AGS, the expression of PTBP1 at mRNA level of PTBP1 knockdown group was lower than that of blank control group and negative control group (SGC7901: 0.78±0.11 vs.3.10±0.19 and 2.99±0.23; AGS: 0.80±0.09 vs. 3.55±0.24 and 3.50±0.18), and the differences were statistically significant ( tSGC7901=10.57 and 8.08, tAGS=10.91 and 13.42; all P<0.01). The results of Western blotting indicated that in GC cell lines SGC7901 and AGS, the expression of PTBP1 at protein level of PTBP1 knockdown group was lower than those of blank control group and negative control group (SGC7901: 0.38±0.04 vs. 1.42±0.05 and 1.35±0.09; AGS: 0.17±0.02 vs. 1.52±0.08 and 1.38±0.45), and the differences were statistically significant ( tSGC7901=15.94 and 10.57, tAGS=16.60 and 20.80; all P<0.01). The results of MTT showed that at 48, 72 and 96-hour after transfection the absorbance values of PTBP1 knockdown group decreased by 0.25±0.01, 0.38±0.02, and 0.84±0.04 as compared with those of negative control group, and the decrease was the most significant at 96-hour after transfection, and the differences were statistically significant ( t=10.21、14.32, both P<0.01). The results of transwell experiment demonstrated that the number of invasion and migration cells of PTBP1 knockdown group were both less than that of the blank control group and the negative control group (SGC7901: 42.00±5.91 vs. 116.40±10.23 and 114.40±10.43; 39.60±6.77 vs. 125.80±11.51 and 122.40±5.90; AGS: 40.20±7.25 vs. 115.60±14.63 and 117.40±9.12; 36.00±5.20 vs. 122.40±12.10 and 125.40±12.74), and the differences were statistically significant ( tSGC7901=14.07, 13.50, 14.43 and 20.62; tAGS=10.27, 14.75, 14.68 and 16.76; all P<0.01). The results of Western blotting showed that the expression of E-cadherin of PTBP1 knockdown group was higher than that of the blank control group and the negative control group (SGC7901: 1.42±0.05 vs. 0.53±0.05 and 0.57±0.03; AGS: 1.34±0.04 vs. 0.54±0.03 and 0.61±0.01), however the expression levels of N-cadherin and vimentin were both lower than those of the blank control group and the negative control group (SGC7901: 0.50±0.03 vs. 1.64±0.05 and 1.46±0.07; 0.32±0.07 vs. 1.42±0.07 and 1.33±0.07; AGS: 0.37±0.06 vs. 1.47±0.04 and 1.36±0.04; 0.41±0.04 vs. 1.53±0.06 and 1.37±0.04), and the differences were statistically significant ( tSGC7901=11.63, 13.19, 18.83, 11.68, 11.43 and 10.43; tAGS= 15.02, 16.23, 14.67, 12.97, 14.45 and 17.18; all P<0.01). Conclusions:The expression levels of PTBP1 increase in GC tissues and cells, which may be involved in regulating the proliferation, metastasis and EMT of GC cells.

Objective To detect the expressions of hTERT and MMP-7 mRNA in the peripheral blood of patients with colorectal cancer and their correlation,and discuss the clinical significance of peripheral blood micro-metastasis.Methods Total RNA was extracted from blood samples by TRIzol and was reverse transcribed into cDNA.The mRNA expressions of MMP-7 and hTERT were detected by real-time PCR.Results Positive mRNA expression of hTERT was found in 63 of 181 patients with colorectal cancer,while positive mRNA expression of MMP-7 was seen in 61 cases.The positive rates of hTERT and MMP-7 mRNA had no significant differences in the patients' sex and age (P ＞ 0.05).The positive rate of MMP-7 mRNA in the patients with poorly differentiated adenocarcinoma (43.9 ％) was higher than that in the patients with moderately and well differentiated adenocarcinoma (27.8 ％) (x2 =4.873,P ＜ 0.05).The positive rate of HTERT mRNA in the patients with stage Ⅲ-Ⅳ (42.5 ％) was higher than that in the patients with stage Ⅰ-Ⅱ (24.0 ％) (x2 =6.591,P ＜ 0.05).The positive rate of MMP-7 mRNA in the patients with stage Ⅲ-Ⅳ (39.6 ％) was higher than that in the patients with stage Ⅰ-1Ⅱ (25.3 ％) (x2 =4.014,P ＜ 0.05).The positive rate of hTERT mRNA in the patients with distant metastasis (53.3 ％) was higher than that in the patients without distant metastasis (28.7 ％) (x2 =9.059,P ＜ 0.05).The positive rate of MMP-7 mRNA in the patients with distant metastasis(46.7 ％) was higher than that in the patients without distant metastasis (29.4 ％) (x2 =4.506,P ＜ 0.05).The positive rate of hTERT mRNA in the patients with positive CEA expression (52.6 ％) was significantly higher than that in the patients with negative CEA expression (30.1％) (x2 =6.735,P ＜ 0.05).The positive rate of MMP-7mRNA in the patients with positive CEA expression (60.5 ％) was significantly higher than those in the patients with negative CEA expression (26.6 ％) (x2 =15.490,P ＜ 0.05).In the patients without distant metastasis,any of hTERT and MMP-7 mRNA expression was thought to be blood micro-metastasis.The recurrence and metastasis rates after 6 months in the patients with blood micrometastasis (36.58 ％) were higher than those in the patients without blood micro-metastasis (7.14 ％)(x2 =13.027,P ＜ 0.05).The recurrence and metastasis rates after 12-month follow-up in the patients with blood micro-metastasis (68.29 ％) were higher than those in the patients without blood micro-metastasis (12.5 ％) (x2 =31.948,P ＜ 0.01).The sensitivity of combined detection was 80.00 ％,and the specificity was 79.03 ％ after 12-month follow-up.Spearman correlation analysis revealed that significant positive correlation existed between hTERT and MMP-7mRNA expression (r =0.341,P =0.000).Conclusion Detection of hTERT and MMP-7 mRNA expression can be used as reliable markers to predict peripheral blood micrometastasis in colorectal cancer patients.

Objective To detect the expression profile of transcription factor C/EBPβ in human immortalized normal hepatic cell lines and hepatocellular carcinoma cell lines so as to determine the correlation between C/EBP3 with cell death mediated by endoplasmic reticulum stress in hepatocellular cells.Methods We cultured the human immortalized normal hepatic cells lines HHL5 and HL7702 and hepatocellular carcinoma cell lines SMMC7721;Bel7402,HepG2 and Hep3B.Hep3B cells were used as the cell model in tunicamycin-induced endoplasmic reticulum stress.Cellular morphology was observed under an inverted optical microscope.MTT assay was used to assess the inhibition of cell growth.To detect cell apoptosis,the cells were dyed with Hoechst 33258 and observed using a fluorescence microscope.RToPCR and Western blotting were used to detect the expression of at mRNA and protein levels,respectively.Results We found that normally the mRNA and protein isoform of C/EBPβ,C/EBPβ-1,were both expressed in all of the four hepatocellular cell lines and the two immortalized normal hepatic cell lines,while C/EBPβ protein isoform C/EBPβ-3 was only expressed in the two immortalized normal hepatic cell lines.Tunicamycin increased the expressions of both mRNA and protein of C/EBPβ in Hep3B cells and the increase of protein isoform C/EBPβ-3 was the most remarkable.In Hep3B cells,cell death was induced by tunicamycin through endoplasmic reticulum stress activity.Apoptosis as well as paraptosis was observed in tunicamycin-induced cell death.Conclusion C/EBPβ-3,one of the protein isoforms of C/EBPβ,is only expressed in normal hepatic cell lines,but not in hepatocellular cell lines.C/EBPβ is involved in cell death mediated by endoplasmic reticulum stress activity in hepatocellular carcinoma cells.

OBJECTIVETo establish a microwave extraction and UPLC method for simultaneous determination of polydatin, resveratrol, anthraglycoside B, emodin and physicion contained in Polygoni Cuspidati Rhizoma et Radix, in order to provide scientific basis for improving quality standards of Polygoni Cuspidati Rhizoma et Radix.

METHODThe test solutions were prepared in a MDS-8 closed microwave system at 160 degrees C with methanol as the solvent. The UPLC analysis was performed in a Waters Acquity UPLC system. A BEH C18 column (2. 1 mm x 100 mm, 1.7 microm) was adopted for gradient elution with acetonitrile and water as the mobile phase. The temperature of column was 30 degrees C, and the detection wavelength was 226 nm.

RESULTThe five active components can be completely extracted in 10 minutes and separated completely in 12 minutes according to UPLC analysis, with a good linearity (r > or = 0. 999 6) within the linear ranges. The average recovery rate was 97.00%-103.7% with RSD < or = 2. 2%. Despite a large difference in content among tested components from Polygoni Cuspidati Rhizoma et Radix, the total content of the five major constituents was relatiely stable (3.683 3%-7.1031%).

CONCLUSIONThe microwave extraction-ultra performance liquid chromatography method in simultaneous determination for contents of five major bioactive components contained in polygoni cuspidati rhizoma et radix is so rapid and highly reproducible that it can be used for quality control and assessment of Polygoni Cuspidati Rhizoma et Radix.

Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; chemistry ; Fallopia japonica ; chemistry ; Medicine, Chinese Traditional ; Microwaves ; Plant Roots ; chemistry ; Plant Stems ; chemistry ; Quality Control

OBJECTIVE: To compare the efficacy and safety of combined radiofrequency ablation (RFA) and transcatheter arterial chemoembolization (TACE) with RFA alone for hepatocellular carcinomas (HCC). MATERIALS AND METHODS: Randomized controlled trial (RCT) studies that compared the clinical or oncologic outcomes of combination therapy of TACE and RFA versus RFA for the treatment of HCC were identified through literature searches of electronic databases (Pubmed, Embase, Cochrane Library, China Biology Medicine disc, China National Knowledge Infrastructure, and Google Scholar). Hazard ratios (HRs) or odds ratios (ORs) with their corresponding 95% confidence interval (CI) were combined as the effective value to assess the summary effects. The strength of evidence was rated by the Grading of Recommendations Assessment, Development, and Evaluation system. RESULTS: Six RCTs with 534 patients were eligible for inclusion in this meta-analysis. The meta-analysis showed that the combination of TACE and RFA is associated with a significantly longer overall survival (HR = 0.62, 95% CI: 0.49-0.78, p < 0.001) and recurrence-free survival (HR = 0.55, 95% CI: 0.40-0.76, p < 0.001) in contrast with RFA monotherapy. The seemingly higher incidence of major complications in the combination group compared with RFA group did not reach statistical significance (OR = 1.17, 95% CI: 0.39-3.55, p = 0.78). CONCLUSION: In patients with HCC, the combination of TACE and RFA is associated with significantly higher overall survival and recurrence-free survival, as compared with RFA monotherapy, without significant difference in major complications.
Carcinoma, Hepatocellular/*surgery ; Catheter Ablation/adverse effects/*methods ; Chemoembolization, Therapeutic/adverse effects/*methods ; China ; Combined Modality Therapy ; Disease-Free Survival ; Humans ; Liver Neoplasms/*surgery ; Odds Ratio ; Proportional Hazards Models ; Treatment Outcome

Objective:To evaluate the feasibility and effectiveness of magnetic anchor-guided endoscopic submucosal dissection (MAG-ESD).Methods:A total of 36 patients with gastrointestinal tumors at different sites who underwent MAG-ESD in the First Affiliated Hospital of Xi'an Jiaotong University from March 2020 to October 2022 were enrolled. The anchor success rate, en bloc resection rate, the anchor time, the procedure time, and the complication incidence were observed and analyzed.Results:Among the 36 patients, there were 9 lesions in stomach, 2 in duodenum, 6 in cecum and 19 in colorectum. Thirty-five (97.2%) patients successfully underwent magnetic anchor, and en bloc resection of lesions were completed. No adverse events such as bleeding or perforation occurred. The anchor time and procedure time was 4.0 (2.0-9.5) min and 36 (16-82) min, respectively.Conclusion:MAG-ESD is feasible and effective for gastrointestinal tumors at different sites, with a high anchor success rate and en bloc resection rate, and shorter operation time, especially for difficult submucosal dissection.