Objective To probe a selective cultural method for bovine retinal endothelial cells (BREC) and pericytes (BRP) in vitro. Methods With the isolation of active retinal blood vessels, BREC were cultured in a fibronectin coated substrate and Dulbecco′s Modified Eagle′s Medium (DMEM) supplemented with 10% human serum and 100 ?g/ml heparin, while homogeneous cultures of retinal pericytes were obtained when isolated microvessels were seeded to uncoated dishes and grown in DMEM supplemented with 20% fetal bovine serum. BREC were identified by acetylated-low density lipoprotein (Dil-Ac-LDL) incorporation and immunohistochemical method of Von Willebrand factor, while BRP were identified by immunohistochemical method of ?-isoform of smooth-muscle actin. Results The purity of selectively cultured BREC and BRP was more than 98%, being reproducible. BREC got together around the microvessel fragments with the small-cyprinoid-like configuration at first,and could phagocytize Dil-Ac-LDL with the expression of fluorescence in cytoplasm. The expressions of Von Wllebrand factor and ?-isoform of smooth-muscle actin were positive and negative respectively in BREC, while were negative and positive respectively in BRP. Conclusion BREC and BRP with high purity can be obtained by using selective culture and coated-dishes respectively which are simple and repeatable methods.

Objective To study the effects of advanced glycation end (AGEs) products induced by bovine serum albumin (BSA) on the survival and the morphology of bovine retinal endothelial cells (BREC) and pericytes (BRP). Methods BSA with the final concentration of 50 mg/ml was incubated in PBS, containing 500 mmol/L D-glucose, for 12 weeks under 37℃. AGEs-BSA was purified by Sephacryl S-300 column chromatography and was confirmed by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). The concentration of AGEs-BSA was determined by the method of commassie protein assay. In order to detect the toxic effects of AGEs-BSA on cultured BREC and BRP, groups of AGEs-BSA and BSA with different concentration and untreated control were set up. Phase contrast microscope was used to observe the effect of AGEs-BSA and BSA (with the concentration of 500 ?g/ml and actuation duration of 48 hours) on morphology of BREC and BRP. Results[WTBZ] As the dosage of AGEs-BSA increased, the number of inhibited cells increased. When the concentration of AGEs-BSA was 500 ?g/ml, the inhibited BREC in AGEs-BSA group was (72.8?15.9)% of which in untreated control group, and the inhibited BRP was (64.8?9) % of which in untreated control group. AGEs-BSA with low concentration promoted the proliferation of endothelial cells, but there was no significant difference between AGEs-BSA and the control group (P=0.231). Inhibited proliferation and abnormal morphology were seen under the phase contrast microscope while the normal morphology of cells was found in BSA and control group. Conclusion AGEs-BSA with the high concentration may inhibit the growth of both BREC and BRP, which leads the loss of BRP and damage of vascular function. These results suggest that nonenzymatic glycosylation plays a major role in diabetic complications.

Objective To investigate the clinical features , treatment regimen , and prognosis evaluation of tertiary peritonitis (TP). Methods Seventy-eight cases with TP were randomly enrolled into 2 groups, including the simple western medicine-treated group (32 cases) and the integrated traditional Chinese and western medicine-treated group (46 cases). The prognoses were evaluated according to the acute physiology and chronic health evaluationⅢ (APACHEⅢ, APⅢ) scoring. Results The mortality rate was 71.9% (23 of 32) in patients received the simple western medicine and was 32.6%(15 of 46) in patients received the integrated traditional Chinese and western medicine with significant difference between these two groups (P < 0.01). There was a significant correlation between AP Ⅲscore and actual mortality (r=0.73,P<0.01), and predicted mortality (r=0.76, P<0.01). Conclusions The therapeutic effect is acceptable and satisfactory for the TP patients received the integrated traditional Chinese and western medicine. The AP Ⅲ scoring system can be used to predict the prognosis of TP patients.

Objective To investigate if insulin can affect the expression of vascular endothelial growth factor (VEGF) in the retina of streptozotocin-induced diabetic rats. Methods A total of 60 male Sprague-Dawley rats were randomly divided into sodium citrate buffer control group (CIT-CON, n= 30) and STZ-induced diabetic group (STZ-DM, n=30). At the 16th week, 24 rats from CIT-CON group at random were randomly divided to group A (sodium citrate buffer control group, n = 12) and group B (sodium citrate buffer plus insulin group, n= 12). The remaining 6 rats from as CIT-CON group served as negative control. At the same time, 24 rats from STZ-DM group at random were randomly divided to group C (STZ-induced diabetic group, n= 12) and group D (STZ-induced diabetic plus insulin group, n= 12). The remaining 6 rats from STZ-DM group also served as negative control. 4 IU of insulin was injected subcutaneously to rats of group B and D. Immunohistochemistry, Western blot and Real-time polymerase chain reaction (RT-PCR) were used to measure the expression level of VEGF protein and mRNA respectively. RESULTS Insulin significantly increased the VEGF mRNA (7.71 ± 0.25 vs 5.36 ±0. 37, t test P< 0. 05) and protein expression (0. 4925 ± 0. 0122 vs 0. 4272 ± 0. 0110, t test P< 0. 05) in the retina of CIT-CON rats.However, in retina of STZ-DM rats, insulin had no effect on VEGF mRNA (8. 92±0. 27 vs 9. 05±0. 28, t test, P>0. 05) and protein expression (0. 5152±0. 0109 vs 0. 5099±0. 0100, t test P>0.05). Conclusions Insulin had no effect on VEGF expression in the retina of STZ-DM rats.

BACKGROUND:The microcosmic and submicroscopic organizations of tissue engineering scaffold matedals’superficial structure have all important effect on the eell adhesion and growth.By means of nano.Technique and three-dimensional porous technique,the resultant nano-hydroxyapatite/collagen(n-HAC)call imitate the component and microstructure of natural bone.OBJECTIVE:To observe the biocompatibility of human bone m arrow mesenchymal stem cells(MSCs)cultured in vitro with nHAC.DESIGN,TIME AND SETTING :Single samples observation was performed in the Experimental Center of School ofMedical Technology,Jiangsu University from September 2005 to December 2006. MATERIALD:nHAc was provided by the Material Science and Engineering Department of Tsinghua University.Humanbone marrow mesenchymal stem cells were derived from healthy adult volunteers.All the subiects signed the informedconsents. METHODS:Whole bone marrow culture and successive adherence method was used to culture MSCs in vitro,and the cells were then induced to differentiate into the phenotype of osteoblasts by the revulsants(methylprednisolone,vitamin C,β-glycerophosphate and basic fibroblast growth factor).MSCs at passage 3 were co-cultured with nHACfor 14 days.MAIN OUTCOME MEASURES:The cytological characteristics of the osteoblast were identified throue,alkalinephosphatase immunohistochemistry method and Von Kossa stain.The growth condition with or without nHAC wasevaluated through invert microscope and scanning electron microscope,respectively.RESULTS:The cultured MSCs proliferated into uniform fibroblast-like cells rapidly.MSCs reached confluence and started to form multilayers averaging from 10 to 12 days,passaged stably as well.Then the MSCs passaged from 7 to 9 days.Cytochemistry evaluation showed that MSCs in induced culture were positive for alkaline phosphatase and Von Kossa stain,and deposited calcified matrix.It showed a typical ostcoblast feature in morphology and biology.In coculture model ofMSCs with nHAC,cells would attach to the inner surface of nHAC.At 8 days,the osteoblasts proliferated in the nHAC and the secretion of the matrix was observed.Lots ofcells adheredon the surfaceand pores of nHAC at 14 days.There wereextensive prominent connections among cells. CONCLUSION:THE nHAC is suitable for MSCs to adhere,grow and proliferate,with a good compatibility.

Objective To investigate the effects of total nutrient admixture (TNA) on the recovery of patients with gastric cancer after radical gastrectomy.Methods The clinical data of 50 patients with gastric cancer who were admitted to the Affiliated Hospital of Luzhou Medical College between March 2013 and March 2014 were retrospectively analyzed.Among 50 patients receiving radical gastrectomy,26 patients receiving TNA were allocated to the experimental group and 24 patients receiving conventional fluid infusion were allocated to the control group.Patients in the experimental group received the nutritional support therapy using TNA at preoperative day 5 and at postoperative days 1-5,and patients in the control group received the postoperative intravenous rehydration including water,glucose,electrolyte,vitamins and micro elements.The nutritional indexes [albumin (Alb),prealbumin,transferrin and hemoglobin (Hb)],time to anal exsufflation,incidence of complications (wound infection,anastomotic leakage,blooding and intestinal obstruction) and duration of hospital stay were observed before nutritional support therapy and at postoperative day 8.The count data were analyzed using the chi-square test.The chi-square value of correction for continuity was used when 1 ≤ minimum theoretical frequency ≤ 5.The measurement data with normal distribution were presented as (x) ±s and analyzed using the t test or repeated measures ANOVA.The ordinal data were analyzed by the analysis of variance.Results The Alb,prealbumin,transferrin and Hb in the experimental group were (38.6 ± 2.0) g/L,(281 ± 33) mg/L,(2.5 ± 0.9) g/L and (111 ± 20) g/L before nutritional support therapy and (38.2 ± 1.9) g/L,(277 ± 16) mg/L,(2.3 ± 1.1) g/L and (112 ± 37) g/L at postoperative day 8,respectivley.The Alb,prealbumin,transferrin and Hb in the control group were (38.3 ±2.4) g/L,(287 ± 34) mg/L,(2.4 ± 1.1) g/L and (107 ± 21) g/L before nutritional support therapy and (30.3 ±2.3) g/L,(190 ± 41) mg/L,(1.6 ± 0.3) g/L and (93 ± 22) g/L at postoperative day 8,respectivley.There were significant differences in the nutritional indexes at postoperative day 8 between the 2 groups (F =174.042,95.637,9.529,4.919,P ＜ 0.05).The time to anal exsufflation in the experimental group were (52 ± 11) hours,which was significantly different from (70 ± 12) hours in the control group (t =-5.176,P ＜ 0.05).The incidence of complications was 15.4％ (4/26) in the experimental group,which was significantly different from 58.3％ (14/24) in the control group (x2=6.460,P ＜0.05).Patients with complications in the 2 groups were cured by anti-infective or symptomatic treatment.The duration of hospital stay was (9 ± 3) days in the experimental group and (12 ± 4) days in the control group,with a significant difference between the 2 groups (t =-2.912,P ＜ 0.05).Conclusion TNA can improve the nutritional status of patients after radical gastrectomy in a short time.It could help patients to get through the perioperative period smoothly,and enhance the postoperative recovery.

Objective To compare the efficacy of total pelvic floor reconstruction with mesh patch and vaginal hysterectomy with anterior and posterior vaginal wall repair for presby-pelvic floor dysfunction patients.Methods We randomly chose 92 pelvic floor dysfunction inpatients of our hospital from September 2007 to February 2009 and divided them into 2 groups:total pelvic floor reconstruction group(n=70) underwent total pelvic floor reconstruction with mesh patch[12 patients of this group complicated with stress incontinence,4 patients received tension free vaginal tape-obturator(TVT-O) simultaneously];control group(n=22) received vaginal hysterectomy with anterior and posterior vaginal wall repair(3 patients of this group complicated with stress incontinence).A follow-up for 3 to 18 months after the surgery were taken in all the patients.Results In total pelvic floor reconstruction group,no recidivate case was found and the FOP scores and cure rate were 0 and 100% respectively.In control group,there were 7 recidivate cases,all of who were primary vagina anterior wall bulge patients.The cure rate of control group was 68%;For the patients complicated with stress incontinence,they were all cured by TVT-0 or by ascending the front patch in total pelvic floor reconstruction group,but 1 case was as before operation and 2 cases were even worse in control group.Conclusion Total pelvic floor reconstruction with vicarious materials had a definite efficacy for presby-pelvic floor dysfunction.

OBJECTIVE To determine the distribution of bacterial flora in hospital infection and to provide laboratory(evidence) for controlling hospital infection and selecting rationally antibiotics in clinic practice.METHODS All(isolates) were identified by routine procedure.MRSA and ESBLs-producing rate of Escherichia coli and Klebsiella pneumoniae were(examined.) RESULTS Among all these clinical infectious specimens,there were 202 strains of Gram negative bacilli,(accounting) for 40.9%(202/495);166 strains of fungi,accounting for 33.5%;621 strains of Gram positive cocci,for 20.6%(102/495).Candida albicans,E.coli,Pseudomonas aerugionosa,C.tropicalis and C.glabrata took the first five bacteria in infection.Analysis of drug resistant bacteria suggested that the isolated rate of ESBLs-producing strains in Staphylococcus aureus be 47.6%,be CNS in MRCNS 78.1% and MRSA in SA be 42.3%.CONCLUSIONS Multidrug resistance and fungus infection are the main risk factors in our hospital.We must improve means of treatment on clinical work and use antibiotic rationally to reduce the infection rate.

Objective To investigate the precision and trueness of results from six imported commercial systems for measurement of gamma-glutamyltransferase (GGT) in serum in order to provide reference for the clinical laboratories to verify the target accuracy. Methods Five fresh frozen human serum samples that differed in catalytic concentration were analyzed in two candidate domestic reference laboratories and the target values for GGT were assigned using IFCC reference measurement procedure. The same samples were tested by six commercial systems which were calibrated using the matched calibrator. Each system consisted of five instruments in five laboratories, which had been well maintained before measurement. The data was collected. Precision from the same manufacturer and different manufacturers and biases between target values and mean values from each system were calculated. Results The differences of the mean values for five levels of commercial systems varied from 16. 1% to 35.4%. For the five levels, the coefficients of variation (CVs) of the results from all measurement system were from 5.3% to 8. 8% , and CVs from each level were A 2. 17%- 5.07%, B 4. 21%-10. 98%, C 0. 52%-2. 38%, D 1.35%-2. 59%, E 0. 23%-1..54%-), F 1.83%-2. 38%. Biases between the mean values of each commercial systems and the target values were A 0. 43% -8.41% ), B -1.49% - -13.04% ), C 11.20% -17.73% ), D 0. 19% -4. 62% ), E -0. 30% - -2. 63% ), F 4). 46%-7.90%, respectively. The investigation showed that biases of two manufacturers were less than a quarter of the total allowable error (TAE) of The Clinical Laboratory Improvement Amendments of 1988 (CLIA'88) in the whole range of investigated concentrations and the other two manufacturers' biases could meet a quarter of TAE in a relative limited range. The biases of two manufacturers were near or more than half of TAE in most levels. It also revealed that the biases of more than half of manufacturers were more than a quarter of TAE in the low or high level of investigated concentrations. Conclusions The mean values of each manufacturer were significantly different. The variances of commercial systems from different manufacturers were much higher than those from the same manufacturer. Some imported commercial systems for measurement GGT should be better calibrated with the reference method, especially in the whole measurement linearity.

To enhance the immunogenicity of DNA and adenoviral vector vaccines expressing HIV-1 subtype B gp160, human interleukin 15 (hIL15) DNA adjuvant (pVR-hIL15) was constructed. BALB/c mice received DNA prime/protein boost immunization with pVR-HIVgp160/Ad5-HIVgp160 alone or combined with pVR-hIL15. Cellular and humoral immune responses were evaluated by IFN-gamma enzyme-linked immunosorbent spot assay and enzyme-linked immunosorbent assay, respectively. Compared with those immunized with vaccines alone, the mice immunized with vaccines combined with pVR-hIL15 had significantly increased specific cellular response and antibody titer (P < 0.05). It suggests that the IL15 DNA adjuvant can enhance the immune responses induced by prime-boost regimen using DNA and adenoviral vector encoding HIV-1 subtype B gp160.
Adenoviridae ; genetics ; Adjuvants, Immunologic ; Animals ; Antibodies, Viral ; immunology ; Antibody Specificity ; Female ; Genetic Vectors ; genetics ; HIV Envelope Protein gp120 ; immunology ; HIV Envelope Protein gp160 ; genetics ; immunology ; HIV Envelope Protein gp41 ; immunology ; Humans ; Immunity, Cellular ; Immunity, Humoral ; Interleukin-15 ; genetics ; Mice ; Mice, Inbred BALB C ; Vaccines, DNA ; genetics ; immunology