OBJECTIVETo study the effect of hypoxia on cofilin activation in intestinal epithelial cells and its relation with distribution of tight junction protein zonula occludens 1 (ZO-1).

METHODSThe human intestinal epithelial cell line Caco-2 was used to reproduce monolayer cells. The monolayer-cell specimens were divided into control group (no treatment), hypoxic group ( exposed to hypoxia), and normoxic group (exposed to normoxia) according to the random number table. Western blotting was used to detect the protein expressions of cofilin and phosphorylatedl cofilin (p-cofilin) of cells in normoxic group and hypoxic group exposed to normoxia or hypoxia for 1, 2, 6, 12, and 24 h and control group, with 9 samples in control group and 9 samples at each time point in the other two groups. The other monolayer-cell specimens were divided into hypoxic group (exposed to hypoxia) and control group (no treatment) according to the random number table. Cells in hypoxic group exposed to hypoxia for 1, 2, 6, 12, and 24 h and control group were obtained. Morphology and distribution of F-actin was observd with laser scanning confocal microscopy, the ratio of F-actin to G-actin was determined by fluorescence method, and distribution of ZO-l and cellular morphology were observed with laser scanning confocal microscopy. The sample number of last 3 experiments was respectively 3, 6, and 3 in both hypoxic group (at each time point) and control group. Data were processed with paired ttest, analysis of variance of repeated measurement, and LSD-t test.

RESULTSThe protein expressions of cofilin and p-cofilin of cells between normoxic group exposed to normoxia for 1 to 24 h and control group showed no significant changes (with values from -0.385 to 1.701, t(p-cofilin)values from 0. 040 to 1.538, P values above 0.05). There were no obvious differences in protein expressions of en filmn of cells between hypoxic group exposed to hypoxia for 1 to 24 h and control group ( with values from 1.032 to 2.390, P values above 0.05). Compared with that in control group, the protein expressions of p-cofilin of cells were greatly reduced in hypoxic group exposed to hypoxia for 1 to 24 h (with values from 4.563 to 22.678, P values below 0.01), especially exposed to hypoxia for 24 h. The protein expressions of cofilin of cells between normoxic group and hypoxic group at each time point were close ( with t values from -0.904 to 1.433, P values above 0.05). In hypoxic group, the protein expressions of p-cofilin of cells exposed to hypoxia for 1, 2, 6, 12, and 24 h were 0.87 +/- 08, 0.780 .05, 0.89 +/- 0.07, 0.68+0. 07, and 0.57 +/- 0.06, respectively, significantly lower than those in normoxic group (0.90 +/- 0.07, 0.97 +/- 0.06, 1.00 +/- 0.06, 1.00 +/- 0.05, and 0.99 +/- 0.05, with t values from 3.193 to 16.434, P values below 0.01). In control group, F-actin in the cytoplasm was abundant, most of it was in bunches. The trend of F-actin was disorderly in hypoxic group from being exposed to hypoxia for 1 h, shortened in length or even dissipated. The ratios of F-actin to G-actin of cells in hypoxic group exposed to hypoxia for 12 and 24 h (0.89 +/- 0.12 and 0.84 +/- 0.19) were obviously decreased as compared with that in control group (1. 00, with t values respectively 3. 622 and 3. 577, P values below 0.01). There were no obvious differences in the ratios of F-actin to G-actin of cells between hypoxic group exposed to hypoxia for 1, 2, and 6 h and control group ( with values from 0.447 to 1.526, P values above 0.05). In control group, cells were compact in arrangement, and ZO-1 was distributed continuously along the cytomnembrane. From being exposed to hypoxia for 2 h, cells became irregular in shape in hypoxic group. ZO-1 was distributed in discontinuous fashion along the cytomembrane with breakage in hypoxic group exposed to hypoxia for 24 h.

CONCLUSIONSHypoxia may cause the disorder of dynamic balance between F-actin and G-actin by inducing cofilin activation, which in turn leads to the changes in distribution of tight junction protein ZO-1 in intestinal epithelial cells.

Actin Depolymerizing Factors ; Actins ; Blotting, Western ; Caco-2 Cells ; drug effects ; physiology ; Epithelial Cells ; cytology ; drug effects ; Humans ; Hypoxia ; metabolism ; Intestinal Mucosa ; drug effects ; metabolism ; pathology ; Intestines ; Oxygen ; pharmacology ; Tight Junctions ; drug effects ; metabolism ; Zonula Occludens-1 Protein ; metabolism

Background Clinical researches showed that antimicrobial-resistance of bacterium is gradually serious.It is very important to master pathogen distribution and antimicrobial susceptibility after eye surgery.Objective This study was to investigate the distribution and drug sensitivity of pathogenic bacteria from ocular operative infection in 3-year duration to provide a reference for reasonable selection of antibiotics.Methods Sixtyfive positive specimens were obtained from 65 patients due to postoperative infection in Department of Ophthalmology of Shenzhou Hospital and Shengjing Hospital from 2009 January through 2011 December.The pathogenic bacteria were isolated and then the drug sensitivity test of conventional antibiotics was performed.Results Among the 65 ophthalmic specimens,35 strains of Gram-positive bacteria were isolated with the constituent ratio 53.8％,and 15 strains of Gram-negative bacteria accounted for 23.1％.Fungus was found in 15 cases with the constituent ratio 23.1％.Staphylococcus epidermidis accounted for 42.9％ in Gram-positive bacteria,and Pseudomonas aeruginosa accounted for 33.3％ in Gram-negative bacteria.Aspergillus was the main component of fungus,which accounted for 60.0％ of the proportion 3 strains of methicillin-resistant Staphylococcus aureus (MRSA) in 8 strains of Staphylococcus aureus,but no vancomycin-resistant Enterococcus (VRE) and vancomycin-resistant Staphylococcus (VRS) were seen.The in vitro drug sensitivity test showed that the main Gram-positive bacteria,Staphylococcus epidermidis and Staphylococcus aureus,were sensitive to linezolid,vancomycin,levofloxacin and ciprofloxacin with the sensitive rates ＞50.0％,and these bacteria were lowly sensitive to penicillin and erythromycin with the sensitive rates ≤30.0％.Pseudomonas aeruginosa appeared to be the main Gram-negative bacteria,showing a sensitive rate ≥ 80.0％ to tobramycin,amikacin,ceftazidime,cefepime,cefoperazone/sulbactam,piperacillin/tazobactam,imipenem and meropenem.Conclusions Staphylococcus epidermidis,Staphylococcus aureus,Pseudomonas aeruginosa and Aspergillus are primary pathogens in ocular infection after operation.Antibiotics such as penicillin,erythromycin,levofloxacin,cefotaxime seem to be relatively high resistant to eye infection.To choose sensitive antibiotics timely and reasonably is a key to the prevention and control of ocular infection after operation.

Objective:To investigate the relationship between clinical effects of acupuncture for elderly patients with sensorineural deafness and ear distending sensation.Methods:A total of 120 elderly patients with sensorineural deafness were randomly divided into a comprehensive treatment group,an acupuncture group and a Western medicine group,with 40 cases in each group.The acupuncture group received acupuncture treatment,the Western medicine group received oral mecobalamin tablets and the comprehensive treatment group received acupuncture plus acupoint injection and auricular acupoint sticking.The values of pure tone hearing threshold test of the three groups were observed before and after treatment,and the relationship between clinical effects and ear distending sensation was compared.Results:The total effective rate of the comprehensive treatment group was 82.5％ versus 67.0％ in the acupuncture group and 62.5％ in the Western medicine group.The inter-group comparisons showed statistically significant differences in the comprehensive treatment group versus the Western medicine group (P＜0.01) and the acupuncture group (P＜0.05).In the comprehensive treatment group,there were 23 cases (57.5％) with ear distending sensation,the clinical total effective rate was 86.9％;there were 17 cases (42.5％) without the sensation,the clinical total effective rate was 76.5％.In the acupuncture group,there were 24 cases (60.0％) with ear distending sensation,the clinical total effective rate was 71.0％;there were 16 cases (40.0％) without the sensation,the clinical total effective rate was 63.0％.In the Western medicine group,there were 21 cases (52.5％) with ear distending sensation,the clinical total effective rate was 66.7％;there were 19 cases (47.5％) without the sensation,the clinical total effective rate was 57.9％.The total effective rate of patients with ear distending sensation were higher than the rates of those without ear distending sensation in the three groups,but the differences were insignificant (all P＞0.05).Conclusion:The comprehensive therapy is one of the effective methods to treat elderly patients with sensorineural deafness.In the three groups of elderly patients with sensorineural deafness,the relief of ear distending sensation and the hearing loss were basically simultaneous,and the hearing recovery in the patients with ear distending sensation may be slightly better than that in those without ear distending sensation.Nevertheless,further research is needed.

AIM:Targeting of focal adhesion kinase (FAK) gene,we aim to construct FAK shRNA lentiviral vector and to identify its function on the growth of leukemic cells.METHODS:FAK shRNA was chemically synthesized,and inserted into a GFP-lentiviral plasmid through molecular biology methods.After packaged and concentrated,the lentiviral-FAK-shRNA-vector was transduced into a human leukemic cell line.FAK gene expression was detected by reverse transcriptional PCR and Western blotting.Cell apoptosis was measured by Annexin V labeling.RESULTS:The results showed that FAK shRNA was successfully inserted into the lentival vector,and the infection efficiency varied from 10% to 25%.Compared to the control vector (lentival vector without FAK shRNA),FAK shRNA inhibited the expression of FAK mRNA and protein by 40% and 70%,respectively.Moreover,the results of apoptosis experiment showed that the percentages of Annexin V+ cells in control vector group and FAK shRNA group were (4.19 ? 0.36) % and (8.48 ? 0.58) % respectively,the difference was statistically significant (P

Objective:To investigate the effects of sensitized sera on bone marrow transplantation and clarify the role of antibody in the process of rejection.Methods:Two hundred microlitres sensitized sera or non-sensitized sera were injected into normal BALB/c one day before transplantation.Ten millions (1?107) bone marrow cells from C57BL/6 were transfused to the recipients after lethal irradiation.The donor-reactive antibodies in recipients before transplantation were measured by complement-dependent cytotoxicity (CDC) method.Moreover,the survival analysis and engraftment evaluation of the recipients were carried post transplantation.Results:The CDC results showed that donor-reactive antibodies existed in the recipients which had received sensitized sera transfusion.Eighty percent (80%) of the recipients received sensitized sera transfusion died of bone marrow failure about 10 days post transplantation,while the recipients received non-sensitized sera transfusion were long-term alive.Furthermore,the hematopoietic recovery and percentage of donor chimera analysis declined along with time in the sensitized sera transfusion recipients,and there were significant differences compared with those in the non-sensitized sera transfusion recipients (P

This article introduced the neuromuscular activation characteristics of patients with chronic ankle instability during dif-ferent movement patterns, and explained the reasons of deficits of neuromuscular control in lower extremity muscle ac-tivity, kinetics, and kinematics, which aimed at further clarifying the mechanism of chronic ankle instability, and provid-ing theoretical basis for its rehabilitation training.

The reaction conditions of baicalin hydrolyzed into baicalein by a kind of thermophilic and sugar-tolerant beta-glucosidase were studied in this paper. The beta-glucosidase could catalyze baicalin into baicalein well in the acetic acid-sodium acetate buffer. The optimal enzyme activity was at 85 degrees C and pH 5.5. The enzyme was stable at the temperature less than 85 degrees C and pH range of 5-7.5. The maximum reaction rate V. and michaelis constant K. were 0.41 mmol x L(-1) x min(-1) and 3.31 mmol x L(-1) respectively. Different metal ions had different effects on the activity of enzyme. Na+ existing in acetic acid-sodium acetate buffer had an activation effect on enzyme. The enzyme activity was enhanced by the concentrations of glucose below 0.6 mol x L(-1), and was gradually inhibited when monosaccharide concentration was over 0.6 mol x L(-1). When the monosaccharide concentration reached 1.2 mol x L(-1), the inhibition rate of enzyme activity was about 50%, which showed good glucose tolerance. The good reaction conditions through the experiment have been determined as follows, the substrate: enzyme dose was 1 g: 0.2 mL, acetic acid-sodium acetate buffer pH 5.5, reaction temperature 85 degrees C, reaction time 10 h, and the enzymatic hydrolyzation ratio could reach 97%.
Biocatalysis ; Enzyme Stability ; Flavanones ; chemistry ; Flavonoids ; chemistry ; Glucose ; chemistry ; Hot Temperature ; Hydrolysis ; Kinetics ; beta-Glucosidase ; chemistry

The study was aimed to investigate the strategy of transfusion of allogeneic hematopoietic stem/progenitor cells (HS/PC) into marrow cavity of mouse model in sensitized transplantation. A sensitized BALB/c mouse model was established by repeated transfusion of allogeneic spleen cells. The normal BALB/c mice were used as non-sensitized controls. The non-sensitized or sensitized recipients were transplanted by transfusion of allogeneic HS/PCs into bone marrow cavity. The survival rate and hematopoietic recovery were monitored. Moreover, non-sensitized and sensitized sera were obtained and incubated with allogeneic HS/PC respectively, the percentage of dead cells was calculated using complement-dependent cytotoxicity (CDC) tests. The results showed that non-sensitized recipients got long-term survival after the transfusion of HS/PC into marrow cavity, and the hematopoietic recovery increased along with time. However, among the sensitized recipients, one mouse died of anesthetic accident, the other 9 mice (9/10) died within 2 weeks after the transfusion of HS/PC in marrow cavity, and the hematopoietic recovery declined along with time. Histopathologic analysis demonstrated that the sensitized recipients died of bone marrow failure. The results of CDC tests showed that the percentage of dead cells in non-sensitized and sensitized group was 7.80 ± 1.93% and 50.80 ± 3.12%, respectively, and the differences were statistically significant (p < 0.05), indicating sensitized sera were capable of impairing allogeneic HS/PC. It is concluded that the strategy of the marrow cavity transfusion of HS/PC can not enhance engraftment of allogeneic donor cells in sensitized recipients.
Animals ; Bone Marrow ; Hematopoietic Stem Cell Transplantation ; methods ; Hematopoietic Stem Cells ; cytology ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Transplantation, Homologous

Osteoarthritis (OA) is a complex chronic progressive disease attacked by biological and mechanical factors and a result from the anabolic and catabolic imbalance in chondrocyte, subchondral bone and extracellular matrix(ECM). Etiology and pathological of OA are not yet entirely clear. The degradation and destruction of collagen II caused by matrix metalloproteinase -13 (MMP-13) is considered the core factor in the occurrence and development of OA. The research of MMP-13 inhibitor provide ideas and methods for the treatment of OA. In this article,the role and determination of MMP-13 in OA and the development prospect of MMP-13 inhibitor in the treatment of OA research progress were reviewed.
Animals ; Collagen ; metabolism ; Humans ; Matrix Metalloproteinase 13 ; analysis ; physiology ; Matrix Metalloproteinase Inhibitors ; therapeutic use ; Osteoarthritis ; drug therapy ; etiology

Adult ; Decompression, Surgical ; Female ; Follow-Up Studies ; Humans ; Intervertebral Disc Displacement ; surgery ; Laser Therapy ; Lumbar Vertebrae ; surgery ; Magnetic Resonance Imaging ; Male ; Middle Aged ; Traction ; methods