BACKGROUND:Hepatocyte transplantation as an effective method for liver failure has been confirmed by animal models and clinical application.However,limited source and poor proliferation of hepatocyte graft limit its development.Studies have shown that bone marrow mesenchymal stem cells(MSCs)have potentials to differentiate into hepatocyte and bile epithelial celts,with strong proliferation.OBJECTIVE:To explore the therapeutic effect of bone marrow MSCs transplantation on liver failure of New Zealand white rabbits.METHODS:Adult male New Zealand rabbits were treated with D-galactosamine,and 3 mL hepatocyte suspension(1×109/L)was injected into the liver of transplantation group,but the control group was injected with the same volume of culture solution with no bone marrow MSCs.Alanine aminotransferase(ALT)and aspartate aminotransferase(AST)activity was detected 48,72 hours,1,4 weeks following transplantation,and pathological detection was performed at 4 weeks.RESULTS AND CONCLUSION:The liver functional index following transplantation of bone marrow-derived MSCs transplantation was significantly decreased,and ALT and AST activity at 4 weeks was significantly less than the control group(P ＜ 0.05).At 4,the transplantation group displayed disorderly hepatic cord,hepatocyte swollen and degeneration,necrosis,accompanied by bleeding and inflammatory cell infiltration.In addition,the hepatic lobule structure was detectable,and regenerative hepatocyte increased among necrotic hepatocyte;small cells with large ratio of nucleus and cytoplasm at header,central vein and surrouding necrosis focus extended to the liver tissues.

OBJECTIVE To investigate the antibiotics-resistant genes in enterococci isolated from the 98th Hospital of PLA,Huzhou,Zhejiang Province,China.METHODS The antibiotics-resistant genes of TEM,aac(6′)/aph(2″),aph(3′)-Ⅲ,ant(2″)-Ⅰ,ant(4′,4″),ant(6)-Ⅰ,ermB,mefA,tetM,vanA,and vanB were analyzed by polymerase chain reaction(PCR) and verified by DNA sequencing in the 15 isolates of Enterococcus faecalis and 9 isolates of E.faecium.RESULTS The positive rate of the resistance genes of TEM,aac(6′)/aph(2″),aph(3′)-Ⅲ,ant(2″)-Ⅰ,ant(4′,4″),ant(6)-Ⅰ,(ermB,) mefA,tetM,vanA,and vanB in the 24 strains of enterococci tested were 37.5%,70.8%,25.0%,0.0%,0.0%,41.7%,75.0%,0.0%,41.7%,4.2%,and 4.2%,(respectively.) CONCLUSIONS The multidrug resistance of enterococci was a serious issue,and harbored antibiotics-resistance genes were the very important reasons of resistance to antibiotics in enterococci.

OBJECTIVE To investigate the ?-lactamase genes in 4 kinds of nonfermenting Gram-negative bacilli isolated from the 98th Hospital of PLA. METHODS Sixty strains of Acinetobacter baumannii,30 strains of Pseudomonas aeruginosa,19 strains of Stenotrophomonas maltophilia,and 15 strains of Flavobacterium were isolated from hospitalized patients.Nine kinds of ?-lactamases genes of TEM,SHV,OXA,CTX-M,PER,VEB,IMP,VIM and GES were analyzed by PCR and verified by DNA sequencing. RESULTS In A.baumannii and(P.aeruginosa),the positive rates of gene of TEM were 100.0% and 66.7%,respectively.SHV gene was positive in 18 of 60 strains of A.baumannii tested,17 of which were SHV-12 subtype ESBLs.The other was a new SHV type ?-lactamase nominated SHV-48.OXA gene was positive in 1 of 30 strains of P.aeruginosa tested,it was an OXA-10 subtype ESBLs.But the rest of genes were all negative. CONCLUSIONS There exist 4 kinds of(?-lactamase) genes at least in nonfermenting Gram-negative bacilli including TEM-1,SHV-12,SHV-48,and(OXA-10.)

OBJECTIVE To investigate the antimicrobial-resistant genes and consanguinity in multi-resistant Pseudomonas aeruginosa(MRPA) isolated from the 98th Hospital of PLA,Huzhou,Zhejiang Province,China.METHODS Thirty strains of MRPA were isolated from hospitalized patients between Sep 2003 and Oct 2004.Twenty four kinds of genes of blaTEM,blaSHV,blaOXA-10 group,blaPER,blaVEB,blaIMP,blaVIM,blaGES,blaCARB,blaDHA,blaMIR,aac(3)-Ⅰ,aac(3)-Ⅱ,aac(3)-Ⅲ,aac(3)-Ⅳ,aac(6′)-Ⅰ,aac(6′)-Ⅱ,ant(3″)-Ⅰ,ant(2″)-Ⅰ,aph(3′)-Ⅵ,oprD,qacE△1-sul1,catB,and cml1 were analyzed by PCR and verified by DNA sequencing.Resistant-genes cluster analysis was performed by Average.RESULTS In 30 strains of MRPA the positive rate of genes of blaTEM,blaOXA-10 group,blaCARB,aac(3)-Ⅱ,aac(6′)-Ⅰ,aac(6′)-Ⅱ,ant(3″)-Ⅰ,ant(2″)-Ⅰ,qacE△1-sul1,and cml1 were 66.7%,3.3%,3.3%,76.7%,3.3%,33.3%,53.3%,26.7%,83.3%,and 3.3%,respectively,and the deficiency rate of oprD gene was 90.0%.The gene of blaOXA-10 group was sequenced and determined to be blaOXA-10 subtype ESBL gene.But the rest of genes were all negative.According to the cluster analysis of resistant-gene,30 strains of MRPA isolated could be classified into four subgroups,which were caused by the infection in hospital.CONCLUSIONS At least 10 kinds of antimicrobial-resistant genes exist in MRPA isolates,and the deficiency rate of oprD gene is very high.MRPA can induce clone transmitted hospital infection and it has fulminant prevalence.

Objective To investigate the structure-activity relationship for 21 anthocyanins in inhibiting oxidized injury of endothelial cells,and explore the structural characteristics of anthocyanins closely related to their effects. Methods Endothelial cells were treated by ox-LDL at different concentrations of 50,100,150 or 200 ?g/ml,and MTT assay was used to determine IC50. After pre-incubated for 2 h with different concentrations ( 50,100 or 200 ?mol/L) of anthocyanins and then treated with 100 ?g/ml ox-LDL for another 24 h in endothelial cells,MTT assay was used to detect the cellular viability. After pre-treated for 2 h with different anthocyanins with 100 ?mol/L and treated with ox-LDL for another 24 h,MDA and NO level in the culture media were both measured according to the methods of assay kits. Structure-activity relationship was analyzed according to the respective cellular viability,MDA and NO level. Results Cellular viability was significantly inhibited by ox-LDL in a dose-dependent manner,and the IC50 was 100 ?g/ml. A significant correlation was observed among the effect of anthocyanins on cell viability,MDA production and NO release. The inhibitory effects of anthocyanins in ox-LDL-injured endothelial cells were positively related to the total number of hydroxyl groups and hydroxyl substitutions in B ring. 3′,4′-ortho-dihydroxyl substitution on B-ring and a 3-hydroxyl group on C-ring significantly enhanced the inhibitory effect of anthocyanins,yet methoxylation or glycosylation significantly decreased the effect. 6-hydroxylation substitution might attenuate the inhibitory effect of anthocyanins,while substitution at C5 or C5′ showed no significant influence on the effect of anthocyanins. Anthocyanin with monosaccharose substitution was much stronger than that with disaccharose substitution,while there was no significant difference between anthocyanins with glucoside and that with galacotoside substitution. Delphinidin and delphinidin-3-glucoside were respectively the most effectual anthocyanidin or anthocyanin. Conclusion 3′,4′-ortho-dihydroxyl substitution on B-ring and a 3-hydroxyl group on C-ring are the main structural requirements for anthocyanins in suppressing ox-LDL-induced injury in endothelial cells.

This study was purposed to investigate the role of monocrotaline-inducing mouse liver sinusoid endothelial cell (SEC) injury in hepatic veno-occlusive disease. BALB/c mice were randomly divided into 2 groups: control group and monocrotaline group, mice were orally administrated with normal saline or monocrotaline with concentration of 200 mg/kg at days 0, 1, 2, respectively. At days 3, 4, 6, 8 and 10 after oral administration with normal saline or monocrotaline, the liver function (ALT, TBIL, AKP) and liver index were examined, and the percentage of activated platelets were detected by flow cytometry. The SEC, vascular endothelial cells and hepatic fibrosis were observed by staining with hematoxylin-eosin and Masson. Transmission electron microscopy was used to observe sinusoidal endothelial cell damage and platelet adhesion. The results showed that compared with control group, mice in monocrotaline group were characterized by severe damage of SEC, numbers of platelet aggregation and adhesion, central number and sinusoidal fibrosis. The percentage of activated platelets and liver index increased (P < 0.05). The characterization of portal hypertension was presented later, such as dysfunction of liver and ascites. It is concluded that SEC injury induced by monocrotaline may be the first step of hepatic veno-occlusive disease, and this kind of SEC injury is self-limiting, but fibrosis is always observed.
Animals ; Endothelial Cells ; pathology ; Endothelium ; cytology ; Hepatic Veins ; cytology ; pathology ; Hepatic Veno-Occlusive Disease ; chemically induced ; pathology ; Liver Cirrhosis ; chemically induced ; pathology ; Male ; Mice ; Mice, Inbred BALB C ; Monocrotaline ; adverse effects ; Platelet Adhesiveness

Hybridoma cells were cultured by continuous perfusion in Fibra-Cel of 5L packed-bed bioreactor for 22 days in low serum or serum-free media. The corresponded amino acids were fed and serum concentration was decreased by analyzing glucose concentration, oxygen uptake rate, secretary antibody amount and amino acids concentration in culture supernatant. Comparing with continuous perfusion culture that amino acids were not fed, antibody amount of production was increased about 2-3 times. The inoculated cell density was 2.5 x 10(5) cells/mL, while the final cell density was 8.79 x 10(8) cells/mL. Antibody production was reached 295 mg/L/d at average level, and the highest level was reached 532 mg/L/d. These results provided a primary mode of enlarge culture for monoclonal antibody industralization.
Animals ; Antibodies, Monoclonal ; biosynthesis ; Cell Division ; Cells, Cultured ; Glucose ; pharmacology ; Hybridomas ; metabolism ; Mice ; Mice, Inbred BALB C ; Oxygen Consumption ; Perfusion

OBJECTIVETo examine the effects of endothelial progenitor cell (EPC) on hematopoietic reconstitution in allogeneic hematopoietic stem cell transplantation (allo-HSCT) mouse model.

METHODSAllo-HSCT mouse model was established with condition of BU/CY, in which C57BL/6 (H-2b) and BALB/c (H-2d) mice were used as donors and recipients respectively. Recipients were randomly divided into 4 groups: untreated group, BU/CY condition group, bone marrow transplantation (BMT) group and transplantation of BM cells combined with EPCs (combined transplantation) group. The pathological changes of BM cells following transplantation were dynamically observed. Changes of BM sinusoidal endothelium and angiogenesis were observed by MECA-32 antibody immunohistochemical staining. The proportion of intramedullary stem and progenitor cells and serum cytokines were analyzed by flow cytometry. The numbers of peripheral blood cells were also counted.

RESULTS(1) Injuries of BM hematopoietic tissue, sinusoidal endothelium and vascular were less severe in combined transplantation group than of BMT group. (2) EPC infusion significantly increased BM hematopoietic stem cells 21 days after transplantation. The percentage of BM hematopoietic stem cells in combined transplantation group peaked on day +14, which was higher than of BMT group (0.1743 vs 0.0787) (P<0.05). The continuously increased percentage of BM hematopoietic progenitor cells in combined transplantation group was significantly higher than in BMT group on day +21 (0.4550 vs 0.3905) (P<0.05). (3) The number of peripheral white blood cells in combined transplantation group was always higher than of BMT group, which reached the peak on day +14 (0.74×10⁹/L to 0.47×10⁹/L) (P<0.05). The peak number of peripheral blood platelets on day +14 in combined transplantation group was significantly higher than of group BMT (1228.9×10⁹/L to 977.12×10⁹/L) (P<0.05).

CONCLUSIONAllo-HSCT combined with EPC infusion accelerated hematopoietic reconstitution compared with BMT alone in allo-HSCT mouse model.

Animals ; Endothelial Cells ; cytology ; Hematopoietic Stem Cell Transplantation ; methods ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Stem Cells ; cytology ; Transplantation, Homologous

To verify the effect of echinacoside on replication and antigen expression of hepatitis B virus (HBV) by using HBV-transfected HepG2. 2. 15 cells as the in vitro model. The ELISA method was used to determine HBeAg and HBsAg levels in cellular supernatants. The effect of echinacoside on HBV replication was studied by using HBV transgenic mice as the in vivo model. First of all, the HBV DNA level in hepatic tissues was quantified with PCR method. Meanwhile, the serum transaminase levels and hepatic pathological changes were also evaluated. Subsequently, HBV transgenic mice were divided into five groups: the control group, the lamivudine group (50 mg · kg(-1)) and echinacoside high, medium and low dose group (50, 25 and 12.5 mg · kg(-1)). The mice were orally administered with drugs once per day for 30 days. At the 31st day, the mice serum was separated to measure HBsAg, HBeAg and HBV DNA. Additionally, the liver HBV DNA level and histopathological change were detected. The results indicated that echinacoside at 50 and 100 mg · L(-1) suppressed significantly HBsAg and HBeAg expressions on the sixth day, with the maximum inhibition ratios of 42.68% and 46.29%; And echinacoside at 100 mg · L(-1) also showed an inhibitory effect on HBV DNA. Besides, echinacoside at 50 mg · kg(-1) inhibited significantly HBsAg and HBeAg expressions of HBV transgenic mice, with the inhibition ratios of 42.82% and 29.12%, and reduced markedly the serum HBV DNA level in HBV transgenic mice. In conclusion, the study suggested that echinacoside has a strong effect against HBV replication and antigen expression.
Animals ; DNA, Viral ; blood ; Female ; Glycosides ; pharmacology ; Hep G2 Cells ; Hepatitis B Surface Antigens ; blood ; Hepatitis B e Antigens ; blood ; Hepatitis B virus ; drug effects ; physiology ; Humans ; Male ; Mice ; Mice, Inbred C57BL ; Virus Replication ; drug effects

By using Zinc nitrate as precursor and hydraZine hydrate as reducing agent, polydiallyl dimethyl ammonium chloride modified reduced graphene oxide/Zinc oxide composite materials ( PDDA-rGO/ZnO) were prepared by simultaneous reduction of graphene oxide ( GO) and Zinc nitrate.The composite materials were characteriZed by Fourier transform infrared ( FTIR ) spectroscopy, X-ray diffractometer ( XRD ) and transmission electron microscopy ( TEM) , and their electrochemical catalytic activity for uric acid was studied by cyclic voltammetry ( CV ) and linear sweep voltammetry ( LSV ) measurements.The result showed that PDDA-rGO/ZnO modified glassy carbon electrode prepared here was sensitive, reproducible and stable, and had significant electrocatalytic activity for UA.When using linear sweep voltammetry for detection of UA, the responses of modified electrode were linear with UA concentration in the ranges of 0.02-0.1 mmol/L and 0.1-1.0 mmol/L respectively, with detection limit of 15.9 nmol/L (S/N=3).