Objective· To identify the effect and potential mechanism of gambogic acid (GA) on natural killer/T-cell lymphoma (NK/TCL) cell lines.Methods · SNK-1,SNK-6 and SNT-8 were incubated with various concentrations of GA for 24 h,and cell viability was detected with CCK-8 assay.Cell apoptosis was examined by Annexin V-FITC/PI staining assay.Levels of proteins regulating cell apoptosis and phosphorylation levels of proteins in key signaling pathways were detected by Western blotting.Results · GA showed a potent effect on reduction of cell viability of NK/fCL cell lines in CCK-8 assay.GA increased the percentages of Annexin V positive cells and induced activation of caspase-3 and caspase-9,cleavage of PARP as well as the reduction of Bcl-xl.GA also inhibited the phosphorylation levels of STAT3 in SNK-1 and SNT-8,and ERK1/2 in SNK-1 and SNK-6 significantly.Conclusion· GA induces cell apoptosis in NK/TCL cell lines SNK-1,SNK-6 and SNT-8.Anti-apoptosis protein Bcl-xl and signaling pathway JAKs/STATs and MEK/MAPK might be involved in this process.

Objective To explore the expression of miRNA-181a (miR-181a) in patients with multiple myeloma (MM) and its effect on biological features of MM cells. Methods CD138+cells of bone marrow from 25 MM patients and 10 patients with hematological non-malignancies were purified by using immunomagnetic separation, and the expression of miR-181a in CD138+cells and MM cell lines including RPMI 8226, H929 and U266 were detected by real-time quantitative PCR. The effects of down-regulation and up-regulation of miR-181a expression on the biological characteristics of MM cells were studied with miR-181a antagomir and agomir. Results Compared with patients with hematological non-malignant diseases, the expression of miR-181a in CD138+ cells was upregulated in MM patients. Compared with CD138+ cells in hematological non-malignancies, high expressions of miR-181a were observed in RPMI 8226 and U266 myeloma cell line, while low expressions of miR-181a were observed in H929 cells. Down-regulation of miR-181a with 100 nmol/L miR-181a antagomir could inhibit the proliferation of U266 cells at 24,48 and 72 h [(67.1 ± 3.3) %vs. (50.5 ± 4.1) %, (71.5 ± 3.6) % vs. (52.3 ± 2.2) %, (78.1 ± 5.4) % vs. (69.5 ± 4.3) %, P < 0.05 respectively], whereas up-regulation of miR-181a with 100 nmol/L miR-181a agomir could significantly promote the proliferation of H929 cells at 24 h and 48 h [(21.2 ± 2.4) %vs. (38.5 ± 3.6) %, ( 61.3 ± 5.4) %vs. (82.2 ±6.9)%, P<0.01 respectively]. Cell cycle analysis showed that miR-181a antagomir made U266 cell cycle arrest in the G0/G1 phase. Meanwhile, susceptibility test results indicated that the apoptosis of U266 cells induced by doxorubicin, paclitaxel and 5-fluorouracil was increased when the proliferation of miR-181a expression was down-regulated with miR-181a antagomir. In migration assay, the data showed that down-regulation of miR-181a with miR-181a antagomir could inhibit the migration of U266 cells, and the proportion of migrated cells in the experimental group (62 ± 10) %was lower than that in the control group (89 ± 12) %(P< 0.05), whereas up-regulation of miR-181a with miR-181a agomir could improve the migration of H929 cells, and the proportion of migrated cells in the experimental group (242 ± 9) % was higher than that in the control group (98 ± 8)%(P<0.01). Conclusions The high expression of miR-181a expressed highly by MM cells may promote the proliferation, migration and drug resistance of myeloma cells, indicating that miR-181a could be an important prognostic biomarker candidate, and the application of gene silencing may improve the prognosis of MM.

Objective:To put forward relevant policy recommendations for strengthening the quality control of clinical trial drug production in China. Methods:The objective differences of clinical trials and marketing drugs in the production and management as-pects were in-depth analyzed, and the lessons in clinical trial drug production quality supervision and management experience were drawn from the FDA and the European Union EMA. Results and Conclusion:Based on the particularity of clinical trial drug produc-tion management,it is suggested to formulate the administrative rules and technical standards for the quality control of clinical trial drug production and amend relevant laws and regulations timely so as to achieve the purpose of strengthening the quality supervision of drug production in clinical trials.

Objective To investigate the sensitization pattern of Blomia tropicalis in patients with aller-gic respiratory disease and the association between Blomia tropicalis with allergic rhinitis and / or asthma. Meth-ods 330 allergic rhinitis and / or asthma patients sensitized with mites were selected in Guangzhou,and sIgE lev-el in the serum of all recruited patients of Blomia tropical,Dermatophagoide pteronyssinus and Dermatophagoide fa-rina was measured. Results The sensitization rate of Blomia tropicalis was significantly lower than that of Derma-tophagoide pteronyssinus and Dermatophagoide farina(P < 0.001,both). 80.0% Patients were sensitized with both three mites,and only 0.3% patients were of monosensitization to Blomia tropicalis. The sIgE of these three mites were significantly positive correlated with each other(P < 0.001),Dermatophagoide pteronyssinus and Derma-tophagoide farina have strong correlation(r = 0.906),Blomia tropicalis has moderate correlation with Dermatopha-goide pteronyssinus and Dermatophagoide farina(r = 0.540 and r = 0.512,respectively). With the increase of Blomia tropicalis sIgE,the severe sensitization rate(sIgE level:class 5~6)of patients sensitized combined Der-matophagoide pteronyssinus or Dermatophagoide farina significantly increased(P < 0.001). The sIgE level of Der-matophagoide pteronyssinus and Dermatophagoide farina in allergic asthma and rhinitis patients were significantly higher than that of patients with rhinitis or asthma alone(P < 0.001),and the sIgE level of Blomia tropicalis in pa-tients with both allergic asthma and rhinitis or with asthma alone were also significantly higher than that of patients with rhinitis alone(P = 0.006 and P = 0.020,respectively). Conclusion The Blomia tropicalis sensitized pa-tients usually sensitize together with Dermatophagoide pteronyssinus or Dermatophagoide farina,and the degree of Blomia tropicalis sensitized in asthma patients are higher.

Objective:To analyze the relationship between perineural invasion and other clinicopathological factors and its effect on the prognosis of gastric cancer.Methods:The clinicopathological data of 665 patients with gastric cancer were retrospectively analyzed. According to the presence of perineural invasion, the patients were divided into perineural invasion positive group and perineural invasion negative group. The relationship between perineural invasion and other clinicopathological factors and its effect on the prognosis of gastric cancer were analyzed. After eliminating the potential confusion bias between the two groups by propensity score matching (PSM) , the differences of 5-year cumulative survival rate between the two groups of gastric cancer patients were compared.Results:The incidence of perineural invasion was 17.0% (113 cases) . The binary logistic regression analysis showed that the depth of tumor invasion and vascular tumor thrombus were independent factors influencing the occurrence of gastric cancer perineural invasion (all P<0.001) . Univariate analysis showed that age (>60 years) , tumor diameter (>4 cm) , borrmann classification, depth of invasion, lymph node metastasis, TNM stage, degree of differentiation, vascular tumor thrombus, perineural invasion, tumor nodule, tumor site, resection site, and surgical operation were the influencing factors for the prognosis of patients with gastric cancer ( P<0.05) , but multivariate analysis showed that age (>60 years) , tumor diameter (>4cm) , depth of invasion, lymph node metastasis, and positive vascular tumor thrombi were independent risk factors affecting the prognosis of gastric cancer patients ( P<0.05) .However, perineural invasion cannot be an independent factor influencing the poor prognosis of gastric cancer in a multivariate analysis. Survival analysis was performed after propensity matching scores, and it was found that there was no statistically significant difference in the five-year survival rate between the perineural invasion positive group and the perineural invasion negative group (34.6% vs 43.0%; χ2=1.713; P=0.191) ,and there was no significant difference in the survival curve analysis between the two. Conclusion:Most patients with gastric cancer of perineural invasion have poor prognosis, but perineural invasion cannot be an independent prognostic factor for the prognosis of gastric cancer.

Objective:To study the effect of enhanced recovery after surgery (ERAS) on intestinal function and gut microbiota changes in patients who underwent laparoscopic gastrectomy.Methods:From Aug. 2018 to Dec. 2019, 80 patients who underwent laparoscopic radical D2 gastrectomy for gastric cancer in the first Department of Gastrointestinal Surgery of Yantai Yuhuangding Hospital were selected. According to whether it adopts ERAS treatment or not, patients were divided into 2 groups (n=40) : ERAS group and traditional perioperative treatment group. The time of postoperative bowel sounds, the time of first exhaust and defecation, the proportion of antibiotic-related diarrhea and surgical site infection (SSI) were recorded. Stools were collected before operation, first time after operation, 1, 2 weeks and 1 month after operation. 16S rRNA sequencing method was used to identify the diversity and species of gut microbiota. The diversity index of intestinal flora in the perioperative period and changes in the proportion of probiotics (bifidobacterium and lactobacillus) were compared.Results:The appearance time of bowel sounds, the first exhaust and defecation time [ (16.25±6.41) h, (23.95±6.02) h, (34.95±9.34) h] in ERAS group were significantly earlier than those in the traditional treatment group [ (22.3±6.49) h, (28.45±7.12) h, (48.1±15.64) h], and the difference was statistically significant ( P<0.05) . The incidence of antibiotic-related diarrhea was higher in the traditional treatment group (3/40) than in ERAS group (1/40) , but the difference was not statistically significant ( P>0.05) . The ratio of postoperative SSI was slightly higher in ERAS group, but the difference was not statistically significant ( P>0.05) . In the perioperative period, the intestinal flora diversity index (Chao1 and Shannon index) and the proportion of probiotics (lactobacillus acidophilus and bifidobacterium) were not significantly different between the two groups before surgery ( P>0.05) ; while at the first time, one week, 2 weeks after the operation, and 1 month after the operation, ERAS group was higher than the traditional group ( P<0.05) ; and at each postoperative time point, the traditional group decreased significantly than the ERAS group. The first time decrease was the largest, ( P<0.05) ; With the passage of time after operation, the diversity of intestinal flora and the proportion of probiotics gradually recovered. By 1 month after operation, the two groups did not return to the preoperative gut microbiota diversity state or proportion. Conclusion:The concept of enhanced recovery after surgery (ERAS) promotes the recovery of intestinal function in patients with gastric cancer, does not reduce the proportion of antibiotic-associated diarrhea (AAD) or surgical site infections (SSI) , and maintains the diversity of gut microbiota balance and stability.

Objective:To analyze the efficacy and safety of cranial approach priority, counterclockwise sequential comple mesocolic excision in laparoscopic right hemicolectomy.Methods:From Jan. 2020 to Dec. 2020, 30 patients with right colon cancer in Department of Gastrointestinal Surgery were retrospectively analyzed. Laparoscopic radical right hemicolectomy was performed via the approach of complete mesocolic excision. The general clinicopathological data of the patients, perioperative data such as operation time, intraoperative blood loss, number of cases of hemorrhage caused by Henle trunk and subordinate branch injury, whether or not converted to open surgery, postoperative pathological data (TNM staging, total number of dissected lymph nodes and the number of metastatic lymph nodes) , postoperative recovery (exhaust time, the time of fluid intake, drainage tube removal and hospital stay) , and complications (such as bleeding, anastomotic leakage, secondary surgery, lymphatic leakage, pulmonary infection, abdominal infection, incision infection, etc) were recorded. Follow-up was performed by telephone or outpatient in 1 year after surgery.Results:The total operation time was (197.80±31.20) minutes, ranging from 150 to 275 minutes, and the intraoperative blood loss was (58.33±30.30) ml, ranging from 10 to 100 ml. There were no cases of intraoperative Henle stem and branch injury bleeding or conversion to open surgery. Postoperative exhaust time was (2.97±0.67) d, ranging from 2 to 4d; postoperative fluid intake time was (3.67±0.76) d, ranging from 3 to 5d; postoperative drainage tube removal time was (6.60±4.00) d, ranging from 4 to 25 days; postoperative hospital stay was (7.87±3.94) days, ranging from 5 to 26 days. pTNM staging: 9 cases of stage I, 5 cases of stage IIA, 1 case of stage IIB, 6 cases of stage IIIA, 4 cases of stage IIIB, and 5 cases of stage IIIC. The total number of lymph nodes dissected was (29.50±8.18) , ranging from 19 to 51; the number of metastatic lymph nodes was (1.40±1.77) , ranging from 0 to 6. Postoperative complications included incision infection in 1 case, anastomotic leakage in 1 case, lymphatic leakage in 2 cases, and lung infection in 1 case. No tumor recurrence or metastasis was found during follow-up, and no patient died.Conclusion:Cranial approach priority, counterclockwise sequential complete mesocolic excision is safe and effective in laparoscopic right hemicolectomy.

Objective: To identify the expression pattern of TIM-3 in natural killer/T-cell lymphoma (NK/TCL) cell lines, and to investigate the effect and mechanism of its ligand galectin-9 (GAL-9) inducing apoptosis of NK/TCL cell lines. Methods: Expression of TIM-3 in NK cell of peripheral blood from healthy donors and NK/TCL cell lines (SNK-1、SNK-6、SNT-8) was detected by Western blotting. After being treated with rhGAL-9 at various concentrations for 24h, the cell proliferation ability was analyzed with CCK-8 assay. Apoptosis ratio of the cells was determined by flow cytometry. Expressions of caspase-3, PARP and their cleavages were detected by Western blotting; moreover, phosphorylation levels of proteins in MAPK signaling pathway were also detected by Western blotting. Results: The expression of TIM-3 in SNK-1, SNK-6 and SNT-8 cell lines was significantly higher than that of NK cells from healthy donors (P<0.05). CCK-8 result showed that rhGAL-9 obviously inhibited the proliferation of NK/TCL cell lines in a concentration dependent manner. Flow cytometry showed that rhGAL-9 induced the apoptosis of NK/TCLcells; and Western blotting proved that the expression of cleaved caspase-3, cleaved-PARP, and p-JNK in MAPK signaling pathway were significantly elevated. Conclusion: TIM-3 was over-expressed in NK/TCL cell lines, and its ligand galectin-9 induced cell apoptosis probably through the activation of JNK kinase pathways.