Objective: To isolate and identify tumor-like stem cells from human osteosarcoma cell line U2-OS. Methods: Tumor sphere cells were isolated from osteosarcoma cell line with serum-free culturing techniques. Flow cytometry was used for detecting the positive rates of Stro-1 protein in U2-OS cell line. The proliferation ability of sarcospheres was estimated by MTT assay. Stro-1+ cells were isolated using magnetic activated cell sorting system. RT-PCR was used to detect the transcription of Stro-1 mRNA in Stro-1+ cells. The Stro-1+ cells and Stro-1- cells were inoculated into NOD/SCID mice. Results: The results of flow cytometry showed that the percentage of Stro-1+ cells was 5.67% in U2-OS cells. Stro-1+ cells had capability of proliferation and differentiation. The over-expression of Stro-1 mRNA was observed in Stro-1+ cells. Inoculation of Stro-1+ cells induced osteosarcoma formation in NOD-SCID mice. Conclusion: Osteosarcoma-like stem cells exist in human osteosarcoma cell line U2-OS. They have self-renewal and multi-directional differentiation abilities.

Gambogic acid-loaded polylacticacid nanoparticles (GA-PLA-NPs) were prepared by modified emulsification solvent diffusion. The shape of nanoparticles was observed by transmission electron microscope (TEM).The size distribution and mean diameter were measured by laser particle size analyzer. The entrapment efficiency and content of drug loading were determined by Ultraviolet Spectrophotometer after ultracentrifugation. GA-PLA-NPs release behavior in vitro was carried out. The acute toxicity were carried out to study the security of GA-PLA-NPs. The preparation process adapted to the formulation was as follows: the volume ratio of the aqueous and organic was 2∶1(v/v), the surfactant concentration in aqueous was 0.5%,the drug concentration in organic was 0.1%(w/v), GA∶PLA was 1∶4(w/w). The mean diameter was 51.36nm for the nanoparticles prepared by above conditions.The entrapment efficiency and content of drug loading were 98.87 % and 13.3 %. The release behavior of drug in vitro showed an initial burst effect with subsequently a slower rate stage. The LD50 value of GA-PLA-NPs on mouse was 26.3 mg/kg. The results showed that the GA-PLA-NPs were well prepared with stable quality and high dispersion. PLA-NPs might be used as a new carrier for gambogic acid.

Objective: To investigate the effects of Qingyi Huaji (QYHJ) decoction, a compound traditional Chinese herbal medicine, on tumor inhibition rate and serum levels of interleukin-6 (IL-6), interleukin-8 (IL-8) and tumor necrosis factor-alpha (TNF-alpha) in nude mice with transplanted tumors of human pancreatic cancer. Methods: The tumor-bearing mice model was established by subcutaneously inoculating with xenografts of pancreatic cancer into the right armpit of 40 BALB/c nude mice. After successful modeling, the mice were randomly divided into untreated group (Arabic gum), capecitabine group, low-dose QYHJ decoction group (36 g/kg) and high-dose QYHJ decoction group (72 g/kg), with 10 mice in each group. Citrate buffer solution (containing 5% Arabic gum), capecitabine suspension and QYHJ decoction were administered to four groups by gavage respectively. After 5-week treatment, the concentrations of serum IL-6, IL-8 and TNF-alpha were examined by enzyme-linked immunosorbent assay (ELISA) using blood sample from eye socket. Then the mice were euthanized by cervical dislocation. Tumor weight and the tumor inhibition rate were calculated. Results: Tumor weight in the low-dose QYHJ decoction group decreased significantly as compared with the untreated group (P<0.05). Serum levels of IL-6 and TNF-alpha in low- and high-dose QYHJ groups were extremely significantly lower than those in the untreated group (P<0.01). Serum level of IL-8 in the low-dose QYHJ group was significantly lower than that in the untreated group (P<0.05). Correlation analysis showed that transplanted tumor weight of the mice was linearly positively correlated with serum levels of IL-6, IL-8 or TNF-alpha (P<0.01). Conclusion: Conventional dose of QYHJ decoction is effective in suppressing pancreatic carcinoma in nude mice. The mechanism may be related to down-regulation of serum cytokines such as IL-6, IL-8 and TNF-alpha.

Objective:To compare the immunogenity of the AST artificial antigen with two different carrier proteins, bovine serum albumin(BSA) and keyhole limpet hemocyanin(KLH).Methods: The hapten Astragaloside Ⅳ(AST) was linked to carrier protein BSA or KLH as immunogen,and linked ovalbumin(OVA) as coating antigen by the sodium periodate method.Then the 6-8 weeks BALB/c mice were immunized with AST-BSA or AST-KLH.The titer and specificity of the antiserum were detected by indirect enzyme-linked immunosorbent assay ( iELISA ) and indirect competitive enzyme-linked immunosorbent assay ( icELISA ) respectively.Results:The antiserum of the mice immunized with AST-KLH had better specificity than which immunized with AST-BSA.The IC50 of AST-KLH antiserum was about 5 μg/ml in the detection by icELISA.The high titer of both antiserums was about 1∶51 200 after immunizing mouse.Conclusion:The conjugate of AST-KLH was testified to have a better immunogenity in comparison with AST-BSA,this investigation would be the foundation of the further research of AST immunoassay.

Mycotoxins are secondary metabolites produced by certain genera of toxigenic fungus and frequently oc-cur in food worldwide. Humans and animals can be simultaneously exposed to different mycotoxins through the diet. As most mycotoxins are highly toxic, carcinogenic, teratogenic and mutagenic, they have posed grave health threats to consumers. Determination of mycotoxins and their main metabolites in blood, urine, bile, milk or faeces can serve as biomarkers and can facilitate effective exposure assessment, crucial to estimate mycotoxin related dis-ease risk. According to reason mentioned above, the study of metabolism and evaluations of mycotoxins in biologi-cal fluids have been paid increasing attention since the results may offer valuable indications on the real risk for consumers. Therefore, it is important to develop proper analytical methods for the rapid quantitative and qualita-tive measurement of mycotoxins and key metabolites in vivo. This paper reviewed some biomarkers and their harm to animals and humans, systematically summarized the research progress of analytical methods and prospected the development trends.

Objective To prepare the preparation of ginkgolic acids gel and to study its antibacterial activity in vitro.Methods The gel was prepared with ginkgolic acids as main component and with Carbopol 940 as base material.The content of ginkgolic acids in the gel was determined by high performance liquid chromatography and the minimum inhibitory concentration (MIC) of the gel against staphylocococus aureus and propionibacteria was measured in vitro by means of liquid-doubling dilution.Results The ginkgolic acids gel has inhibitory effects on staphylocococus aureus and propionibacteria to various extents.And the MIC was 31.756 mg?mL-1 for staphylocococus aureus and 0.992 mg?mL-1 for propionibacteria.Conclusion The preparation of the gel is feasiblye and the the properties of the gel were stable.The ginkgolic acids gel possesses good antibacterial effects on staphylocococus aureus and propionibacteria.

ObjectiveTo explore the ultrasound characteristics of coronary artery fistula to improve its prenatal diagnosis.MethodsImaging data for 12 fetuses with coronary artery fistula who underwent fetal echocardiography at our hospital from June 2005 to December 2013 were retrospectively analyzed to summarize the ultrasound characteristics of coronary artery fistula.ResultsOf the 12 fetuses, 7 were confirmed with coronary artery fistula by echocardiography after birth, and the remaining 5 were aborted due to other cardiac abnormalities (since an autopsy was not performed, the diagnosis of coronary artery fistula could not be confirmed). Of the 7 confirmed cases, 1 had left coronary artery-to-right atrium fistula, 3 had left coronary artery-to-right ventricle fistula, and 3 had right coronary artery-to-right ventricle fistula; 1 had left coronary artery-to-right ventricle fistula with muscular ventricular septal defect, and 6 had simple coronary fistula. Two-dimensional ultrasound examination revealed that coronary arteries were displayed in all cases, and the diameter ranged from 1.8 to 4.0 mm. Color Doppler ultrasound clearly showed the fistula size, course and the situation fistulas drained into heart chambers. Diastolic regurgitation signal could be detected in the ascending aorta and aortic arch.ConclusionCoronary artery fistula can be diagnosed by color Doppler ultrasound combined with spectral Doppler ultrasound when the fetal coronary artery expands.

The isolated 24 strains-producing hydantoinase & carbamoylase were first identified by Biolog microbial identification system and 16S rDNA sequence analysis.The results suggested that the hydantoinase & carbamoyalse-producing bacteria belonged to Bacillus,Geobacillus,Brevibacillus,Aneurinibacillus,Microbacterium,Pseudomonas,Kurthia and Empedobacter,and so on.Especially,Kurthia and Empedobacter were new hydantoinase & carbamoylase-producing genera.Furthuremore,it was found that D-hydatoinase & carbamoyalse-producing bacteria belonged to Pseudomonas and Agrobacterium,while most of L-hydantoinase & carbamoyalse-producing bacterial belonged to Bacillus,Geobacillus and Microbacterium.The distribution feature of D-hydantoinase & carbamoyalse-producing bacteria and L-hydantoinase & carbamoyalse-producing bacteria showed some genera tendency.This research work will provide the biomaterial of different hydantoinase and carbamoylase and contribute to study the structure and function,molecular evolution of the two enzymes.

Objective To study the expression of cyclooxypenase-2(COX-2),prognosis of patients and the correlation between COX-2 and CD_(105)in osteosarcoma.Methods The expression of COX-2 and CD_(105)in human osteosarcoma was detected with immunohistochemistry.Then to count the microvessl density (MVD)were marked with CD_(105),The prognosis of the patients with osteosarcoma was analyzed by Cox multi- variate survival analysis.Results The positive rate of COX-2 expression was 77.5 %,The expression of COX-2 was positively correlated with surgical grade and metastasis of osteosarcoma;There were significant difference between surgical grade Ⅰ and Ⅱ b、grade Ⅰ and Ⅲ(P0.05);Metastatic cases had higher MVD than those without metastasis(P

AIM: To study the isolation,purification and characterization of Ramulus Mori polysaccharide. METHODS: Ramulus Mori was extracted by boiling water. The raw extract was precipitated fractionally by alcohol deproteinized,passing through DEAE ion exchange cellulose ( DEAE-52) and SephadexG-100,obtained RMPS1 and RMPS2. The composition and characterization of Ramulus Mori polysaccharide were researched by TLC、IR、 GC、HPLC and smith degradation. RESULTS: The molecular weight of RMPS1 and RMPS2 were 5. 8 ? 105 and 6. 5 ? 105; RMPS1was made up of rhammose、arabinose、glucose and galactose with the molarity rate of 1. 08 ∶ 1 ∶ 1. 40 ∶ 1. 57; RMPS2 of rhammose、glucose and galactose with the molarity rate of 11. 38 ∶ 1 ∶ 1. 35. Smith degradation showed that the main linkage form in RMPS1 and RMPS2 was 1→2 and 1→4 glycosidic linkages,But some 1→3 glycosidic linkages also existed in the molecules; infrared spectrum showed that both had the polysaccharide characteristic absorption peaks. CONCLUSION: The structures of RMPS1 and RMPS2 are first determined from Ramulus Mori.