Many researches show that the prognosis of gastrointestinal stromal tumor (GIST) are relatedtonumerousvariationsofbiomarkerssuchasnumericchromosomalaberrationsor nuclear/mitochondrialmicrosatelliteinstability.Inaddition, thechangesofproto-oncogeneand tumor-suppressor gene are found to influence the prognosis of GIST.The kit and platelet-derived growth factor receptor gene mutation are correlated with GIST progression,metastasis and targeted drug sensitivity,and then influence the prognosis of GIST.

Background and purpose:Several reports demonstrated that the expression of STAT3 has been found to be an oncogene in solid tumors such as head and neck squamous cell carcinoma,esophageal carcinoma and prostate carcinoma.This study was done to explore the effects of X-ray irradiation combined with RNAi against STAT3 on proliferation and apoptosis of human esophageal carcinoma cell line Eca-109.Methods:Three pairs of DNA template coding siRNA were synthesized against STAT3 to reconstruct pRNAT-U6.1-siRNA-STAT3,which was transfected into Eca-109 cells,the positive cell clones were screened with G418.Inhibitory effect of STAT3 mRNA and protein in Eca-109 cells was detected by RT-PCR and Western blot,respectively.The transfected cells were exposed to 0,2,4,6 and 8 Gy X-ray respectively;the survival fraction of Eca-109 cells was analyzed by clone formation assay,and flow cytometry was applied to analyze cell apoptosis at the dose of 4 Gy.Results:pRNAT-U6.1-siRNA-STAT3 was reconstructed and identifi ed as correct by sequencing.RT-PCR and Western blot analyses demonstrated that STAT3-siRNA could obviously reduce the expression of STAT3 mRNA and protein in Eca-109 cells.Clone formation assay and flow cytometry results showed that irradiation at different doses combined with STAT3-siRNA could inhibit the proliferation of Eca-109 cells,irradiation with 4 Gy X-ray could induce apoptosis.Conclusion:X-ray irradiation combined with RNAi against STAT3 could inhibit the proliferation of Eca-109 cells and induce apoptosis.

Objective To explore the effects of X-ray irradiation combined with RNAi against signal transducer and activator of transcription 3(STAT3)on the radiosensitivity of human esophageal carcinoma cells.Methods Human esophageal carcinoma cells of the line Eca-109 were euhured.Three pairs of DNA template aiming at the base sequences of the coding regions 2037-2055,1243-1261,and 455-473 of the STAT3 mRNA were synthesized(siRNAI,siRNA2,and siRNA3),and a negative sequence was synthesized to be used as control.STAT3-siRNA positive recombinant plasmids(pRNAT-U6.1-siRNAI,pRNAT-U6.1-siRNA2, and pRNAT-U6.1-siRNA3), and a STAT3-siRNA negative recombinant plasmid (pRNAT-U6.1-negative)were thus constructed and then transfected into the cultured Eca-109 cells,which were divided into transfection reagent control group,pRNAT-U6.1-siRNAl-3 transfection groups,and pRNAT-U6.1-negative centrel group.The positive eell clones were screened.RT-PCR and Westem blotting were used to detect the STAT3 mRNA and protein expression.The transfected Eca-109 cells were exposed to 0,2,4,6,and 8 Gy of X-rays,respectively,and the survival fraction of the cells was analyzed by clone formation assay.Flow cytometry was applied to analyze the cycle arrest and cell apoptosis 4 Gy post-irradiation.Results Agarose gel electrophoresis confirmed the successful construction of the plasmid pRNAT-U6.1-siRNA.RT-PCR and Western blotting demonstrated that the mRNA and protein expression levels of STAT3 transfected with sTAT3-siRNA3 were both significanfly lower than those of the control groups.At 2-8 Gy, the survival fractions of the siRNA3 group were aU significantly lowered than those of the control group(t＝-0.228--0.051,P<0.05).Flow cytometry showed that the percentage of the cell cycle G0/G1 phase and the apoptosis rate of the siRNA3 group were both significantly higher than those of the control groups at 4 Gy post-irradiation(t＝-13.137-16.350,P<0.01).Conclusions X-ray irradiation combined with RNAi against sTAT3 could inhibit the proliferation of the human esophageal carcinoma cells,induce cell cycle arrest and apoptosis,improve the radiosensitivity in Eta-109 cells.

Objective: To study the cell proliferation, cell cycle and apoptosis of esophageal carcinoma Eca-109 cells treated with RNA interference technique to silence signal transducers and activators of transcription 3 (STAT3) gene. Methods: Three pairs of DNA template coding siRNA specific for human STAT3 gene mRNA were designed and synthesized. The annealed oligonucleotide fragments were subcloned into pRNAT-U6.1/neo plasmid to construct STAT3-siRNA expression vector which was then transfected into Eca-109 cells. The expression of STAT3 mRNA and protein in cancer cells was detected by RT-PCR and Western blot, respectively. The cell proliferation, cell cycle distribution and apoptosis were examined by MTT and flow cytometry. Results: STAT3-siRNA expression vector was successfully constructed and identified by sequencing. The results of RT-PCR and Western blot demonstrated that STAT3 expression in Eca-109 cells transfected with STAT3-siRNA expression vector was significantly higher than that in the control group (P＜ 0.01). MTT showed that after transfection of the siRNA vector into Eca-109 cells, cell proliferation was obviously reduced and the cell growth inhibition ratio in the siRNA3 group was 35.68%, significantly higher than that in the control group (P＜0.01). Flow cytometry results suggested that cell cycle arrest and more apoptosis were observed in the siRNA3 group. Cell cycle was arrested at G_0/G_1 phase, and the rate of apoptosis was 13.26%, much higher than that in the control group (P＜0.01). Conclusion: Silencing STAT3 gene by RNA interference technique can effectively inhibit STAT3 expression, suppress the proliferation of Eca-109 cells, induce cell cycle arrest at G_0/G_1 phase, and promote apoptosis.

Objective To study the safety and efficacy of intraoperative implanting slow-released 5-Fu after hepatectomy in patients with hepatocellular carcinoma (HCC). Methods 59 HCC patients undergoing hepatectomy from January 2008 to January 2009 were divided into two groups:to receive slowrelaesed 5-Fu in treatment group and nothing in control group.Postoperatively no patients in both group took chemotherapy.The serous value of WBC,ALT,AST,TBIL and AFP were measured in all cases on 1 day before and 3 weeks after the operation.During the first six months,AFP was measured and imaging studies were done one month after discharge,and every 3 months since then.CT-guided biopsy was used to confirm the recurrence of HCC. Disease-free survival rate and overall survival rate in two groups calculated respectively 6-,12-,18- and 24-months after the operation. Results The serous value of WBC,ALT,AST,TBIL was not different between treatment group and control group 3 weeks after the operation (separately t =0.801,- 0.854,- 1.948,- 0.503,all P ＞ 0.05).AFP was (361.58 ± 431.06) μg/L,( 17.02 ± 15.55 ) μg/L,(43.61 ± 58.03 ) μg/L in treatment group and (495.50 ± 441.63 ) μg/L,(26.82 ±60.46) μg/L,(127.48 ±229.79) μg/L in control group preoperatively,3 weeks and 6 months after the operation.AFP was lower in treatment group than that in control group ( t =- 2.065,P ＜ 0.05 ).The disease-free survival rate in treatment group was 95.8％,91.7％,79.2％,75.0％ at 6-,12-,18- and 24-months after the operation,in control group it was 94.3％,71.4％,60.0％,48.6％ ( x2 =4.035,P ＜0.05).The overall survival rate was respectively 100％,95.8％,91.7％,83.3％ in treatment group and was 100％,94.3％,77.1％,60.0％ in control group( x2 =3.931,P ＜0.05).Conclusions Intraoperativeimplanting slow-released 5-Fu during the hepatectomy was safe and effective method to reduce the recurrence rate of hepatocellular carcinoma and prolongs postoperative disease free and overall survival in HCC patients undergoing hepatectomy.

Objective To investigate the effect of Huoxuetongluo Decoction on expression of CD54 and polymorphonuclear adhesion during human umbilical vein endothelial cells(HUVEC) anoxia/reoxygenation injury,and explore its possible mechanisms.Methods Rabbits serum containing Huoxuetongluo Decoction was prepared.Resuscitate and culture HUVEC,then establish the anoxia/reoxygenation injury model of HUVEC.The model cells were divided into five groups:normal control,model group and group of high,medium,low dose of Huoxuetongluo Decoction.After dealt seperately,the morphologic change of cells were observed through the microscope,the expression of CD54 and the polymorphonuclear adhesion were determined.Results The group of low,medium and high dose of Huoxuetongluo Decoction inhibited the expression of CD54 and decreased the adhesion between polymorphonuclear and HUVEC,the effects were strengthened with increasing the dose of Huoxuetongluo Decoction.The difference between group of medium,high dose of Huoxuetongluo Decoction and model group had statistical significance(P

Objective To investigate the allergic reaction of Shuanghuanglian Injection(SI)and chlorogenic acid.Meth- ods The sensitization of SI was evaluated by observed the systemic active allergic reaction on guinea pigs.The homogeneous and heterogenic passive skin allergic reactions in rats,mice and guinea pigs were observed to compare the sensitationz of SI and chlorogenic acid.Results The possibility of SI causing allergic reaction was higher than that of chlorogenic acid.Con- clusion It is suggested that the possibility of SI as a compound arising adverse reactions be higher than that of chlorogenic acid.

Objective: To establish the method for determination of taurocholic acid in the Traditional Chinese Medicine Shedanchuanbei Oral Liquid and snake bile. Methods: The sample was prepared as mixed solution containing methanol and KH 2PO 4. The mixed solution was injected into Sep Pak C 18 cartridge for the purpose of sample purity. In this processing, the substances which having strong retain action and could harm analytic column were hold in the Sep Pak C 18 cartridge. The eluting solution that the Sep Pak C 18 cartridge had be over loading for taurocholic acid was used as the test solution. The test solution was measured by RP HPLC. The chromatographic conditions were as followed: Supelcosil LC 8 column(150mm?4.6nm,5?m) as analytic column, detect wavelength at 203nm, and MeOH 0.4%KH 2PO 4 mixed solution(56∶44, V/V ) as mobile phase. The inject volume was 50?L. Results: The linear response range of sodium taurocholate was from 0.0253mg?mL -1 to 0.253mg?mL -1 , and the correlation coefficient was 0.9999. The average recovery rate was 101.3%, RSD was 0.40%( n =6). Conclusion: This method was simple, efficient and suitable to the quality control for Shedanchuanbei Oral Liquid and snake bile.

BACKGROUND: At present, studies on repair of cartilage defect have been focused on tissue engineering technique. Growth factors are one of the most important parts. However, the effect and security of growth factors have not been confirmed. Studies have shown that Weilingxian can maintain and promote the synthesis of proteoglycan, collagen Ⅱof chondrocyte, and it also can promote proliferation of chondrocyte and expression of transforming growth factor (TGF)-β1 mRNA.OBJEGTIVE: To investigate the feasibility of injectable chitosan/β-glycerol phosphate (C/β-GP) encapsulating allograft chondrocytes on the repair of articular cartilage defects and the intervention effect and possible mechanisms of Weilingxian.METHODS: A 0.4-mm defect was established on knee articular cartilage. Expeirmental New Zealand rabbits were randomly divided into three groups: Weilingxian, common culture media, and model groups. In the common culture media group, the samples were treated with C/β-GP and chondrocyte suspension (1 mL); at 2 days after gel injection, Weilingxian or common culture media (1 mL) were respectively given into joint cavity, once a day, for 7 successive days. The samples in the model group were not treated. Gross, histological (HE staining, TB staining), type Ⅱ collagen immunohistochemical, and Wakitani score examinations were performed on 6 and 12 weeks after surgery.RESULTS AND CONCLUSION: Defects of articular surface were well filled in Weilingxian and common culture media groups, and hyaline cartilage-like structure was formed. The surface flatness and degree of integration with surrounding tissue of Weilingxian group was better than common culture media group. Formation of cartilage-like and secretion of cartilage matrix and specificity of collagen type Ⅱ were found in histological slices. Defects in the model group were not repaired, while tissue proliferative degeneration was observed. Integration of.repair tissue with surrounding tissue, histology and amount of type Ⅱ collagen secretion in Weilingxian group were better than common culture media group. Wakitani scores of Weilingxian group and common culture media group were significantly lower than model group (P < 0.01), andscores of Weilingxian group was significantly lower than common culture media group (P<0.05). Injectable chitosan/β-glycerolphosphate gel encapsulating allograft chondrocytes could repair articular cartilage defects, and Weilingxian was able to promote the process of it, this manifested the role like growth factor in tissue engineering technique repairing articular cartilage defects.

BACKGROUND:Studies have shown that Weilingxian can maintain and promote the synthesis of proteoglycan and collagen Ⅱ of chondrocyte,and protect artlcular cartilage and postpone the development of osteoarthdtis by inhibiting the level of intedeukin-1(1L-1)possibly.OBJECTIVE:Based on the previous studies,to observe the effect of Weilingxian on the proliferation of rabbit knee articular chondrocyte and transforming growth factor-β1 mRNA expression,and then to explore the role and possible mechanism of Weilingxian in the treatment of osteoarthdtis.METHODS:Knee cartilage was shredded after harvested from New Zealand white rabbits under sterile conditions,and chondrocytes were isolated and cultured by the way Of enzymatic digestion.After identifying by toluidine blue staining,the third-passage calls in the logarithmic growth phase were cultured in vitro and randomly divided into two groups after adherence.The experimental groups were cultured in DMEM with 0.01,0.05,0.1,0.5,and 1.0 mg/mL Weilingxian,while the control group was given with normal medium alone.Chondrecytes morphology was observed under an inverted phase contrast microscope,and the phenotype was identified by toluidine blue staining;Methyl Thiazolyl Tetrazolium(MTT)assay method was adopted to observe the influenca of Weilingxian with difierent concentrations on the proliferation of chondrocytes,and anti-transcription-polymerase chain-type reaction(RT-PCR)was used to assay the expression changes of transforming growth factor-β1 mRNA.RESULTS AND CONCLUSlON:Primary cultured chondrocyte was round-shaped,and most of It adhered after 24 hours,the appearance was polygonal and irregular-shaped;after passage,cell growth was faster than before,the typical appearance was slabstone-like;long spindle-shaped chondrocytes appeared after four generations;after six generations,most cells showed long spindle-shaped fibroblast-like appearance,the rate of growth also slowed down.Extracellular matrix of chondrocytes was stained to be blue by toluidine blue staining,and the nucleus was dark blue.Different concentrations of Weilingxian could promote the proliferation of chondrocytes,effect of 0.5 mg/mL group was significantly,and the peak of proliferation was on the third day.0.05,0.1,0.5,and 1.0 mg/mL Weilingxian group could promote the expression of transforming growth factor-β1 mRNA.and there was no significant difference between four groups(P＞0.05),but the peak was at 0.5 mg/mL group.Weilingxian can promote proliferation of chondrocyte and transfonlling growth factor-β1 mRNA expression,and these may be one of the possible mechanisms that Weilingxian can work in the treatment of osteoarthritis.