ObjectiveTo explore the tumor igenic property of side population cells (SP) from human gallbladder carcinoma cell line GBC-SD. Methods SP and non-SP cells were isolated from GBC-SD staining with Hoechst33342 dye by fluorescence-activated cell sorting (FACS). The soft agar clonal assay and xenograft assay were performed to characterize tumorigenic property of side population cells in vitro and in vivo, respectively. The percentage of SP cells was analyzed by FACS in 5 hu man gallbladder carcinoma specimens. ResultsThe percentage of SP cells accounted for approximately 0.87 ％ of GBC-SD cells. The clone-formed rates of SP was more frequent than that of non-SP cells (14.74％ ± 3.53％ vs 5.17％ ± 1.05％), there was statistically significant difference (t =2.75,P＜0. 05). SP cells could generate tumors with as few as 5 × 103 cells (four of seven animals), whereas at least 1 × 105 non-SP cells were needed to form a tumor (one of seven animals). Re-analysis of SPderived tumors by FACS showed that SP cells under in vivo conditions also have the capacity to regenerate the SP and non-SP fractions. Besides, analysis of Hoechst33342 revealed s small fraction of SP cells, ranging from 0. 27％ to 2.3％ in gallbladder carcinoma specimens. ConclusionSP cells from GBC-SD are highly tumorigenic similar as the cancer stem cells.

Objective To investigate the drug resistance of side population cells in human gallbladder cancer cell line GBC-SD and explore its mechanism. Methods Drug sensitivity assays of 5chemotherapeutic agents were performed on side population cells (SP) and non-SP cells of GBC-SD.GBC-SD was cultured and then treated with the chemotherapeutic agent gemcitabine. The frequency of SP by FACS was measured. RT-PCR and Western blotting were used to detect the expression of AB-CG2 in both the SP and the corresponding non-SP subsets. Results After 1 d treatment with 4 chemotherapeutic agents (gemcitabine, cisplatin, 5-fluorouracil and mitoxantrone) in IC50 concentration to GBC-SD cell line, the reproductive ability of SP was higher than that of non-SP (P<0.05). However, statistical significance was not achieved when compared with epirubicin (P>0.05). The percentage of SP in GBC-SD treated with chemotherapeutic agent gemcitabine after 3 weeks was sharply elevated by FACS (8.02% ±0.13% vs 0.62% ±0.08%, P<0.05), and the expression of ABCG2mRNA and protein were increased in SP as compared with non-SP. Conclusion SP from human gallbladder cancer cell line GBC-SD, like stem cell, showed a heighten resistance to drugs. Increased expression of ABCG2 was largely responsible for the multi-drug resistance.

Objective To evaluate a modified technique for digestive tract reconstruction after pancreaticoduodenectomy(PD).Methods 171 admitted patients were enrolled from January 2012 to January 2014 at our department.According to the preoperative CT scan and intraoperative exploration,pancreaticogastrostomy was performed in cases of soft pancreas texture,while pancreaticojejunostomy was performed in fibrotic pancreas after PD.Bypassed biliary-pancreatic reconstruction were applied on all cases.Results For the digestive tract reconstruction after PD,92 patients underwent pancreaticogastrostomy,79 patients underwent pancreaticojejunostomy.The median time for the surgery was 240.0 minutes (ranging from 186 to 414 min).Operative mortality was zero,and morbidity was 18.1％ (n =31),including hemorrhage (n =4),biliary fistula (n =3),pulmonary infection (n =2),adipose liquefaction and operative incision infection (n =0),delayed gastric emptying (DGE) (n =6),abdominal abscess (n =4).Fout patients developed a pancreatic fistula (type A in 2,type B in 2).Conclusions Modified pancreaticogastrostomy,pancreaticojejunostomy and biliary-pancreatic bypass is safe for digestive tract reconstruction after pancreaticoduodenectomy.

Objective: To investigate the expression of blood vessel epicardial substance (BVES) in gallbladder carcinoma and the effects of BVES overexpression on the proliferation and invasion of gallbladder cancer GBC-SD cells. Methods: Immunohistochemistry was used to detect the expression of BVES in gallbladder carcinoma tissues and the para-cancerous tissues. The BVES overexpressed lentivirus was infected into gallbladder cancer GBC-SD cells, while the GBC-SD cells infected with empty carrier lentivirus were used as the negative control, and the uninfected GBC-SD cells were used as the blank control. The expression of BVES protein in GBC-SD cells was detected by Western blotting. The proliferation and invasion of GBC-SD cells in different groups were detected by colony formation assay and Transwell assay, respectively. Results: The positive expression rate of BVES in the gallbladder carcinoma tissues was lower than that in the para-cancerous tissues (P < 0.01). The expression level of BVES protein in GBC-SD cells infected with BVES overexpressed lentivirus was higher than that in the negative control group or the blank control group (both P < 0.05). The proliferation and invasion abilities of GBC-SD cells after the infection of BVES overexpressed lentivirus were significantly decreased (both P < 0.05). Conclusion: The positive expression rate of BVES is low in gallbladder carcinoma. Overexpression of BVES can inhibit the proliferation and invasion of gallbladder carcinoma GBC-SD cells.

Mounting evidence has shown that side population (SP) cells are enriched for cancer stem cells (CSCs) responsible for cancer malignancy. In this study, SP technology was used to isolate a small subpopulation of SP cells in human gallbladder cancer cell line GBC-SD, and SP cells which had superior potential for proliferation in vitro and tumorigenesis in vivo were identified. Importantly, the abundance of GBC-SD SP cells was increased by a transforming growth factor-β (TGF-β)-induced epithelial-mesenchymal transition (EMT), and this effect was accompanied with a strong up-regulation of ABCG2 mRNA expression, and a decreased sensitivity to mitoxantrone. SP cells were restored upon the removal of TGF-β and the reversion of the cells to an epithelial phenotype, and smad3-specific siRNA reduced SP abundance in response to TGF-β. In conclusion, TGF-β-induced EMT by smad-dependent signaling pathway promotes cancer development and anti-cancer drug resistant phenotype by augmenting the abundance of GBC-SD SP cells, and a better understanding of mechanisms involved in TGF-β-induced EMT may provide a novel strategy for preventing cancer progression.

Objectives To isolate cancer stem cells (CSCs) in pancreatic cancer cell lines PANC1 and ASPC-1 with serum-free medium( SFM ), and to detect the expression of miR-590-3p in CSCs. Methods PANC1 and ASPC-1 cells was cultured in serum-free medium. The monoclonal formation, differentiation and cell cycle, half inhibitory concentration ( IC50 ), and the expression of the surface markers CD24 + , CD44 + were detected. qRT-PCR was used to detect the expression of miR-590-3p. Results After SFM culture, (0.94 ±0.53 ) ％ of ASPC-1 and (0.57 + 0. 12 ) ％ PANC1 survived, and they formed spheres, and could continuously passage in vitro. Cell spheres differentiation recurred when serum was supplemented in SFM. The G0/G1 stage proportion, CD24+ , CD44 + , CD24+ CD44+ cells proportion, IC50 in ASPC-1 cell were (75.3 ± 5.4)％,0.96％ ～ 2.01％, 27.52％ ～ 34.47％, 0.35％ ～ 0.44％ and (224.37 ± 5.71 ) μg/ml, which were significantly higher than that those in parent cell [ (43.7 ± 3.8 ) ％, 0. 38％ ～ 0.42％, 17.65％ ～ 18.25％,0.05％ ～0.08％, (11.43 ±2.10)μg/ml, P＜0.05]. The G0/G1 stage proportion, CD24+ ,CD44+ ,CD24 +CD44 + cells proportion, IC50 in PANC 1 cell were ( 80. 1 ± 4.7) ％, 5.31％ ～ 9.84％, 72.05％ ～ 93.06％,4.91％ ～5.21％, (296.58±4.27) μg/ml, which were significantly higher than that those in parent cell [ (46.1 ±5.3)％, 4.09％ ～4.97％, 47.71％ ～55.66％, 1.48％ ～2.63％, (26.17 ±3.81)μg/ml, P＜0.05]. The expression of miR-590-3p in ASPC-1, PANC1 spheres was 4.67 and 4.52 times higher than the expression in parent cell lines. Conclusions Pancreatic cancer cell spheres can be isolated from ASPC-1, PANC1 by culture with SFM. miR-590-3p is up-regulated and may play an important role in regulating biological characteristics of pancreatic cancer stem cells.

Objective: To establish a method for determination the S-phenylmercapturic acid in urine by dispersive solid-phase extraction using Humic Acid/Fe₃O₄ magnetic nanocomposite as adsorbent. Methods: The 5 ml of urine samples were adjusted to pH 1.0 and extracted by Fe3O4@HA. Then the analytes were separated on EC-C₁₈ capillary column and detected by HPLC-VWD. The S-phenylmercapturic acid was characterized by the retention time and quantified by peak area and external standard method. Results: The standard curves of SPMA showed significant linearity between 0.04~1.00 mg/L (r=0.999 7) . The average recovery was 94.2%~102.4%. The intra-day and inter-day precisions (RSD) were 2.9~6.7% (n=6) and 3.1~7.5% (n=6) respectively. The detect limit of SPMA was 0.012 g/L (S/N=3) . Conclusion This method is simple, rapid, sensitive and accurate. It is applicable for determination of SPMA in the urine of works who were exposed to benzene.

Mounting evidence has shown that side population (SP) cells are enriched for cancer stem cells (CSCs) responsible for cancer malignancy.In this study,SP technology was used to isolate a small subpopulation of SP cells in human gallbladder cancer cell line GBC-SD,and SP cells which had superior potential for proliferation in vitro and tumorigenesis in vivo were identified.Importantly,the abundance of GBC-SD SP cells was increased by a transforming growth factor-β (TGF-β)-induced epithelial-mesenchymal transition (EMT),and this effect was accompanied with a strong up-regulation of ABCG2 mRNA expression,and a decreased sensitivity to mitoxantrone.SP cells were restored upon the removal of TGF-β and the reversion of the cells to an epithelial phenotype,and smad3-specific siRNA reduced SP abundance in response to TGF-β.In conclusion,TGF-β-induced EMT by smad-dependent signaling pathway promotes cancer development and anti-cancer drug resistant phenotype by augmenting the abundance of GBC-SD SP cells,and a better understanding of mechanisms involved in TGF-β-induced EMT may provide a novel strategy for preventing cancer progression.

Lower extremity deep vein thrombosis (DVT) is one of the main complications in patients with traumatic fractures, and for severe patients, the DVT can even affect arterial blood supply, resulting in insufficient limb blood supply. If the thrombus breaks off, pulmonary embolism may occur, with a high mortality. The treatment and rehabilitation strategies of thrombosis in patients with lower extremity fractures have its particularity. DVT in traumatic fractures patients has attracted extensive attention and been largely studied, and the measures for prevention and treatment of DVT are constantly developing. In recent years, a series of thrombosis prevention and treatment guidelines have been updated at home and abroad, but there are still many doubts about the prevention and treatment of DVT in patients with different traumatic fractures. Accordingly, on the basis of summarizing the latest evidence-based medical evidence at home and abroad and the clinical experience of the majority of experts, the authors summarize the clinical treatment and prevention protocols for DVT in patients with traumatic fractures, and make this consensus on the examination and assessment, treatment, prevention and preventive measures for DVT in patients with different fractures so as to provide a practicable approach suitable for China ′s national conditions and improve the prognosis and the life quality of patients.