Animals ; Dementia, Vascular ; blood ; psychology ; Erythrocyte Count ; Learning ; drug effects ; Male ; Memory ; drug effects ; Peptides ; pharmacology ; Rats ; Rats, Sprague-Dawley

Animals ; Ear Canal ; surgery ; Female ; Gene Expression ; Hypothalamus ; metabolism ; Nerve Block ; Pregnancy ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley ; Receptors, N-Methyl-D-Aspartate ; genetics ; metabolism

Animals ; Female ; Gene Expression Regulation, Developmental ; Inferior Colliculi ; metabolism ; N-Methylaspartate ; genetics ; metabolism ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley ; Receptors, N-Methyl-D-Aspartate ; genetics ; metabolism

Objective To evaluate the ability to kill human urothelial carcinoma T24 cells selectively in vitro by celecoxib combined with ZD55-IL-24 and explore the effectiveness for this combination use.Methods The EGFP expression of cells was observed by fluorescence microscope.The human umbilical vein endothelial cells (HUVEC) were used as crotrols.The expression of IL-24 mRNA was detected by RT-PCRwhen the cells were transfected by ZD55-EGFP or ZD55-IL-24.After transfected by ZD55-IL-24 and treated by celecoxib,the inhibition effect on cells was measured by MTT assay,and the apoptosis rate was examined by flow cytometry.Results The fluorescence in T24 and HUVEC can be observed 24h after ZD55-EGFP transfection and the fluorescence intensity was increased corresponding with the times.Fluorescence intensity in T24 cells showed higher than that in HUVEC group at the same times.The result of RT-PCR showed that the T24 cells expressed higher level of IL-24 mRNA than HUVEC group at the same time when the cells were transfect by ZD55-IL-24 (P ＜ 0.01).The inhibition rate of ZD55-IL-24 combined with celecoxib group was significantly higher than other groups (P ＜ 0.001).The inhibition rate of T24 cells in each group was significantly higher than HUVEC group (P ＜ 0.01).The flow cytometry results indicated that celecoxib combined with ZD55-IL-24 had the highest apoptosis rate on T24 cells than other single use group.Apoptosis rate of T24 cells showed a higher than HUVEC cells (P ＜ 0.01).Conclusion Celecoxib combined with ZD55-IL-24 can inhibit T24 cells proliferation at a greatest degree and this effect may be contributed to apoptosis.

AIM: To investigate the use of therapy ultrasound to enhance nonviral gene delivery. METHODS: Endothelial cells (EC) and vascular smooth muscle cells (VSMC) were cultured in 6-well plates. Plasmid (pcDNA3.1/His/LacZ) with or without microbubbles at the different concentrations was transfected into the cells with the use of ultrasound for 1 min at 2 MHz, 1.8 mechanical index (MI). Additional controls included ultrasound alone, microbubble alone and microbubble plus plasmid. The rate of blue cells and the activities of ?-Gal were measured. In addition, cell viability was detected with different time from 1 to 30 min of ultrasound irradiation and the different concentrations of microbubbles. RESULTS: In the group of ultrasound with microbubble, the rate of blue cells and activity of ?-Gal markedly increased by 60% and 9-fold, respectively. Microbubbles at concentration of 10% led to the highest transfection effect. Ultrasoud exposure at 1 to 30 minute had no cell toxic effects, while microbubbles at the concentration of 50% had significant effect on cell survival. CONCLUSIONS: Albumin-coated microbubbles markedly enhance gene delivery by therapeutic ultrasound-mediated microbubble destruction, which can be used as a safe and practicality vectors in gene therapy.

Objective Insulin autoantibody (IAA) is known to exist in sera of type 1 diabetes mellitus (T1DM) patients and pre-T1DM individuals. The aim of this study was to establish a novel microtiter plate radioimmunoassay (RIA) for IAA and evaluate its clinical value. Methods Diluted 125Ⅰ-insulin was mixed with 5 ul serum samples in a 96-well microtiter plate and then incubated for 72 h on an orbital plate shaker (4℃). The immunocomplexes were transferred to another protein a coated Millipore plate, and then the plate was washed with Tri-Buffered Saline Tween-20 (TBT) buffer. Counts per minute (CPM) was measured with liquid scintillation and luminescence counter. The positive cut-off point of IAA index was defined as ≥0.06 based on the 99-percentile of the distribution in 317 healthy individuals. The specificity and sensitivity of the assay were calculated from the samples provided by the fourth Diabetes Autoantibodies Standardization Program (DASP 2005). The IAA levels were determined in 71 T1 DM and 551 newly diagnosed type 2 diabetes (T2DM) patients, and 317 healthy controls. The t test, non-parametric test, x2 test and linear correlation analysis were performed on the data using SPSS 11.5 software. The concordance rate was estimated with Kappa value. Results (1) The optimized testing condition was described as 2×104 CPM of 125Ⅰ-insulin, 5 ul serum sample and slowly horizontal shaking for 72 h. (2) The intra-assay CV was 4.8%-8.9% and inter-assay CV was 6.4%-10.5%. Based on DASP 2005 samples, the specificity and sensitivity of the assay were 97% (97/100) and 50% (25/50), respectively. Ninety-six serum samples with different IAA levels were selected and tested to compare between our new method and a domestic IAA RIA kit. The results showed that the IAA indices from the two methods were positively correlated (r= 0.678, P<0.001). The concordance rate was 72.9 %(Kappa value=0.402). There were 25 samples with discordant results, which were positive for IAA titer using the corresponding microtiter plate RIA but negative using the novel RIA kit. (3) In TIDM group the positive rate of IAA was 19.7% (16/71), higher than the healthy controls (0.9%, x2=54.36, P<0.001). The subgroup of T1DM children (with 0-9 years) showed the highest IAA positive rate (55.6% ,x2=4.85, P<0.05). In T2DM group the frequency of IAA was 1.5% (8/551), which had no significant difference comparing with that of healthy controls (x2= 0.95, P >0.05). Conclusions Our proposed microtiter plate RIA method for IAA is highly sensitive and specific, likely to be feasible for clinical application. The frequency of IAA is high in children with T1DM.

OBJECTIVETo summarize the clinical application result of the selective nerve root blocks in limited operation of the lumbar spine.

METHODSFrom January 2008 to October 2012,68 patients with lumbar spinal canal stenosis with multiple levels were underwent the selective nerve root blocks in limited operation of the lumbar spine,including 47 males and 21 females with an average age of 56 years old ranging from 45 to 80. After never roots blocks,64 cases were positive for limited operation of the lumbar spine; the other 4 cases were negative and abort the operation.

RESULTSThe nerve roots block operation smoothly and no complications related to the nerve roots block occurred. There was no neurologic injury complication in this study. Follow-up period ranged from 16 to 45 months postoperatively (means, 32 months). The recovery effect was calculated with Macnab scores, the result was excellent in 44 cases, good in 18 cases, poor in 1.

CONCLUSIONOperative treatment for lumbar spinal canal stenosis with multiple levels is focused on the areas causing symptomate neural compression rather than prophylactic decompression at areas of nonsymptomatic disease. Application of selective nerve root blocks can accurately judge the responsible vertebral body and pain source and improve the curative effect of limited operation of the lumbar spine

Aged ; Aged, 80 and over ; Female ; Humans ; Lumbar Vertebrae ; surgery ; Male ; Middle Aged ; Nerve Block ; methods ; Spinal Nerve Roots ; Spinal Stenosis ; surgery

It was constructed that a genomic DNA library from Lactobacillus sp. MD-1 yielding D, L-lactic acid. The gene encoding L-lactate dehydrogenase (L-LDH) was cloned from the genomic library of strain MD-1 by complementation in E. coli FMJ144 which was lactate dehydrogenase and pyruvate-formate lyase double defective mutant. The nucleotide sequence of the ldhL gene predicted a protein of 316 amino acid starting with ATG. The putative molecular weight of the L-LDH amino acid sequence was 33.84kD. A putative typical promoter (-35 and -10 boxes) had been observed in the 5' noncoding region. An rho-independent transcriptional terminator has been observed in the 3' noncoding region. Three highly conserved regions (Gly13 approximately Asp50, Asp73 approximately Ileul00 and Asn123 approximately Arg154) with several conserved residues had been identified. Gly13 approximately Asp50 was NADH-binding site domain. Asp73 approximately Ileu100 and Asn123 approximately Arg154 were reported to be the active site domains. The ldhL and the L-LDH of Lactobacillus sp. MD-1 showed the low identity and similarity with other Lactobacilli, and the highest percentage were 61.9% and 68.9% respectively. All the above indicated this gene is a novel ldhL.
Amino Acid Sequence ; Base Sequence ; Cloning, Molecular ; L-Lactate Dehydrogenase ; chemistry ; genetics ; physiology ; Lactobacillus ; genetics ; Molecular Sequence Data

The aim of this study was to investigate the inhibitive effect of artesunate (ART) on CML cell line K562 and its influence on VEGF expression in vitro. Human CML cell line K562 cells were cultured in RPMI 1640 medium supplemented with 10% heat-inactivated fetal calf serum. All cells were cultured in a humidified atmosphere of 5% CO2 at 37.0 degrees C. K562 cells in logarithmic growth phase were collected and seeded in RPMI-1640 medium, and were treated with ART. At the indicated time points, viable cells were counted by trypan blue exclusion method. Each assay was triplicated. K562 cells were treated with ART at different concentrations. Morphological changes were observed with invert microscope. VEGF expression in K562 cells treated with ART at different concentrations and in the control group were detected by enzyme-linked immunosorbent assay (ELISA). The results indicated that ART obviously induced growth inhibition in K562 cells. The relationship between cell inhibition rates and the concentrations of ART showed a dose-dependent manner (p < 0.01). VEGF expression of K562 cells treated with ART at different concentrations decreased significantly (p < 0.01). No significant change of VEGF expression in control group was observed (p > 0.05), while VEGF expression was down-regulated significantly in experiment groups (p < 0.01). The inhibition rate of K562 cells increased in time and concentration-dependent manners. In K562 cell lines treated with ART, VEGF expression was up-regulated at first and then down-regulated to a lower level. It is concluded that ART inhibits k562 cell proliferation in a dose and time dependent manner. The mechanism underlying the inhibitive effect of ART on K562 cells may be realized through down-regulation of VEGF expression.
Antineoplastic Agents, Phytogenic ; pharmacology ; Artemisinins ; pharmacology ; Cell Proliferation ; drug effects ; Down-Regulation ; drug effects ; Humans ; K562 Cells ; Vascular Endothelial Growth Factor A ; genetics ; metabolism

OBJECTIVETo summarize the clinical application results of the repair soft tissue defect in contralateral leg with a cross-leg soleus muscle flap pedicle transplantation.

METHODSFrom January 2008 to January 2013, 8 patients with soft-tissue defect in lower leg underwent reconstruction with a cross-leg soleus muscle flap pedicle transplantation (without microvascular anastomoses). There were 7 males and 1 female, aged from 20 to 49 years old with an average of 31.8 years. The operative time after injury was from 2 to 8 weeks with the mean of 46 days. The soleus muscle flap was transposed across to the contralateral leg defect area, then immediate to perform the coverage of the muscle flaps by a meshed split-thickness skin graft. The donor site was closed directly.

RESULTSAll the muscle flaps had survived completely. In one case, recipient area edge had a less exudate from drainage hole everyday, the incision spontaneously was healed after 2 week's changing dressing. Follow-up period ranged form 1.5 to 4 years with an average of 2.5 years postoperatively. The tibia and fibula fractures were healed well. A good contour was achieved at the recipient area. According to LEM standard, 2 cases got excellent results, 5 good and 1 fair.

CONCLUSIONSoleus flap pedicle transplantation is very suitable to repair the soft tissue defect of the injuried leg only one main blood vessel, and can reduce the damage of donor area.

Adult ; Female ; Humans ; Leg Injuries ; surgery ; Male ; Middle Aged ; Muscle, Skeletal ; Soft Tissue Injuries ; surgery ; Surgical Flaps