Objective To investigate the effect of hydrogen on endotoxin-induced expression of zonula occludens-1 (ZO-1) in human colon epithelial cells (Caco-2 cells).Methods Caco-2 cells were cultured routinely,seeded in Transwell chambers or wells,and randomly divided into 4 groups (n =45 each) using a random number table:control group (group C);hydrogen-rich culture medium group (group H);endotoxin group (group E);hydrogen-rich culture medium + endotoxin group (group HE).The cells were cultured in high-glucose DMEM culture medium in group C.The cells were incubated in hydrogen-rich culture medium containing hydrogen 0.6 mmol/L in group H.The cells were incubated in highglucose DMEM culture medium containing 50 μg/ml lipopolysaccharide in group E.The cells were incubated in hydrogen-rich culture medium containing 50 μg/ml lipopolysaccharide and 0.6 mmol/L hydrogen in group HE.Transepithelial electrical resistance (TEER) was measured before incubation or culture,and at 6,12 and 24 h of incubation or culture.The viability of Caco-2 cells was measured by methyl thiazolyl tetrazolium assay at 24 h of incubation or culture.The expression of ZO-1 mRNA in Caco-2 cells was determined using real-time reverse transcriptase polymerase chain reaction at 6,12 and 24 h of incubation or culture.The distribution of ZO-1 in Caco-2 cells was observed by immunofluorescence at 24 of incubation or culture.Results Compared with group C,TEER was significantly decreased at 6,12 and 24 h of incubation or culture,and the expression of ZO-1 mRNA was significantly down-regulated in E and HE groups (P＜0.05),and no significant change was found in the parameters mentioned above in group H (P＞0.05).Compared with group E,TEER was significantly increased at 6,12 and 24 h of incubation or culture,and the expression of ZO-1 mRNA was significantly up-regulated in group HE (P＜0.05).The distribution of ZO-1 protein in cell membrane became discontinuous,and the distribution of ZO-1 protein in cytoplasm was significantly increased in group E.Compared with group E,the distribution of ZO-1 protein in cell membrane was significantly increased and gradually became continuous,and the distribution of ZO-1 protein in cytoplasm was significantly decreased in group HE.Conclusion The mechanism by which hydrogen reduces the damage to human colon epithelial cell barrier is related to up-regulation of ZO-1 expression and improvement in the redistribution of ZO-1 protein.

Objective To explore the feasibility of carrying out the common imprecision range among different biochemical systems in different laboratories.Methods Biochemical professionals visited 18 third-grade class A hospitals in Hebei Province,investigated the internal quality control data of 21 biochemical tests and made classification according to certain parameter criterias.Data were collected from April to September,2010 and calculated for cumulative mean,standard deviation (s) and CV.Data were compared according to 1/3,1/4 of TEa established by CLIA'88 and allowable CVb％ derived from biological variants.Results Among 18 hospitals,5 (27.8％) set their target value by mean value of 6 months,5(27.8％) by continuous 20 days and 8(44.4％) by the given value of the supplier.CVs of 21biochemical tests were quite different among 18 biochemistry laboratories,in which LDH was 6.79 times and CK was 76.79 times different from one another.35.5％-94.1％ biochemical tests met the requirement of CV ＜ 1/3TEa and 0.0％-91.7％ met the requirement of CV ＜ 1/4TEa.0.0％-94.1％ of tests were below allowable CVb％ 16/21 (76.2％) tests could satisfy the requirement of CV ＜ 1/3TEa.The top five tests which didn't meet the requirements were Na,Urea,TBIL,ALT and Glu.Conclusions The internal quality control among biochemistry laboratories in Hebei Province has not been standardized yet.According to the survey data,biochemistry laboratories of third-grade class A hospitals may set 75％ point of imprecision as a reference.After a period of improvement,we will set common imprecision range among biochemistry laborotories in Hebei Province and even in China.

AIM To study the effects of matrine (Ma) on the action potential and contractile force in guinea pig papillary muscles. METHODS Conventional microelectrode technique was used to record the fast action potentials (FAP) and slow action potentials (SAP) induced by histamine and BaCl_2 of guinea pig papillary muscles. RESULTS Ma 10,25,50 ?mol?L -1 dose-dependently prolonged the action potential duration at 50%, 90% repolarization (APD_ 50 , APD_ 90 ) and effective refractory period (ERP) of FAP, and lengthened the APD_ 50 , APD_ 90 of SAP induced by histamine and BaCl_2 when perfused with KCl 25 mmol?L -1 Tyrode's solution. The maximal upstroke velocity (V_ max ) of FAP, SAP and contractile force (Fc) were not affected by matrine 10,25,50 ?mol?L -1 . CONCLUSION It was suggested that Matrine could block K + channels in the myocardium.-

OBJECTIVE:To determine the content of levofloxacin by PVC membrane selective electrode. METHODS: A glass electrode coated with levofloxacin PVC membrane was prepared for the first time with the molecular complex of levofloxacin iodide and bismuth iodide as electric active material, and a series study was performed on its responsibility. RESULTS: The electrode gave Nernst response to levofloxacin over the concentration range of 5.0?10-3～1.0?10-5 moL?L-1 with a slope rate of 56.5mV?pC-1, a suitable pH of 2.5～4.0, and an average recovery rate of 98.31%～101.6%(2.80%～4.90%). CONCLUSION: The electrode was simple in operation, and it has a rapid response and good reproducibility, and it is applicable for the quantitative determination of levofloxacin.

Objective To investigate the role of Rho kinase (ROCK) in the protective effects of hydrogen on intestinal epithelial barrier function in sepsis. Methods Caco-2 cells were cultured routinely, and divided into 6 groups randomly (n=3):control group (C group), hydrogen-rich medium group (H group), lipopolysaccharide (LPS)-treatment group (L group), hy?drogen+LPS-treatment group (HL group), Rho kinase inhibitor (Y-37632) treatment group (Y group) and Rho kinase inhibi?tor Y-27632+LPS-treatment group (YL group). H group was treated with 0.6 mmol/L hydrogen-rich media. The concentra?tion of LPS and Y-27632 were 50 mg/L and 25μmol/L separately. After the Caco-2 monolayer model was established, the transepithelial electrical resistance (TEER) values were measured regularly. When the TEER value reached 800Ω·cm2, the treatment was administered. Then TEER values were measured at 6 h, 12 h and 24 h, and FITC-dextran permeability was de?tected at 24 h. Cells were seeded on 6-well plates. After cell density reached 80%-90%, treatments were given randomly. The real time-polymerase chain reaction (RT-PCR) was conducted to assess mRNA levels of ZO-1 and ROCK mRNA. ZO-1 and ROCK protein expression levels were detected by Western blot assay. Results Compared with C group, TEER values were elevated in 12 h and 24 h in H group (P<0.05). There were no statistical significances in FITC-dextran permeability,
protein expression levels of ZO-1 and ROCK between C group and H group (P>0.05). TEER values were elevated at 6 h, 12 h and 24 h in Y group (P<0.05). There was no significant difference in FITC-dextran permeability between C group and Y group (P > 0.05). The mRNA expression of ZO-1 increased and mRNA expression of ROCK decreased in Y group (P <0.05). The TEER values reduced at 6 h, 12 h and 24 h in L group. The FITC-dextran permeability increased significantly, mRNA and protein expressions of ZO-1 significantly decreased, mRNA and protein expressions of ROCK significantly in?creased in L group (all P<0.05). Compared with L group, TEER values increased significantly at 6 h, 12 h and 24 h in YL group, FITC-dextran permeability decreased, mRNA expressions of ZO-1 increased, mRNA expressions of ROCK de?creased in YL group (P<0.05). Compared with L group, TEER values increased at 6 h, 12 h and 24 h in HL group, FITC-dextran permeability reduced markedly, protein expressions of ZO-1 increased at each time point, protein expressions of ROCK decreased at each time point in HL group (P<0.05). Conclusion Hydrogen can protect intestinal barrier function against sepsis, ameliorate the integrity and permeability of intestinal epithelium and increase the expressions of intercellular tight junction proteins. The suppression of Rho kinase over-expression induced by LPS may be involved in these protective effects of hydrogen.

Objective To investigate the biological effects of Pseudomonas aeruginosa quorum sensing molecule OdDHL on murine mast cells. Methods The molecule structure and purity of synthesized OdDHL were confirmed by mass spectrum or proton nuclear magnetic resonance (NMR) and high-performance liquid chromatography, respectively. Its biological activity was checked using a quorum sensing sensor bacterial strain. The viability, apoptosis and intracellular calcium changes of P815 cell line in response to different concentration of OdDHL were determined. Results The biological active OdDHL was synthesized successfully. OdDHL inhibited proliferation of P815 cells in a dose, and time dependent manner. It also induced apoptasis and intracellular calcium release in P815 cells. Conclusion Psendomonas aeruginosa quorum sensing molecule OdDHL induces apoptosis and intracellular calcium release in murine mast cell line P815.

BACKGROUND:There is no effective drug for idiopathic pulmonary fibrosis (IPF), because of a lack of the animal model imitating the complete pathogenesis of human IPF. Therefore, it is critical to establish an ideal animal IPF model used for investigating the underlying pathogenesis and developing a kind of effective drug. OBJECTIVE:To establish an animal model that can mimic more characters of human IPF. METHODS:Seventy male C57BL/6 mice were randomly divided into two groups, fol owed by subjected to the intraperitoneal injection of bleomycin (35 mg/kg) on days 1, 4, 8, 11, 15, 18, 22, and 25, twice (group A) or once (group B) a week. Mice were sacrificed at 2, 4, 6, 8, and 10 weeks after the eighth injection, and the lung tissues were moved used for hematoxylin-eosin, Masson and immunohistochemical stainings. RESULTS AND CONCLUSION:There were various degrees of alveolitis and pulmonary fibrosis in the two groups at different time points after the last injection. The scores of alveolitis and pulmonary fibrosis in the group A began to gradual y increase from the 2nd week and reached the highest level at the 6th-8th weeks until the 10th week. In contrast, the scores of alveolitis and pulmonary fibrosis in the group B peaked at the 2nd week, then fluctuately decreased, and were significantly lower than those in the group A at the 6th week (P<0.05). Immunohistochemistry showed that type I col agen deposition was mainly distributed in the subpleural region, peri-vascular region and alveolar septa, which was consistent with Masson staining findings. The expression levels of transforming growth factorβ1 (TGF-β1) andα-smooth muscle actin (α-SMA) in the regions developing alveolitis and pulmonary fibrosis were significantly increased. In the group A, the expression levels of type I col agen, TGF-β1,α-SMA, and the hydroxyproline content in the lung tissues reached the peak level at 6-8 weeks. However, in the group B, al above indicators reached the highest level at the 2nd week, but gradual y decreased thereafter. At the 4th week, the expression Levels of TGF-β1 andα-SMA in the group B were significantly lower than those in the group A (P<0.05). At the 6th week, the hydroxyproline and type I col agen levels in the group B were significantly lower than those in the group A (P<0.05). In conclusion, the mouse model of pulmonary fibrosis induced by intraperitoneal injection of 35 mg/kg bleomycin twice weekly can be used to mimic the repetitive wound healing process, pathological morphology and cytokine changes of human IPF, which is prone to administration, with better stability and repeatability. This model is of great significance for the study on IPF. Subject headings:Disease Models, Animal;Pulmonary Fibrosis;Bleomycin

Objective To evaluate the effect of inhalation of hydrogen gas (H2) on Rho/Rho kinase (ROCK) pathway in lung tissues of septic mice with acute lung injury.Methods Forty male C57BL/6 mice,weighing 20-25 g,aged 6 weeks,were randomly divided into 4 groups (n=10 each) using a random number table:sham operation group (group Sh),sham operation + inhalation of H2 group (group H2),sepsis group (group S),and sepsis+ inhalation of H2 group (group S+H2).Sepsis was produced by cecal ligation and puncture (CLP).Both H2 and S+H2 groups inhaled 2％ H2 for 1 h starting from 1 and 6 h after CLP.The mice in each group were sacrificed at 24 h after CLP.The bronchoalveolar lavage fluid (BALF) was collected for determination of protein concentrations,polymorphonuclear neutrophil (PMN) count,and concentrations of tumor necrosis factor-alpha (TNF-α) and interleukin-1beta (IL-1β) (by enzyme-linked immunosorbent assay).The lung tissues were obtained for examination of the pathological changes which were scored and for determination of the wet/dry weight ratio (W/D ratio),activities of myeloperoxidase (MPO) and superoxide dismutase (SOD),malonaldehyde (MDA) level,the expression of Rho,ROCK1,ROCK2 and activated caspase-3,and phosphorylation of myosin phosphatase target protein 1 (MYPT-1) (by Western blot).Results Compared with group Sh,the concentrations of protein,PMN count,and concentrations of TNF-α and IL-1β in BALF,lung injury score,W/D ratio,and levels of MPO and MDA in lung tissues were significantly increased,the activity of SOD in lung tissues was significantly decreased,the expression of Rho,ROCK1,ROCK2 and activated caspase-3 was significantly upregulated,and the phosphorylation of MYPT-1 in lung tissues was significantly increased in S and S+H2 groups (P＜0.05),and no significant change was found in the parameters mentioned above in group H2 (P＞0.05).Compared with group S,the concentrations of protein,PMN count,and concentrations of TNF-α and IL-1β in BALF,lung injury score,W/D ratio,and levels of MPO and MDA in lung tissues were significantly decreased,the activity of SOD in lung tissues was significantly increased,the expression of Rho,ROCK1,ROCK2 and activated caspase-3 in lung tissues was significantly down-regulated,and the phosphorylation of MYPT-1 in lung tissues was significantly decreased in group S+H2 (P＜0.05).Conclusion The mechanism by which inhalation of H2 attenuates acute lung injury is related to inhibition of Rho/ROCK pathway activation in lung tissues of septic mice.

Objective To estabhsh a rat portacaval shunt model of portal vein arterialization (PVA)by using right renal artery,andtoinvestigateitsinfluence on portal velnpressure,liver funciton and hepatic pathology.Method Forty-three Sprague-Dawley rats were assigned to PVA group(33 rats)and control group(10 rats).Portal pressure,liver function and pathological changes were evaluated at day 2,2 weeks and 2 months after surgery,respectively.Results The experimental model was successfully established in 30 out of 33(91%)rats.At day 2,2 weeks and 2 months after the operation,the portal pressure increased significanfly(P<0.05)compared with the control group,but no significant differences were found in the senrum level of alanine amlnotransferase,total bilirubin,total bile acid and creatinine.There were no patholgiccal changes from day 2 to 2 weeks after surgery,only postoperative expanslon of the sinusoidal snace was found on 2 months after surgery.Conclusion The rat model of PVA by renal artery was successfully established with the hand-suture mierosurgical techniques.The portal vein pressure maintained at a high level after PVA,meanwhile,no negative effect was found on the liver function and hepatic histological structure changes.

Objective To explore the method of pre expanded deltopectoral flap for repairing post burn faciocervical scars.Methods Anterior axillary incisions were made and appropriate expanders were implanted above anterior chest wall at the first stage.After a 4 6 months' expanding,the flaps based on perforating branches of the internal mammary artery,branches from the thoracoacromi al area,or perforating branches from deltoid muscle,were designed and raised according to scars and dominant vessels.The donor sites were closed at same time without skin graft.Results 43 patients with 51 flaps were operated for reconstruction of post burn faciocervical scars.All flaps and donor sites survived well.Conclusions Pre expanded deltopectoral flap is an ideal donor site for repairing post-burn faciocervical scars.