Objective To detect the level of serum Klotho in chronic kidney disease (CKD) 3-5 stage patients and explore its relationship with the progress of renal function.Methods Twenty-nine patients with CKD 3-5 stage (CKD group) and 10 cases of age-matched non-CKD patients (control group) were enrolled in the study.The level of serum Klotho and fibroblast growth factor 23 (FGF23) were measured by enzyme-linked immunosorbent assay.Results The level of serum Klotho in CKD group was significantly lower than that in control group [(670.07 ± 146.22) ng/L vs.(1 125.50 ± 126.96) ng/L] (P ＜ 0.01).The level of serum FGF23 in CKD group was significantly higher than that in control group[(48.89 ± 44.28) ng/L vs.(10.48 ± 8.93) ng/L] (P ＜ 0.01).Pearson correlation analysis showed that the level of Klotho in CKD patients was positively correlated with estimated glomerular filtration rate (eGFR) and hemoglobin,and Klotho was negatively correlated with log urea nitrogen,log serum creatinine,log FGF23 and phosphate.Multiple regression analysis showed that log FGF23 and serum phosphorus were independent factors for serum Klotho level in patients with CKD.CKD patients were divided into rapid progression of renal function group (10 cases) and renal stability group (19 cases) according to annual rate of eGFR dechne.The patients in rapid progression of renal function group with lower serum Klotho [(580.27 ± 162.15) ng/L vs.(717.33 ± 115.22) ng/L],higher uric acid and more severe proteinuria [(448.00 ±65.21) mmol/L vs.(368.32 ±78.80) mmol/L,(1.99 ± 1.57) g/d vs.(0.60 ± 1.00) g/d] (P ＜0.05).Conclusions Serum Klotho level of CKD 3-5 stage patients decreases with the decline in renal function.Serum phosphate and FGF23 are the independent factors for serum Klotho.Serum Klotho may become the prediction index of renal function progress for patients with CKD.

Objective To evaluate the value of S100B gene on cardiovascular remodeling in rats with abdominal aorta coarctation.Method Sprague-Dawley rats(sanitary degree,male),weighring(220～240)g,were used in the study.They were provided by College of Medicine,Zhejiang University.The abdominal aortas of rats were isolated and constricted so as to establish models of heart failure;the abdominal aortas of another ten rats were isolated but not ligated(sham-operation group).After one week,the 20 rats were alive and randomly divided into 2 groups:operation group(n=10)and Carvidilol-operation group(Car group,n=10).Car Group was treatedwith 2 mg·kg-1per day Carvedilol and operation group was administered with the same doses of normal saline.After 4 weeks,a catheter was inserted through the right jngular artery into the left ventricle to record hemodynamic.Then all rats were euthanized,and the heart tissues were rapidly excised,rinsed with PBS.The left ventricles were then cut into two parts at the equator:the upper sections were fixation,the lower sections were stoted at-80℃,at 100 mg per tube.LV transverse section(4μm thick)was subjected to S100B immunehistoc- hemistry.RNA was iselated from tissues by Trlzol to determine levels of S100BmRNA and β-actinmRNA by RT- PCR.The data was expressed as(x±s);One-way analysis of variance was used for comparison among groups, and Student-Newman-Keulsa test for comparison between two groups.Statistical significance was set at P＜0.05. Results In Operation Group,there was a decrease in maximal rate of systolic and diastolic of left ventricular pressure(±dp/dtmax)[(1543.6±277.9)mmHg/s vs.(2640.4±481.3)mmHg/s and(-1352.5±202.3) mmHg/s vs.(-1873.2±412.3)mmHg/s];and an increase in left ventricular end diastolic pressure(LVEDP) [(24.8±5.2)mmHg vs.(2.1±0.7)mmHg]compared with sham-operation group(P＜0.01,P＜0.01 and P＜0.05).And in Car group,the level of LVEDP was just in the midst of the Operation Group and sham-opera- tion group,and had statistical significance(P＜0.01);while±dp/dtmac[(2372.3±92.6)mmHg/s and (-1786.4±62.6)mmHg/s]were much higher than those in operation group(P＜0.01).There were no any S100B positive cells and expression of S100B mRNA in sham-operation group;while there were much more S100B positive cells in operation group than those in Car group(P＜0.01).The expression of S100B mRNA in operation group was more pronounced than that in Car group.Changes in expression of S100BmRNA were positively correlat- ed with changes in LVEDP(r=0.847,P＜0.01);while changes in expression of S100BmRNA were negatively correlated with changes in±dp/dt max(r=-0.853 and-0.689,beth P＜0.01).Conclusions There was hish expression of S100B in myocytes from rats with experimental heart failure and negative correlation between the expression of S100B and heart function.It indicated that S100B could play a negative role in heart failure.

Objective To investigate the effect of theanine pretreatment on DNA repair function in neurons during brain ischemia-reperfusion (I/R) injury in rats.Methods One hundred and eight male Sprague-Dawley rats,weighing 290-310 g,aged 15 weeks,were randomly divided into 3 groups (n =36 each) using a random number table:sham operation group (S group),I/R group and theanine pretreatment group (T group).Global cerebral I/R was produced by 4-vessel occlusion method.Bilateral vertebral arteries were electrically cauterized,and bilateral common carotid arteries were clamped for 6 min.Theanine 1 g/kg was injected intravenously at 4 h before clamping bilateral common carotid arteries in T group,and the equal volume of normal saline was given in the other two groups.At 2,6,12,24,48 and 72 h of reperfusion,6 rats were selected in each group and sacrificed,the brains were removed,and the hippocampus was isolated for determination of the number of viable neurons in the hippocampal CA1 region (with a light microscope),apoptosis in neurons in the hippocampal CA1 region (by TUNEL),and expression of DNA repair protein X-ray repair cross-complementing group 1 (XRCC1) and Ku70 (by immunohistochemistry).The apoptotic index was calculated.Results Compared with S group,the number of viable neurons was significantly decreased,and the apoptotic index was significantly increased at 6,12,24,48 and 72 h of reperfusion,and the expression of XRCC1 and Ku70 was significantly down-regulated at 2,6,12,24,48 and 72 h of reperfusion in I/R group (P＜0.01).Compared with I/R group,the number of viable neurons was significantly increased at 12,24,48 and 72 h of reperfusion,the apoptotic index was significantly decreased at 6,12,24,48 and 72 h of reperfusion,and the expression of XRCC1 and Ku70 was significantly up-regulated at 2,6,12,24,48 and 72 h of reperfusion in T group (P ＜ 0.01).Conclusion The mechanism by which theanine pretreatment attenuates brain I/R injury is related to enhancement of DNA repair function and reduction of neuronal apoptosis in rats.

Objective To investigate the relations between serum adiponectin and coronary artery calcification score (CACS) and to find the risk factors for coronary artery calcification in maintenance hemodialysis(MHD) patients.Methods Twenty-seven MHD patients(MHD group) and 13 healthy persons (control group) were enrolled in this study.The serum adiponectin was measured by enzyme-linked immunosorbent assay method.CACS was calculated by multi-row spiral CT.The circulating parameters such as hemoglobin (Hb),calcium,phosphate,calcium-phosphate product,intact parathyroid hormone (iPTH),total cholesterol (TC),low density lipoprotein cholesterol (LDL-C),albumin (Alb),high sensitive C-reactive protein (hs-CRP),and so on were detected.Results The level of serum adiponectin in MHD group [(15.00 ± 7.47) mg/L] was significantly higher than that in control group [(2.07 ± 0.83) mg/L],and there was significant difference (P＜ 0.01).Coronary artery calcification(CACS ＞ 0 score) was observed in 88.9％ (24/27) in MHD group and 10/13 in control group.The mean CACS in MHD group was significantly higher than that in control group [655 (0-3 570) scores vs.126 (0-731)scores],and there was significant difference (P ＜ 0.05).Eleven MHD patients and 1 healthy person had severe coronary artery calcifications,(CACS ≥ 400 scores).There was significan t difference in dialysis duration,diastolic pressure,phosphate calcium-phosphate product and adiponectin (P ＜ 0.05 or ＜ 0.01).Spearman analysis showed that CACS of MHD patients was positively correlated with dialysis duration,phosphate,calcium-phosphate product,serum creatinine and adiponectin (P ＜ 0.01 or ＜ 0.05).Only calcium-phosphate product remained as independent predictor of CACS in multivariate analysis (P ＜0.01).Conclusion Coronary artery calcification is common in MHD patients and which is correlated with dialysis duration,serum phosphate,calcium-phosphate product,serum creatinine and adiponectin.

Objective To observe the in vitro inhibitory effects of brucea javanica oil on human washed platelet and explore the possible mechanism.Methods Human washed platelets were mixed withdifferent concentration of brucea javanica oil which were divided into four groups[untreated control group,negative control group,9.0% of brucea javanica oil group,and 22.5%of brueea javanica oil group].The maximal ratio of platelet aggregation induced by adenosine liphosphate(ADP),arachidonic acid(AA),collagen,and thrombin,respectively,was measured with platelet aggregation analyzer.The expressions of fibrinogen receptor(FIB-R)and P-selectin(CD_(62p))on external membrane of activated platelet were determinated with flow cytometry.The F-actin of cytoskeletal structure in activated platelet was detected by SDS-PAGE.Results At 9.0% of brucea javanica oil,the maximal ratio of platelet aggregation[(57.7±4.0)%,(62.2±3.9)%,(66.9±5.0)%and(71.8±5.1)%]induced by ADP,AA,collagen,and thrombin,was significantly lower than that[(75.3±4.1)%,(79.3±4.8)%,(80.6±5.4)%,(84.1±6.2)%]at negative control(0% of brucea javanica oil)(P<0.01),but makedly higher than that[(39.2±3.5)%,(45.8±3.4)%,(51.2±3.9)%and(56.7±4.8%)]at 22.5%,respectively(P<0.01).The inhibitory rate of platelet aggregation(47.9%,42.2%,36.5%and 32.6%)at 22.5%of brucea javanica oil was notably higher than that(23.4%,21.6%,17.0%and 14.6%)at 9.0%,respectively(P<0.001).There was a negative correlation between brucea javanica oil concentration and the aggregation ratio of platelet stimulated by the four agonists,respectively(r=-0.952,-0.961,-0.970,-0.975,P<0.001).At 9.0% of brucea javanica oil,the expression levels of FIB-R[(64.7±4.0)%]and CD_(62p) [(3.91±0.21)%] of platelet activated by ADP were significantly lower than that[(85.5±4.6)%and (5.05±0.27)%]at negative control,but remarkably higher than that[(36.2±3.9)%and(2.34±0.15)%]at 22.5%,respectively(P<0.01).There was a much higher inhibitory rate of platelet aggregation(57.7%)at 22.5%than that(24.3%)at 9.0%(P<0.01).The ratios(1.68±0.10 and 1.77±0.12)of F-actin photodensity at 22.5%and 9.0%to that in blank control were significantly lower than that(2.22±0.15)at negative control(P<0.01)but there was no statistical difference between the ratios in the group of 9.0%and 22.5%brucea javanica(P>0.05).Conclusions brucea javanica oil has special inhibitory effect on activated platelet and thrombosis in a dose-dependent manner.The mechanism is to inhibit the expression of fibrinogen receptor on external membrane of activated platelet,which is also related to the inhibition of F-actin and secretion of platelet.

Objective To analyze the cerebral autoregulation capability in patients with chronic insomnia disorder (CID).Methods Sixty CID patients (54 with generalized anxiety disorder) and 40 healthy controls were enrolled in this study.Polysomnography was done in all the participants.Noninvasive continuous cerebral blood flow velocity of bilateral middle artery and arterial blood pressure were recorded simultaneously using transcranial Doppler and a servo-controlled plethysmograph.Transfer function analysis was used to derive the autoregulatory parameters, including phase difference and coherence function.Results The phase difference values of CID patients with generalized anxiety disorder were significantly lower than that of the healthy controls ((46.89±15.39)°vs (56.00±12.05)°, t=3.439, P=0.001).In the correlation analysis, we further found that there was no correlation among phase difference values and the score of Hospital Anxiety and Depression scale.Conclusions The dynamic cerebral autoregulation was compromised in CID patients with generalized anxiety disorder regardless of the degrees of anxiety and depression.Dynamic cerebral autoregulation may be a potential therapeutic target in improving neurological symptoms in patients with CID.

Objective To observe the effect of the 8-word bandage on the walking ability of stroke patients with knee hyperextension. Methods Fifty patients with stroke combined with knee hyperextension were randomly divided into the observation group and the control group with 25 cases in each group. The control group was treated with conventional methods, including Bobath technology, Brunnstrom therapy and motor relearning primarily rehabilitation training. In addition to the conventional methods, patients in the observation group used 8-word bandage to fix knee joint in walking training. Before and after 8 weeks of treatment, Holden walking function classification, 10-meter maximum walking speed and improved Barthel index were adopted to evaluate the walking ability, maximum walking speed and the life ability of the patients. Results There were no significant differences in the scores before treatment between the two groups. The Holden walking function classification, the 10-meter maximum walking speed and the Barthel index scores were significantly improved after 8-week treatment in both two groups (P<0.05), and patient conditions were more significantly improved in the observation group than those of the control group (P<0.05). Conclusion Using 8-word bandage to fix knee joint can significantly improve knee hyperextension in patients with stroke, so as to improve the walking ability and activities of daily living.

Objective This study is aimed at determining whether anti-von Willebrand factor (VWF) autoantibodies are present in the plasma of idiopathic thrombotic thrombocytopenic purpura (TTP) patients with normal ADAMTS13 activity and undetectable anti-ADAMTS13 antibodies,and at examining whether murine monoclonal antibodies (mAbs) against human VWF decrease the susceptibility of VWF to ADAMTS13 in vitro.Methods Anti-VWF autoantibodies and ultralarge VWF (UL-VWF) multimers were measured in plasma samples of 53 adult patients with idiopathic TTP by enzyme-linked immunosorbent assay and sodium dodecylsulphate-agarose gel electrophoresis,respectively.Moreover,the effects of eight murine mAbs to different human VWF domains on VWF cleavage by ADAMTS13 were evaluated under fluid shear stress and static/denaturing conditions,respectively.Results Anti-VWF antibodies and UL-VWF multimers were detected in two TTP patients with normal ADAMTS13 activity and undetectable anti-ADAMTS13antibodies.The SZ34,an anti-VWF mAb,inhibited VWF proteolysis mediated by ADAMTS13 under flow,but not static conditions.Conclusion Anti-VWF antibody may be one of the causes of idiopathic TTP with normal ADAMTS13 activity and undetectable anti-ADAMTS13 antibodies.

Objective:To analyze the molecular characteristics of Echovirus 11 (Echo11) strains isolated in Xiangyang, Hubei Province from 2016 to 2017 based on the sequences of VP1 gene.Methods:Rectal and throat swab specimens were collected from children with hand, foot and mouth disease (HFMD) in Xiangyang from 2016 to 2017. Echo11 strains were detected by real-time reverse transcriptase PCR (RT-PCR) and isolated after cultured in human rhabdosarcoma (RD) cells. The VP1 regions of Echo11 strains isolated from RD cells and the whole genomes of three representative Echo11 strains were amplified by conventional RT-PCR and the sequences were analyzed. DNAStar7.0 (MegAlign) and MEGA6.0 (Data) were used to analyze the homology and mutation sites in nucleotide and amino acid sequences. Neighbor-joining method was used to construct phylogenetic trees. Recombination analysis was performed with SimPlot software (BootScanning).Results:A total of 11 Echo11 strains were isolated from 3 494 HFMD cases, accounting for 0.31%. They were highly homologous in the VP1 gene. These strains shared 98.4%-100.0% homology in nucleotide sequences and 98.3%-100.0% homology in amino acid sequences. The homology between the 11 Echo11 strains and the prototype strain (Echo11/Gregory, X80059) was 73.9%-74.8% in nucleotide sequences and 87.7%-88.7% in amino acid sequences. All of the Echo11 strains circulating in Xiangyang were classified into lineage D, having a similarity to the strains circulating in some regions of mainland China since 2013. In multiple regions of the genome, the Echo11 strains isolated in Xiangyang were highly similar to the Henan Echo1 strains in 2010 and the Hubei Echo6 strains in 2015, suggesting there was recombination within the genome of Echo11 strains in Xiangyang.Conclusions:The Echo11 strains circulating in Xiangyang from 2016 to 2017 belonged to lineage D and were recombinant strains.

Objective To evaluate the changes in expression of NF-κB, IL-6 and TNF-α in spinal cord in a rat model of bone cancer. Methods Seventy-two female SD rats weighing 150-180 g were randomly divided into 3 groups (n = 24 each): control group (group C);sham operation group (group S) and bone cancer pain group (group BP). Bone cancer was induced by intra-tibial inoculation of 1 × 105 Walker 256 breast cancer cells. Paw withdrawal threshold to mechanical stimulation was measured with yon Frey filaments. The expression of NF-κB p65, IL-6 and TNF-α mRNA in the spinal cord was determined by RT-PCR and the expression of NF-κB p65 by immuno-histochemistry and NF-κB p65 positive cell count was determined. Results The paw withdrawal threshold was significantly lower and the expression of NF-κB p65, NF-κB p65 mRNA, IL-6 mRNA, TNF-α mRNA and NF-κB p65 positive cell count in the spinal cord were significantly higher in group BP than in group C and S ( P ＜0.05 or 0.01 ). Conclusion Intra-tibial inoculation of Walker 256 breast cancer cells activates NF-κB in the spinal cord, leading to the increased release of IL-6 and TNF-α and mechanical hyperalgesia.