Objective To investigate the role and the mechanism of ppk1 gene (coding for polyphosphate kinase 1) in oxidative stress resistance in uropathogenic Escherichia coli (UPEC).MethodsMutant strains with ppk1-deletion (△pk1) and complemented strains (△pk1-C) were constructed based on the UPEC strain CFT073.A comparative analysis was conducted to analyze survival rates of CFT073, △pk1 and △pk1-C strains at different time points while they were under oxidative stress.Differences in protein expression between CFT073 and △pk1 strains were analyzed using mass spectrometric analysis.Differences between CFT073 and △pk1 strains in expression of katG and katE genes were analyzed using real-time quantitative RT-PCR.Results The survival rate of △pk1 strains was lower than that of CFT073 strains at every time point, while the survival rate of △pk1-C strains was basically the same as that of CFT073 strains.Gel image analysis and mass spectrometric analysis revealed that six proteins were down-regulated and one was up-regulated in △pk1 strains as compared with those in CFT073 strains.Expression of the catalase-coding genes katG and katE in △pk1 strains were respectively (20.5±8.2)% and (20.9±6.9)% of those in CFT073 strains (P<0.05).Conclusion The ppk1 gene plays an important role in oxidative stress resistance in UPEC by modulating the expression of catalase-coding genes katG and katE.

Objective To investigate the correlation of plasma matrix metalloproteinase-3 (MMP-3)levels and MMP-3 Lys45Glu (rs679620) polyrnorphism with ischemic stroke and its TOAST subtypes.Methods The patients with large artery atherosclerotic stroke (LAA) and small artery occlusion stroke (SAO)according to TOAST etiological typing (ischemic stroke group) and healthy subjects (control group) were enrolled.The enzyme-linked immunosorbent assay was used to detect plasma MMP-3 level.The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to detect the genotypes of MMP-3 Lys45Glu.Results A total of 233 patients with ischemic stroke were enrolled,in which 162 were LAA and 71 were SAO; 200 healthy subjects were taken as controls.The plasma MMP-3 level in the ischernic stroke group was significantly higher than that in the control goup (253.99 ± 75.02 ng/ml vs.196.38 ± 78.17 ng/ml;t =7.813,P=0.000).The plasma MMP-3 level in the LAA group (262.81 ±69.23 ng/ml) was significantly higher than those in thegroups of SAO (233.85 ± 83.90 ng/ml,P =0.008) and control (P =0.000),and the plasma MMP-3 level in the SAO was also significantly higher than that in the control group (P =0.000).Multivariate logistic regression analysis showed that the increased serum MMP-3 level was an independent risk factor for ischemic stroke (odds ratio [OR] 1.012,95％ confidence interval [CI] 1.008-1.015; P =0.000).There was no significant difference in the frequencies of genotype (x2 =2.085,P =0.353) and allele (x2 =2.29,P =0.130) of MMP-3 Lys45Glu between the ischemic stroke group and the control group.However,there were significant difference in MMP-3 Lys45Glu genotype frequencies among.the groups of LAA,SAO and control (x2 =10.39,P=0.034).The AA + GA genotype frequency in the LAA group was significant higher than those in the groups of SAO (65.4％ vs.49.3％ ;x2 =5.375,P =0.020) and control (65.4％ vs.54.0％ ;x2 =4.84,P =0.028).There was no significant difference in the allele frequencies among the groups of LAA,SAO and control (x2 =3.887,P =0.143).Multivariate logistic regression analysis showed that MMP-3 Lys45Glu polymorphism was an independent risk factor for LAA (OR 1.783,95％ CI 1.183-2.688; P =0.006).The plasma MMP-3 level in patients with the genotypes AA (n =73),GA (n =176) and GG (n =184)were 235.70 ± 70.85 ng/ml,(244.20 ± 85.90 ng/ml and 207.98 ± 77.61 ng/ml.There were significant difference in the plasma MMP-3 levels among the patients with the genotypes AA,GA and GG (F=9.682,P =0.000).The plasma MMP-3 level in the patients with the genotype AA + GA was significantly higher than that in patients with genotype GG (241.71 ± 81.73 ng/ml vs.207.98 ± 77.61 ng/ml; t =4.336,P =0.000).Conclusions The plasma MMP-3 level increased in patients with LAA or SAO,especially in the patients with LAA.The MMP-3 Lys45Glu polymorphism might be associated with the plasma MMP-3 level and LAA.

Objective This study was aimed to analysis the relationship of BK polyomavirus ( BKV) and hemorrhagic cystitis ( HC ) in patients who received hematopoietic stem cell transplantation (HSCT).Methods Data of 80 patients who received HSCT and took regular urine test every week from June 2015 to April 2018 in the Third Xiangya Hospital of Central South University was retrospectively analyzed, they were 35 females and 45 males (aged 20-40 years, median age 30 years), and 31 cases with acute myeloid leukemia ( AML) , 24 cases with acute lymphoblastic leukemia ( ALL) , 15 cases with aplastic anemia ( AA ) , 4 cases with chronic myeloid leukemia ( CML ) , 6 cases with other diseases such as myelodysplastic syndrome ( MDS) in the study population.The positive rate and incidence of HC were analyzed.Patients who infected with the BK virus were divided into HC group and non -HC group according to occurrence of HC.BK viral load were compared in two groups .Urine BK viral load were analyzed after logarithmic transformation.Data that conforms to a normal distribution is expressed as mean ± standard deviation.t test, ANOVA and ROC curve were used to statistical analysis .Skewed data is expressed in median ( interquartile range ) , non-normal distribution parameters were compared by Wilcoxon test .Results Among 80 patients, 43 recipients (53.75％) became urinary BK positive, with 19 patients developed HC (23.75％), all of the 19 HC patients have urinary BK positive , and none of 37 BK-negative patients developed HC;the urine BKV level of the initial time and the peak time in HC group were (7.59 ±2.46) lg copy/ml, (10.56 ±1.71) lg copy/ml, the urine BKV level of the initial time and the peak time in non-HC group were (5.75 ±2.10) lg copy/ml,(7.31 ±2.29) lg copy /ml.The urine BKV level of the initial time and the peak time in HC group was higher than in non-HC group ( t=2.642, P=0.012 and t=5.147, P=0.000 respectively), when analyzing the urine BKV level of the initial time in HC group and non-HC group, the best threshold is 5.23 lg copy/ml(1.68 ×105 copy/ml),with a sensitivity of 84.20％and specificity of 54.17％, when analyzing the urine BKV level of the peak time in HC group and non-HC group, the best threshold is 9.75 lg copy/ml(5.62 ×109 copy/ml),with a sensitivity of 84.20％and specificity of 83.33％, area under curve of each other were 0.728 (95％ CI 0.575-0.881) and 0.875 (95％ CI 0.769-0.981) respectively.Conclusions The BK viral load is closely related with HC in HSCT patients .The cut-off level of 1.68 ×105 copy/ml when analyzing the urine BKV level of the initial time , and the cut-off level of 5.62 × 109 when analyzing the urine BKV level of the peak time , help to forecast or auxiliary diagnose HC .

OBJECTIVECalcium plays an important role in the impairment of heart function and arrhythmia under the condition of acute hypoxia, but the mechanism is different from that of chronic hypoxia. This study aimed to evaluate the effect of chronic hypoxia on the expression of calmodulin (CaM) and calcicum/calmodulin-dependent protein kinase II (CaMKII) and the calcium activity in myocardial cells through an animal model of chronic hypoxia in order to get a deeper sight into the mechanism.

METHODSA chronic hypoxia model of the rat was prepared by hypoxia exposure (FiO2=10%). The expression of mRNA and protein of CaM and CaMKIIgamma and CaMKIIdelta in myocardial cells were measured by RT-PCR and Western Blot in normal rats and hypoxia rats 1 and the 3 weeks after exposure. The cardiac cells of the rats from the control group and the 3-week hypoxia group were cultured. Then the intracellular calcium activity was detected using laser confocal equipment. The effect of CaMKII on the calcium activity in myocardial cells was evaluated by the application of KN-62 (CaMKII specific inhibitor).

RESULTSThe expression of CaM, CaMKIIgamma and CaMKIIdelta mRNA in myocardial tissues increased in hypoxia rats compared with that in normal controls (P<0.01). The CaM and CaMKIIdelta mRNA expression was different between the 1-week and the 3-week hypoxia groups (P<0.01). The laser confocal demonstrated that the amplitude of calcium wave in hypoxic myocardial cells was not different from that in normal controls, but the duration of calcium wave in hypoxic myocardial cells was longer than that in normal controls (P<0.01). After KN-62 use, the amplitude of calcium wave decreased and the duration of calcium wave prolonged significantly.

CONCLUSIONSThe contents of CaM and CaMKII in myocardial cells increased under condition of chronic hypoxia as a compensation to keep calcium homeostasis in a certain time. With more prolonged hypoxia time, abnormal electric activities of heart occurred and the heart function may be impaired.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine ; analogs & derivatives ; pharmacology ; Animals ; Blotting, Western ; Calcium ; metabolism ; Calcium-Calmodulin-Dependent Protein Kinase Type 2 ; genetics ; Calmodulin ; genetics ; Chronic Disease ; Hypoxia ; metabolism ; Male ; Myocardium ; metabolism ; Norepinephrine ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo evaluate the effect of autologous bone marrow-derived stem cell (BMSCs) transplantation in the treatment of liver failure and decompensated hepatic cirrhosis.

METHODSBone marrow was harvested (65-95 ml) from 24 patients in the transplantation group. The BMSCs were isolated and infused into liver or spleen of patients via hepatic or splenic artery. At different time points after the transplantation, the patients' liver function and prothrombin time (PT) were evaluated, and the survival rate and symptoms of the patients were recorded.

RESULTSAll the serum biochemical indexes remained stable 2 weeks after the transplantation, and at 4 weeks after transplantation, albumin level increased significantly in comparison with the preoperative level (P<0.05). At 12 weeks, the albumin level further increased (P<0.01) along with Pre-ALB (P<0.01), while total bilirubin, tolal bile acid, PT and fibrinogen were all significantly lowered (P<0.05), and globulin, ALT, and AST remained unchanged (P>0.05). One week after the transplantation, improved appetite was observed in 22 cases (91.67%), and 21 cases (87.5%) showed better physical strength; at 2 weeks, hepatic face improved in 15 cases (62.5%), and spider telangiectasia was significantly reduced in one case; at 12 weeks, the survival rate of the patients was 62.5%, and 9 died or gave up treatment due to chronic liver failure complicated by spontaneous bacterial peritonitis, hepatorenal syndrome, or DIC. No complications associated with the transplantation occurred in these patients.

CONCLUSIONBMSC transplantation can significantly improve the liver function of patients with terminal liver disease with good safety and effectiveness.

Adolescent ; Adult ; Aged ; Bone Marrow Transplantation ; methods ; Female ; Humans ; Liver Cirrhosis ; physiopathology ; surgery ; Liver Failure ; physiopathology ; surgery ; Liver Function Tests ; Male ; Middle Aged ; Stem Cell Transplantation ; methods ; Transplantation, Autologous ; Treatment Outcome ; Young Adult

BACKGROUNDHereditary spastic paraplegia (HSP) is a group of inherited neurodegenerative disorders with the shared characteristics of slowly progressive spasticity and weakness of the lower limbs. Thirteen loci for autosomal dominant HSP have been mapped.

METHODSA Chinese family with HSP was found in the Shandong province and Inner Mongolia Autonomous Region of China and genomic DNA of all 19 family members was isolated. After exclusion of known autosomal dominant loci, a genome wide scan and linkage analysis were performed.

RESULTSThe known autosomal dominant loci of SPG3A, SPG4, SPG6, SPG8, SPG9, SPG10, SPG12, SPG13, SPG17, SPG19, SPG29, SPG31 and SPG33 were excluded by linkage analysis. The results of a genome wide scan demonstrated candidate linkage to a locus on chromosome 11p14.1-p11.2, over an 18.88 cM interval between markers D11S1324 and D11S1933. A maximal, two point LOD score of 2.36 for marker D11S935 at a recombination fraction (theta) of 0 and a multipoint LOD score of 2.36 for markers D11S1776, D11S1751, D11S1392, D11S4203, D11S935, D11S4083, and D11S4148 at theta=0, suggest linkage to this locus.

CONCLUSIONThe HSP neuropathy in this family may represent a novel genetic entity, which will facilitate discovery of this causative gene.

Adult ; Chromosome Mapping ; Chromosomes, Human, Pair 11 ; Female ; Humans ; Lod Score ; Male ; Spastic Paraplegia, Hereditary ; genetics

OBJECTIVETo screen and identify the proteins that interact with connexin 26 (CX26) and to analyze the expressions of these proteins in cochlea so as to explore the proteins that relate to the trafficking, assembly, localizing and gap junction functions of CX26.

METHODSThe whole coding region of GJB2 (CX26) gene was amplified from normal human genomic DNA by polymerase chain reaction (PCR) and then directionally subcloned into the vector pGBKT7 plasmid of the Match Maker Ga14 Two-Hybrid System 3 as a target to screen the interactive proteins of CX26 from the human fetal brain cDNA library by the yeast two hybrid technique. The false positive clones were discarded from the preys by repeated yeast two hybrid method between CX26 and everyone of the preys respectively. The DNAs of the insert of the identified positive clone were sequenced and BLAST analyzed against the GenBank. Lastly, the mRNA of the gene encoding the identified protein was analyzed in the murine inner ear by reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSThe insert of one positive clone contained 867 bp with the former 525 bp being coding region. The DNA sequence and the open reading frame of the insert were identical to the 525 bp before the stop codes (including the stop codes) and the 238 bp after the stop codes of RTN4 gene which encoded Nogo protein. And the 174 amino acid residues encoded by the insert were those of the C-terminal of Nogo protein: Nogo-A, Nogo-B and Nogo-C. RTN4 mRNA expressed in the murine inner ear was confirmed by RT-PCR method.

CONCLUSIONThe C-terminal of Nogo protein interacts with CX26. Nogo protein expresses in the inner ear and may take part in the trafficking of CX26 or CX26 gap junction function.

Animals ; Base Sequence ; Connexin 26 ; Connexins ; genetics ; metabolism ; Ear, Inner ; metabolism ; Gene Expression ; Humans ; Mice ; Molecular Sequence Data ; Myelin Proteins ; genetics ; metabolism ; Nogo Proteins ; Protein Binding ; RNA, Messenger ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Two-Hybrid System Techniques

OBJECTIVECanine model established for tracheal defect reconstruction, to investigate the outcome of tracheal reconstruction with combination of polypropylene and flap.

METHODSAbout 3.5 to 4 centimeter cervical trachea was resected and replaced with artificial trachea made from monofilament knitted polypropylene and surgical flap. Covered stent was implanted postoperatively. Survival period and quality of life were recorded, bronchofibroscopy, X-ray films and HE sections were performed.

RESULTSSix dogs survived well and another two died. The causes of death were respiratory failure in 1 and infection in another. Stenosis of anastomosis in 1 was recorded during survival period. The dogs started drinking and eating on the second postoperative day, no dyspnea was found. The animals were sacrificed at 2, 4, 8 weeks and 6 months after surgery. Soft tissue growth was found in polypropylene net 2 weeks after surgery and more at 4 weeks. The polypropylene net was covered completely with soft tissue at 8 weeks and 6 months postoperatively, the hardness and sustentation degree were enhanced following the growth and fibrosis of soft tissue. The squamous epithelium and columnar epithelium were observed healing well by HE staining method.

CONCLUSIONSOne-stage operative artificial trachea made from monofilament knitted polypropylene which has good histocompatibility and surgical flap is the closer artificial trachea to native trachea. It has a promising prospect in clinical use.

Animals ; Dogs ; Polypropylenes ; Prostheses and Implants ; Reconstructive Surgical Procedures ; instrumentation ; methods ; Skin Transplantation ; Surgical Flaps ; Trachea ; surgery

Objective To study the new endovascular treatment technique of intracranial wild-necked aneurysm, and preliminarily assess the safety and efficacy of hydrocoil assisted by stent and capsule. Methods We assessed the clinical value of therapeutic embolization of wild-necked aneurysm with hydrocoil assisted by stent and capsule by retrospectively studying the clinical data of 18 cases with wild-necked aneurysms, discussing the technical features, and follow-up visit of therapeutic effects by angiography. Results The therapeutic effects of 18 aneurysms were satisfactory, of which 6 were relapse-free as DSA demonstrated after half a year postoperatively. But long-term follow-up study was needed. Conclusion The endovaseular treatment of intracranial wild-necked aneurysm with hydrocoil assisted by stent and capsule is effective and safe.

Objective: To observe the effect of Yiqi Mingmu Pill on the expression of BaxmRNA protein and caspase-3 mRNA in retina of RCS rats. Methods: Twenty-four RCS rats were randomly divided into three groups: blank group, model group, Yiqi Mingmu Pill group. In the blank group, there were 8 RCS (rdy+/+, p+/+) rats, with 4 males and 4 females, and they received intragastric administration of normal saline. The model group consisted of 8 RCS (rdy-/, p-/-) rats, with 4 males and 4 females, and they received intragastric administration of normal saline. Yiqi Mingmu Pills groupconsisted of 8 RCS (rdy-/, p-/-) rats, with 4 males and 4 females, respectively. They received intragastric administrationof suspension of Yiqi Mingmu Pills. After intragastric administration for 30 days, the structure of retina layers wasobserved by HE staining. BaxmRNA and Caspase-3 mRNA in retinal tissues of each group were measured by RT-qPCRand Western Blot. Results: HE staining showed that the retina thickness of rats in Yiqi Mingmu Pill group wassignificantly thicker than that of the model group. The retinal pigment epithelial band was partially visible, and part of theouter nuclear layer photoreceptor sensory ciliary layer was visible. The number of photoreceptor cell nuclei was higherthan that of the model. There were many groups. The outer layer, outer core layer, and the visual cone layer were clearerand thicker than the model group. RT-qPCR showed that the relative expression of Bax mRNA and Caspase-3 mRNA inYiqi Mingmu Pills group was significantly lower than the model group, and the difference was statistically significant (P＜0.01), WB test showed that the relative expression of Bax protein and Caspase-3 protein in Yiqi Mingmu Pill group wassignificantly lower than that in the model group, and the difference was statistically significant (P＜0.01) . Conclusion: YiqiMingmu Pill has protective effect on the ultrastructure of RP retina. Yiqi Mingmu Pills can reduce the apoptosis of retinalphotoreceptor cells by inhibiting the expression of BaxmRNA and Caspase-3 mRNA on the retina and protect the visualcells.