AIM: To clone P1 and P3 promoters of the human insulin-like growth factor Ⅱ(IGF-Ⅱ) gene. METHODS: According to the complete DNA sequence of IGF-Ⅱ gene, the nested primer PCR was performed for amplifying P1 and P3 promoter fragments of the gene from human L-02 cell line.These PCR products were analyzed by agarose gel electrophoresis, and cloned by using TOPO TA Cloning kit. The positive clones containing P1 and P3 fragments were selected and confirmed by sequencing.RESULTS: The DNA sequences of P1 and P3 promoters cloned were accordant with GenBank data. CONCLUSION: In this study P1 and P3 promoters of the IGF-II gene were cloned successfully.

Objective To investiagate cell apoptosis and expressions of Bax,Bcl-2 andCaspase-3 in gambogic acid-treated colorectal cancer cells. Methods SW480/LOVO colorectal cancer cells were treated by gambogic acid. Cell Counting Kit-8 assay (CCK-8) was used to test cell proliferation. Microscopy was used to check the morphological changes. Immunofluorescence staining technique was used to detect cell apoptosis. Expressions of Bax,Bcl-2 and Caspase-3 protein were detected by Western blot assay. Results Gambogic acid inhibited the proliferation of SW480/LOVO in a dose and time-dependent manner. Gambogic acid could induce cell apoptosis. Gambogic acid increased expressions of Caspase-3 and Bax, increased the ratio of Bax/Bcl-2, and decreased Bcl-2 protein expression. Conclusion Gambogic acid can inhibit proliferation and induce apoptosis of SW480LOVO cells, with the mechanism of up-regulation of Bax/Bcl-2 and activation of Caspase-3.

Objective To investigate the expression of circulating tumor cells (CTC) in peripheral blood of patients with different stages of colorectal cancer (CRC), and to evaluate its significance in early diagnosis of colorectal cancer metastasis. Methods Sixty patients with CRC (42 inⅡ-Ⅲstage and 18 in IV stage ) and 30 patients with benign rectal disease were recruited from January 2014 to December 2015. The CTC in peripheral blood was purified with Immunomagnetic Separation Technologie, and detected by immunofluorescence in situ hybridization (imFISH). The serum levels of CEA were detected by electrochemiluminescence method meanwhile. The correlation between CTC and CEA was analyzed. Results CTC positive rates in CRC patients were significantly higher than those in benign rectal disease controls. CTC positive rates inⅣstage were significantly higher than those inⅡ-Ⅲ stage. The expression of CTC was significantly correlated with CEA (r = 0.6652, P < 0.01). Conclusions The expression of CTC in CRC patients is significantly higher than that in benign rectal disease control group. It is closely related to clinical stages. Detection of peripheral blood CTC has important clinical significance in the early diagnosis of CRC metastasis.

OBJECTIVETo investigate the effect and its possible mechanism of interleukin-17 (IL-17) on invasion and metastasis of human colon cancer cells.

METHODSIL-17 was added into the culture media of human colon cancer cells SW480 and LOVO. Cells were divided into 4 groups: SW480 control group (SW480 cells), LOVO control group (LOVO cells), SW480 experiment group (50 μg/L IL-17+SW480 cells), and LOVO experiment group (50 μg/L IL-17+LOVO cells). Cell growth was measured by CCK-8 assay. The proliferation rate(%)=[(Aexperiment group-Ablank)/(Acontrol group-Ablank)]×100%). The ability of cell invasion and migration was measured by transwell assay. Real time-PCR was used to detect mRNA expression of VEGF and MMP-9. Western blot was performed to detect protein expression of STAT3, p-STAT3, VEGF and MMP-9. Enzyme-linked immunosorbent assay (ELISA) was applied to measure the protein content of VEGF and MMP-9 in the supernatant.

RESULTSAfter cultivation for 24, 48 and 72 hours, CCK-8 assay revealed that the proliferation rate of SW480 was 1.18%±0.07%, 1.42%±0.09%, and 1.62%±0.08%; the proliferation rate of LOVO was 1.13%±0.02%, 1.32%±0.05% and 1.73%±0.02% in experiment group. Transwell experiments showed that after cultivation with IL-17 for 24 hours, number of invasion cell in experimental groups (SW480: 34.00±0.45, LOVO: 41.60±0.51) was higher as compared to corresponding control groups (SW480: 4.53±0.14; LOVO: 3.67±0.33) with significant differences (SW480: t=-76.026, P=0.001; LOVO: t=-81.580, P=0.005). The number of migration cell in experimental groups (SW480: 36.40±0.51, LOVO: 46.40±0.68) was higher as compared to corresponding control groups (SW480: 7.83±0.69; LOVO: 6.67±0.48) with significant differences (SW480: t=-51.542, P=0.003; LOVO: t=-49.265, P=0.005). Real-time PCR results revealed that after cultivation with IL-17 for 24 hours, VEGF and MMP-9 mRNA relative expression levels in experimental groups (SW480: VEGF:1.53±0.12, MMP-9: 2.44±0.23; LOVO: VEGF: 2.96±0.35, MMP-9: 3.38±0.55) were higher than those in control groups (both 1) with significant differences (VEGF: t=3.799, P=0.043; MMP-9: t=5.254, P=0.039). Western blot illustrated that after cultivation with IL-17 for 24 hours, STAT3, p-STAT3, VEGF and MMP-9 proteins relative expression levels in experimental groups were significantly higher that those in control groups (SW480:STAT3: t=3.233, P=0.023; p-STAT3: t=3.954, P=0.032; VEGF: t=3.201, P=0.025; MMP-9: t=3.154, P=0.029; LOVO: STAT3: t=3.788, P=0.012; p-STAT3: t=2.662, P=0.040; VEGF: t=4.118, P=0.035; MMP-9: t=4.268, P=0.030). ELISA indicated that content of VEGF and MMP-9 in the supernatant of experimental groups (SW480: VEGF 5 491.41±63.22, MMP-9: 21.43±1.35. LOVO: VEGF: 8 631.46±129.59, MMP-9: 178.32±3.20) were higher than those in control groups (SW480: VEGF:4 456.32±87.56, MMP-9:18.57±2.44. LOVO: VEGF: 8 122.38±108.66, MMP-9: 163.22±6.89) with significant differences (SW480: VEGF: t=6.993, P=0.037; MMP-9: t=5.587, P=0.040. LOVO: VEGF: t=7.013, P=0.044; MMP-9: t=6.762, P=0.043).

CONCLUSIONIL-17 may be able to activate STAT3 signal transduction pathway in vitro through up-regulation of VEGF and MMP-9 expression, thereby enhancing the invasion and migration of colon cancer SW480 and LOVO cells.

Cell Cycle ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Colonic Neoplasms ; pathology ; Humans ; Interleukin-17 ; pharmacology ; Matrix Metalloproteinase 9 ; metabolism ; Neoplasm Invasiveness ; Real-Time Polymerase Chain Reaction ; STAT3 Transcription Factor ; metabolism ; Signal Transduction ; Up-Regulation ; Vascular Endothelial Growth Factor A ; metabolism

Objective To explore the change and clinical significance of the serum carcinoembryonic antigen (CEA)and carbohydrate antigen 724(CA724)in patients with rectal cancer before and after neoadjuvant ra-diotherapy and chemotherapy.Methods The serum levels of CEA and CA724 of 30 patients with rectal carci-noma were detected by electrochemiluminescence method and were compared with those in 30 healthy people. Results The serum levels of CEA and CA724 in rectal carcinoma patients before neoadjuvant radiotherapy and chemotherapy were significantly higher than those in the healthy people,the difference was statistically significant(P<0.05).The serum levels in complete remission and partial remission patients after eight-week neoadjuvant radiotherapy and chemotherapy were significantly lower than those before treatment,the differ-ence was statistically significant(P<0.05),and the reduction levels were obviously higher than those in the stable group,the difference was statistically significant(P< 0.05).The CEA and CA724 levels in the stable treatment patients were also lower than those of before neoadjuvant radiotherapy and chemotherapy,the difference was statistically significant(P<0.05).The serum levels in the progression group were higher than those of before treatment(P<0.05).The serum levels of CEA and CA724 before neoadjuvant radiotherapy and chemotherapy existed a positive correlation(r=0.862,P=0.000).Conclusion The serum levels of CEA and CA724 in rectal cancer are highly expressed,suggesting that both of them are closely related to the occur-rence and development of rectal carcinoma,the measurement of the serum level changes of CEA and CA724 in patients with rectal cancer before and after treatment contributes to estimate the efficacy of neoadjuvant radio-therapy and chemotherapy and tumor progression.

Objective To investigate the antimicrobial resistance in the A cinetobacter baumannii strains in different parts of China during 2012 .Methods A total of 8 739 clinical isolates of Acinetobacter were collected from 13 general hospitals and two children’s hospitals ,of which most were A . baumannii (89 .6% , 7 827/8 739 ) . Antimicrobial susceptibility testing was carried out by means of Kirby-Bauer method according to the unified protocol . The susceptibility testing data were analyzed by WHONET 5 .6 software according to CLSI 2013 breakpoints .Results Majority (85 .4% ) of the Acinetobacter strains were isolated from inpatients .The remaining 14 .6% were from outpatients and emergency room patients .Of the 7 827 strains of A .baumannii , 10 .9% ,35 .2% ,35 .7% and 43 .4% were resistant to tigecycline ,minocycline ,cefoperazone-sulbactam and amikacin , respectively .The percentage of A .baumannii resistant to imipenem and meropenem was 63 .5% and 68 .2% ,respectively . The antimicrobial resistant pattern varied in different hospitals . The resistance of A . baumannii varied between different clinical departments .A number of pandrug resistant (PDR) (20 .0% ,1 567/7 827) and multidrug-resistant (MDR) (45 .0% , 3 521/7 827 ) A . baumannii were identified . Conclusions A . baumannii is the most popular pathogenic bacteria among Acinetobacter .The antibiotic resistance of A .baumannii is still increasing .Cefoperazone-sulbactam and minocycline has good in vitro antibacterial activity against A .baumannii .The antibiotic resistance of A .baumannii varies greatly with hospital and department .

Objective To investigate the distribution and changing resistance proifle ofSalmonella isolates in hospitals across China during the period from January 2005 to December 2014.Methods Seventeen general hospitals and two children’s hospitals were involved in this program. Antimicrobial susceptibility testing was carried out by means of a unified protocol using Kirby-Bauer method or MIC determination. The results were analyzed according to CLSI 2014 breakpoints.Results The proportion ofSalmonella isolates increased with time from 0.2% in 2005 to 0.7% in 2014. A total of 3 478Salmonella strains were collected from 19 hospitals. The proportion ofSalmonella typhimurium andSalmonella enteritidis was 27.4% and 24.4%, respectively. During the 10-year period, theSalmonella strains showed highest resistance rate to ampicillin (33.3%-64.8%), but low resistance to cefoperazone-sulbactam (0-5.3%) and ciprofloxacin (2.4%-14.3%).S. typhimurium showed higher resistance rate thanS. typhi,S. paratyphi andS. enteritidis. About 76.8% and 50.5% ofS. typhimurium were resistant to ampicillin and trimethoprim-sulfamethoxazole. The average prevalence of multi-drug resistantSalmonellawas 3.9% in the ten-year period, the highest (7.5%) was in 2005, the lowest (1.5%) in 2013.Conclusions During the period from 2004 to 2015, majority of theSalmonella isolates in hospitals across China wasS. typhimurium andS. enteritidis. Ampicillin and trimethoprim-sulfamethoxazole are no longer appropriate for empirical treatment ofS. typhimurium infection due to high resistance rate.Salmonella isolates are relatively more susceptible to third-generation cephalosporins and quinolones. Ongoing monitoring is necessary to identify multi-drug resistant strains ofSalmonella.