AI M:Our purpose wastoinduce MSCs differentiatinginto endothelial cells (EC)invitroandto providetheseed cells for study of cardiovascular tissue -engineering.METHODS :MSCs were separated by gradient centrifugation on Percoll(density 1 073 g/L) fromhuman bone marrow(HBM) ,and incubated for purification and amplification in DMEM(lowglucose)with 10 %fetal bovine serum(FBS) .Then,the MSCs were incubated for orientation differentiated into ECin DMEM(high glu-cose) with20 %FBS,VEGF(10?g/L) ,bFGF(5?g/L) ,L-glutamine (2 mmol/L) ,penicillin (1?105U/L) and streptomycin(100 mg/L) for about 14 -21 days andtheir phenotypic characteristics were analyzed byflowcytometry.Afterwards ,the differenti-ating cells were evaluated by histology andimmunohistochemistry.RESULTS :The quantity of MSCs was increasedfrom5?0?105inthe primary culture to 8?0?1012,or toincrease to 1?6?107times after 15 generations of incubation.The purity of MSCs wasabove 95 %and 98 %homogeneous at passages 2 and 3 ,respectively.About 80 %-90 %of the differentiating cellsfrom MSCs af-ter 14 -21 days were positively stainedfor Ⅷfactor (vWF) related antigen by immunohistochemistry assay,and Weible -paladecorpuscle was also observed bytransmission electron microscopyinthe cytoplasm.CONCLUSION:MSCs from HBMhave the ca-pability of differentiationinto ECsin vitro,which may be a potential source of seed cellsforfabrication of tissue -engineering heartvalve ,particularlyin children with congenital heart disease .

The purpose of this investigation is to improve ethanol production and decrease acetate formation in Saccharomyces cerevisiae strain YS2-?adh2.The strain YS2-?adh2 with deleted alcohol dehydrogenase Ⅱ(adh2) gene was isolated in our lab with higher ethanol production than that of the strain YS2.The ace-taldehyde dehydrogenase Ⅵ(ald6) gene encoded a cytosolic acetaldehyde dehydrogenase,a key enzyme of the pyruvate dehydrogenase(PDH) bypass,transfers acetaldehyde to acetate.To disrupt ald6 gene of the strain YS2-?adh2,ald6 gene targeting cassettes were synthesized by long flanking homology PCR(LFH-PCR) and then were transformed into YS2-?adh2 mutants by LiAc/SS Carrier DNA/PEG method.Positive transformants were selected with G418 and further confirmed by PCR.Once correctly integrated into the genome,the selective marker was rescued by transforming the plasmid pSH65 into the positive transformants and inducing the Cre expression with a Cre/loxP-mediated marker removal procedure.We named the ald6 gene knocked-out strain as YS2-?adh2-?ald6 which has a 12.5% higher ethanol production and a 18% lower acetate formation compared to the strain YS2.

Objective:To observe the outcome after sacral canal injection in patients with disc herniation associated with without sciatica. Methods:From December 2010 to June 2011,65 patients with acute low back pain without sciatica due to lumbar disc herniation or bulging confirmed by CT or MRI were randomly divided into sacral canal injection group (experi-mental group) and lumbar oblique wrench group (control group):the experimental group had 35 cases,including 30 males and 5 females,with an average age of (43.90±1.14) years old ranging from 33 to 56 years old. The control group had 30 cases,in-cluding 27 males and 3 females,with an average age of (44.00±1.19) years old ranging from 34 to 57 years old. The course of morbidity was 1 to 3 days. All patients received sacral canal injection or lumbar oblique wrench method. The visual analog scale (VAS) scores before and at 30 min after treatment were compared between two groups. Results:The symptom of acute low back pain were relieved obviously. The average VAS scores before and after treatment in experimental group were de-creased from 6.63 ±0.97 to 3.06 ±1.51,in control group were from 6.67 ±0.96 to 3.93 ±1.20 respectively. These two methods could improve the VAS score,but the effect of sacral canal injection group was better than that of lumbar oblique wrench group,there was statistically differences (P<0.05). Conclusion:It is effective that the methods of sacral canal injection and lumbar oblique wrench applied to patients with acute low back pain without sciatica due to lumbar disc herniation or bulging confirmed,the former has better effect.

AIMTo amplify mesenchymal stem cells (MSCs) from human bone marrow (HBM) and to induce MSCs differentiated into endothelial cells (ECs) in vitro, which possibility and conditions were to be discussed.

METHODSMSCs were separated by gradient centrifugation on Percoll (density 1.073 g/ml) from HBM, and incubated for purification and amplification in DMEM (low glucose) with 10% fetal bovine serum (FBS). MSCs, which their phenotypic characteristics were analyzed by flow cytometry, were incubated for orientation differentiated into ECs in DMEM (high glucose) with 20% FBS and VEGF (10 ng/ml) for about 14 - 21 days. Afterwards, the cells differentiated were evaluated by immunohistochemistry with Tie-2 monoclonal antibody and by transmission electron microscopy (TEM) for observation Weible-palade corpuscle in the cytoplasm.

RESULTSThe quantity of MSCs was increased from 5.0 - 10(5) in the primary culture to 8.0 x 10(12), or to increase 1.6 x 10(7) times after 15 generations incubated. The purity of MSCs was above 95% and 98% homogeneous at passages 2 and 3, respectively. The typical "cobblestone" of these cells presented from MSCs differentiation culture after 14 - 21 days was observed by light microscopy. More than 90% of the cells were positive stain for Tie-2 related antigen by immunohistochemistry assay. The Weible-palade corpuscle, which form is typical morphology of ECs, was also observed by TEM in the cytoplasm.

CONCLUSIONMSCs from HBM has the capability in differentiation into ECs in vitro, which is possible to provide the seed cells for fabrication of tissue-engineering heart valve.

Adult ; Antibodies, Monoclonal ; Bone Marrow Cells ; cytology ; Cell Differentiation ; Cells, Cultured ; Endothelial Cells ; cytology ; Flow Cytometry ; Humans ; Mesenchymal Stromal Cells ; cytology ; Receptor, TIE-2 ; immunology

OBJECTIVETo explore the anti-tumor efficacy of anti- vascular endothelial growth factor (VEGF) McAb 5-fluorouracil (5-FU) loaded polylactic acid (PLA) nanoparticles (NPS) in human gastric carcinoma xenografts of nude mice.

METHODSAnti-VEGF McAb 5-FU loaded PLA NPS were made by ultrasound emulsification. Nude mice model of human gastric carcinoma xenografts was established. Therapeutic effects of drugs on human gastric carcinoma xenografts and side effects concerned were observed.

RESULTSThe tumor inhibition rates of control group, nanosphere without 5-FU group, 5-FU (20 mg/kg) group, anti-VEGF McAb nanosphere without 5-FU group, anti-VEGF McAb group, nanosphere with 5-FU group, 5-FU (20 mg/kg) combined with anti-VEGF McAb group, anti-VEGF McAb 5-FU loaded nanosphere group was 0, 6.61%, 24.26%, 27.94%, 35.29%, 37.50%, 39.71% and 52.21% respectively, and there were no significant differences between anti-VEGF McAb 5-FU loaded nanosphere group and nanosphere group without 5-FU in WBC count, serum alanine transferase level or creatinine level. Compared with control group and anti-VEGF McAb 5-FU loaded nanosphere group, the 5-FU group decreased by 34.43% and 37.38% respectively in WBC count (P< 0.05), and increased by 93.17% and 66.56% respectively in alanine transferase. There were significant differences between experimental groups and control group in apoptosis index, especially between anti-VEGF McAb 5-FU loaded nanosphere group and control group (P< 0.05). The microvessel density (MVD) of experimental groups containing anti-VEGF McAb was significantly lower than that of control group or groups containing 5-FU (P< 0.05).

CONCLUSIONAnti-VEGF McAb 5-FU loaded nanosphere can increase the tumor inhibitory rate of 5-FU, induce apoptosis by inhibiting tumor angiogenesis with less side effect, and then enhance therapeutic effect, which indicate its potential as a novel, safe nano-tumor-targeting drug.

Animals ; Antibodies, Monoclonal ; administration & dosage ; pharmacology ; Antimetabolites, Antineoplastic ; administration & dosage ; pharmacology ; Cell Line, Tumor ; Drug Carriers ; Fluorouracil ; administration & dosage ; pharmacology ; Humans ; Lactic Acid ; administration & dosage ; pharmacology ; Mice ; Mice, Nude ; Nanoparticles ; Neovascularization, Pathologic ; Polyesters ; Polymers ; administration & dosage ; pharmacology ; Stomach Neoplasms ; drug therapy ; pathology ; Vascular Endothelial Growth Factor A ; administration & dosage ; pharmacology ; Xenograft Model Antitumor Assays

Objective To investigate the incidence of central line-associated bloodstream infection (CLABSI) in patients with hematopoietic stem cell transplantation (HSCT), explore risk factors for the occurrence of CLABSI.Methods Basic information of patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) who underwent HSCT in a hematology department from November 1, 2016 to October 31, 2017 was collected, incidences of original CLABSI (OCLABSI) and modified CLABSI (MCLABSI) were calculated, related risk factors were analyzed by multivariate Cox regression.Results A total of 218 patients with AML and MDS who underwent HSCT were enrolled, 19 of whom had OCLABSI and 10 had MCLABSI.Twenty-one strains of pathogens were isolated from 19 patients with OCLABSI, including 9 gram-positive bacteria, 11 gram-negative bacteria, 1 fungus;9 strains were multidrug-resistant organisms.The main risk factors for OCLABSI included the female (HR=0.088;95％CI:0.017-0.440;P=0.003), age (HR=1.560;95％CI:1.066-2.530;P=0.034), bone marrow cell transplantation only (HR=4.408;95％CI:1.860-22.593;P=0.043), ATG/CSA/MMF/MTXG for preventing graft-versus-host disease (GVHD) (HR=0.101;95％CI:0.015-0.686;P=0.019), and MTX for preventing GVHD (HR=0.097;95％CI:0.011-0.816;P=0.032).Conclusion Definition of MCLABSI can provide more accurate monitoring on deep central venous catheter-related bloodstream infection.Incidence of CLABSI in HSCT patients can be reduced by early detection of high-risk population according to high-risk factors, strict adherence to the prevention and control measures of bloodstream infection, and implementation of immune recombination after enhanced transplantation.

Objective To explore the influence of drug dosage, solvent and other main influencing factors on the successful establishment of alloxan-induced hyperglycemia mouse model and the effect on the stability of this model. Methods 160 6-8-week-old Kunming mice ofSPF grade, (male:female=1:1) were used in this study.The influences of different dosages of alloxan and solvent combinations on the successful establishment rate of the model, survival rate, body weight, fasting blood glucose, blood glucose area under curve, serum insulin level and their stabilities were dynamically observed for six weeks.Results By single intraperitoneal injection of 160 mg/kg bw alloxan ( pH 4.5 citrate sodium as solvent) , we were able to obtain a stable experimental hyperglycemic mouse model with higher levels of successful establishment rate (70%), survival rate (75%), fasting blood glucose (15-20 mmol/L), glucose area under the curve (55-65 mmol/L) and a lower but not loss of serum insulin levels (21 mIU/L).Conclusions In the present study we have carefully considered the influence of main factors such as drug dosages, solvent, etc., on the alloxan-induced experimental hyperglycemic mouse model, and successfully established this model after 6-week period observation of its stability.This model may provide a useful tool in the research of experimental diabetes and hypoglycemic functional studies.

Objective: To explore the association between chronic hepatitis B virus infection and diabetes among adults. Methods: The baseline data of China Kadoorie Biobank ( CKB ) study from Tongxiang of Zhejiang Province was used for analysis. Community residents were investigated in the study from August 2004 to May 2008, including questionnaire survey, physical measurement and biological sample test. Univariate and multivariate logistic regression models were used to estimate the association of chronic hepatitis B virus infection with diabetes. Results: Totally 52 888 participants were included in the final analysis. The overall prevalence of HBsAg-positive was 3.55% ( N=1 877 ). The overall prevalence of diabetes was 5.17% ( N=2 733 ). The prevalence of HBsAg-positive in diabetic patients was 3.51% ( N=96 ). Both univariate and multivariate logistic regression models indicated that there was no association between chronic hepatitis B virus infection and diabetes（ P>0.05 ）. Conclusion No significant association has been found between chronic hepatitis B virus infection and diabetes among adults.

This study was aimed to investigate the characteristics of human bone marrow mesenchymal stem cells during ex-vivo expansion, MSCs were isolated from human bone marrow. At each passage, the characteristics of proliferation kinetics, osteogenic and adipogenic differentiation potential were analyzed, and cell morphology, surface markers were investigated as well. The karyotype analysis was done in different passage cells. The infection HIV, HCV, HBV and TP were detected by ELISA. Mycoplasma contamination in vitro was detected by PCR method. HLA-SBT was used to reanalyze the results of HLA antigens and alleles. STR genetic loci were detected by PCR in the MSC1, MSC2, MSC3 and MSC4. The results indicated that the proliferative ability and osteogenic potential decreased with the increase of passage number during culture expansion. The multiple differentiation potential of MSCs was maintained during their life span. Karyotype analysis showed that MSCs from 4 groups before passage 8 were normal. The expression of CD29, CD44, CD105, CD166 and CD73 were positive. The expression of CD14, CD34, CD45, CD80, CD86 were all negative. SBT was used to identify HLA-A, B, Cw, DRB1, DRPB1, DQ alleles in the MSC1, MSC2, MSC3, MSC4. The genetype of STR in the MSC1, MSC2, MSC3, MSC4 was different. MSC 3 was examined by TP-ELISA to confirm the infectious disease of TP. MSC2 was contaminated by mycoplasma at passage 5. It is concluded that culture expansion causes MSCs to gradually lose their stem cell properties. During ex-vivo expansion of MSCs, the osteogenic differentiation potential is decreased. MSCs before passage 8 can be a valuable subject for basic research and clinical application.
Adipogenesis ; Adult ; Bone Marrow Cells ; cytology ; Cell Differentiation ; physiology ; Cells, Cultured ; Female ; Humans ; Karyotyping ; Male ; Mesenchymal Stromal Cells ; cytology ; Osteogenesis

OBJECTIVETo investigate the effects of Qihong capsule (QH) on HeLa cells infected by coxsackievirus B3 (CVB3) in vitro and its potential antiviral mechanism.

METHODSHeLa cells were infected by CVB3 in vitro. XTT assay and plaque inhibition assay were performed to determine the 50 % effective dose, (ED50), 50 % inhibitory concentration (IC50), and 50% cytotoxicity concentration (CC50) of QH and the control drug, ribavirin. The total therapeutic index (TI) was calculated. Anti-viral time-course experiments were performed to compare the anti-viral effects at different time points. The inhibitory effects of QH on the attachment and penetration of CVB3 were also observed.

RESULTSXTT assay and plaque inhibition assay showed that the ED50 and IC50 were (7.16+/-0.80) mg/L and (2.63+/-0.50) mg/L in QH group and (4.35+/-0.40) mg/L and (1.92+/-0.30) mg/L in ribavirin group, respectively. CC50 was 16-fold higher in QH group than in ribavirin group QH: (1 648+/-219) mg/L vs. Ribavirin: (103+/-14) mg/L. Time-course studies demonstrated that antiviral effect of QH was mainly found 0-4 hours after infection. QH effectively blocked the attachment and penetration of CVB3 into cells.

CONCLUSIONBy inhibiting the attachment and penetration of CVB3, QH can effectively inhibit the invasion of virus in vitro with low toxicity.

Antiviral Agents ; pharmacology ; Capsules ; Drugs, Chinese Herbal ; pharmacology ; Enterovirus B, Human ; drug effects ; HeLa Cells ; Humans ; Inhibitory Concentration 50