Aim To purify L-amino acid oxidase(LAAO) from the venom of Naja atra and study its effect on endothelial cells.Methods The NAV-LAAO was purified by ion-exchange chromatography and affinity chromatography.The MTT assay and Western blot were used to detect the viability and apoptosis of HUVEC.The tubule-forming was used to study the angiogenesis of cells.Results The NAV-LAAO was purified successfully from the venom of Naja atra.The molecular weight of NAV-LAAO was determined to be 58 ku by SDS-PAGE.NAV-LAAO effectively inhibited the growth and tubule-forming of HUVEC,and the 50% inhibitory concentration(IC50) was 21.42 mg?L-1.Compared with control,the levels of caspase-3 and caspase-8 increased in HUVEC treated with NAV-LAAO.Conclusions The NAV-LAAO is purified successfully from Naja atra venom by ion-exchange chromatography and affinity chromatography.The NAV-LAAO inhibits the growth and tubule-forming capacity of HUVEC in a dose-dependent manner.

Objective To investigate the application value of urine microalbumin (mALB) ,retinol binding protein(RBP) and cys-tatin C(CysC) and their combined detection in early diagnosis of type 2 diabetic nephropathy(DN) .Methods Ninety-two inpatients with DN (DN group) and 90 people undergoing the physical examination(control group) in our hospital from June 2014 to Decem-ber 2015 were collected .Urine mALB ,RBP and CysC were detected in all subjects and detection results were analyzed statistically . Results The levels of urine mALB ,RBP and CysC in the DN group were significantly higher than those in the control group ,the differences all had statistical significance (P< 0 .05) .Among 3 indicators ,the positive rate of urine mALB for detecting DN was highest (94 .57% ) ,while which of 3-index combined detection was 97 .83% ,and significantly higher than that of single detection , the difference was statistically significant(P<0 .05) .The sensitivity ,specificity ,positive predictive value ,negative predictive value and Youden index of 3-index combined detection were all higher than those of single index .The ROC curve showed that AUC of u-rine mALB for diagnosing DN was 0 .732 ,the diagnostic cut-off value was 43 .58 mg/L ,AUC of urine RBP was 0 .685 ,the diagnos-tic cut-off value was 1 .47 mg/mL ,AUC of urine CysC was 0 .701 ,the diagnostic cut-off value was 1 .42 mg/L ,while AUC of com-bined detection was 0 .928 .Conclusion Urine mALB ,RBP and CysC are better indexes reflecting renal injury .Their combined de-tection will increase the positive rate ,sensitivity and specificity for diagnosing DN .So monitoring the levels of urine mALB ,RBP and CysC has an important significance to diagnosing the occurrence and development of DN early renal injury and prevention ,treat-ment and delaying progress of DN .

Objective To develop a candidate reference method for the determination of serum creatinine and to evaluate the ac-curacy of conventional detection systems though method comparison to achieve traceability .Methods The candidate reference method was established according to the sarcosine oxidase and the accuracy and reliability of the method was verified through par-ticipation in international reference laboratories EQA activities (IFCC-RELA) .20 fresh single human serum samples with different concentration and calibrator were simultaneously measured by using conventional detection system and candidate reference method . Results The calibration curve for serum creatinine was linear in the concentration range from 50-2 000 μmol/L with a correlation coefficient of 0 .999 9 under the optimum experimental conditions (the linear equation was Y=0 .000 884 2X-0 .000 325 3) and the imprecision was less than 1 .0% .The proposed method has been applied to the determination of RELA samples with satisfactory re-sults .The measured results with conventional detection systems were consistent with candidate reference method ,and the slope of the regression equation was 1 .005 6 .Conclusion The candidate reference method of serum creatinine is successfully established and which can be used for traceability and standardization .It may provide an effective way for conventional detection system traceable to the reference method or reference material .

Objective To observe the effect of pigplacenta ins tead of Ziheche (P lacenta Hominis) for the treatment of senile dementia. Methods Seventy Kunming mice were divided into blank group, model group, pos itive control group, Zih eche low and high dosage groups, and pigplacenta low and high dosage grou ps, wi th 10 in each. The senile dementia models were established with the D-Galactose subcutaneous injection. The blank group was not administered any medicines. The model group was prescribed normal saline instead of the tested medicine; the pos itive control group was given Naofukang by gavage; the Zih eche low and high dosage groups were given 2g/kg and 4g/kg Ziheche respectivel y by gavage; while the pigplacenta low and high dosage groups were treated wi th p igplacenta 4g/kg and 8g/kg respectively by gavage. After treatment for 6 week s, the behavior experimental dark-avoiding test and step-down test were applied to test the effect of the medicines on the learning memory of mice, and acetylchol inesterase and monoamine levels in brain tissues. Results There was no s ignificant difference between the effect of pigplacenta and Ziheche in resi stin g senile dementia. In the latency of dark-avoiding test, the effect of high d osa ge of pigplacenta was significantly better than that of Ziheche (P

Objective To explore the effect of artificial and computer-assisted cognitive training on visuospatial impairment. Methods63 patients with visuospatial impairment were randomly assigned to artificial group (n=21), computer-assisted training group (n=21) and integrationgroup (n=21). The artificial group received conventional cognitive training while the computer-assisted training group used computerto assist cognitive training, and the integration group combined computer with conventional cognitive training. All groups were assessedwith Loewenstein Occupational Therapy Cognitive Assessment (LOTCA). Results All groups improved significantly after training(P<0.001). There were no significant difference between 3 groups in visual perception, spatial perception, and motor praxis after training.Compared to the computer-assisted training group, the integration group and the artificial group improved more significantly in visuomotororganization after training. Conclusion Artificial cognitive training and computer-assisted training can effectively improve the cognitivefunction of patients with visuospatial impairment.

Objective To observe the effect of oral motor exercise on the feeding behaviour of autistic children. Methods 59 children with autism were assigned to control group (n=29) and exercise group (n=30). The control group received diet intervention, while the exercise group received oral motor exercises in addition. Both groups were assessed with questionnaire related to children's diet before and after treatment. Results Bad feeding behaviors such as little eating, overlength eating, picky eating, partial eating, refuse to eat significantly improved after treatment in the exercise group (P<0.05), and improved more than in the control group (P<0.05). Stereotyped feeding behavior improved after treatment in both groups (P<0.05) and there was no difference between 2 groups (P>0.05). Conclusion Oral motor exercises may improve the feeding behaviour of autistic children.

Objective To investigate the effects of surface anesthesia on assisted balloon dilatation when treating dysphagia caused by radiotherapy for nasopharyngeal carcinoma.Methods Fifty-four patients with dysphagia after radiotherapy were divided randomly into an anesthesia group and a non-anesthesia group.The anesthesia group received anesthetics before treatment while the non-anesthesia group did not.All of the patients were treated with low-frequency electrical stimulation and assisted balloon dilatation for 3 weeks.They were then assessed using videofluoroscopy and self-reports of difficulty in swallowing before and after the treatment.Results After the treatment, significant improvement was observed in pharyngeal delay time, in cricopharyngeal opening, and in laryngeal elevation and forwardness.There was also a significant decrease in self-reported swallowing difficulty and failed swallows in both groups compared with before the treatment.The improvements in the non-anesthesia group were significantly greater than in the anesthesia group.After the treatment, the average aspiration rate of the anesthesia group was significantly higher than before treatment and higher than that of the non-anesthesia group.The improvement in oral intake of the non-anesthesia group was significantly better than that of the anesthesia group.Conclusion Balloon dilatation and low-frequency electrical stimulation have a synergistic effect and can improve patients' swallowing after radiation-induced cranial nerve damage, thus promoting survival.Assisted balloon dilatation without anesthesia has a better effect than when surface anesthesia is used.

Objective To study the effects of early application of integrated traditional Chinese medicine (TCM)and western medicine and massage on the incidence and severity of pneumonia and prognosis in the therapeutic course of infants with cytomegalovirus(CMV)infection infantile cholestatic hepatopathy(ICH)accompanied by gastric volvulus(GV). Methods A prospective randomized controlled trial was conducted. 120 infants 1-6 months old with CMV infection ICH and complicated with GV inpatients were divided into treatment group and control group (each,60 cases). The TCM Lidanheji(consisting of artemisiae capillaris 30 g,weeping forsythia 30 g,Chinese goldthread 5 g,prepared rhubarb 5 g,unpeeled root of herbaceous peony 30 g,cassia 5 g,fruit of citron or trifoliate orange 10 g,large-headed atractylodes 10 g,fruit of Chinese magnoliavine 10 g,pangolin scale 3 g and licorice root 5 g)for oral administration or enema and ganciclovir intravenous injection were given to both groups. In the control group,based on the above treatment,postural and diet therapy,prokinetics(domperidone)were given in cases with GV. The massage treatment was only applied for treatment of GV in treatment group. The pneumonia in both groups was treated in accord to the conventional diagnosis and treatment for infantile pneumonia,bronchiolitis,severe pneumonia and the guide and management of infantile community acquired pneumonia. The total number of cases with pneumonia and its severity occurring before and after treatment in the course of the disease,the clinical therapeutic effects of ICH, GV and infantile pneumonia,the clinical physical signs,biochemical indexes and the time of hospitalization were observed and compared between the two groups. Results Compared with control group,after treatment the curative rate and total effective rate of ICH,GV,infant pneumonia in treatment group were significantly higher〔the curative rate of ICH:83.05%(49/59)vs. 71.93%(41/57),total effective rate:96.61%(57/59)vs. 91.23%(52/57);the curative rate of GV:72.88%(43/59)vs. 51.79%(29/56),total effective rate:96.61%(57/59)vs. 78.57%(44/56);the curative rate of infant pneumonia:81.08%(30/37)vs. 67.44%(29/43),total effective rate:100%(37/37)vs. 100%(43/43),P<0.05 or P<0.01〕. In the treatment group,the total number of cases with pneumonia occurring in the therapeutic course and the incidence of severe pneumonia were significantly less than those in the control group〔the occurrence of pneumonia:12 cases vs. 21 cases,the incidence of severe pneumonia:16.67%(2/12) vs. 42.86%(9/21),P<0.05 or P<0.01〕. The enlargement of liver and spleen,the level of total bilirubin(TBil), direct bilirubin(DBil),total bile acids(TBA)and alanine aminotransferase(ALT)in two groups after treatment were significantly lower than those before treatment,the changes being more marked in treatment group(P<0.05 or P<0.01). The time of stay in hospital in treatment group was remarkably shortened(days:21.32±3.26 vs. 27.38±6.09, P<0.05). Conclusion Early interference with combined TCM and western medicine and massage for treatment of infants with CMV infection ICH accompanied by GV can significantly decrease the incidence of pneumonia and its severity occurring in the disease course,therefore this therapeutic method is beneficial to the treatment and prognosis of infants with CMV infection ICH.

Objective To explore the clinical manifestations and the characteristics of neonatal intrahepatic cholestasis caused by Citrin deficiency(NICCD)in Hubei province. Methods The biochemical indicators including liver function,blood lipid,lactic acid,blood ammonia,total bile acid,alpha feto protein,coagulogram,blood amino spec-trum,acylcrnitine spectrum,urine organic acid and SLC25A13 gene analysis of 20 cases with NICCD,who came from Wuhan Children's Hospital,during September 2010 to January 2013,were collected before treatment,then followed up for 1 year. Results Laboratory results of NICCD patients showed high blood bilirubin,elevated liver enzymes and bile acid,hyperlipidemia,high alpha feto protein,high lactic acidosis,high ammonia,hypoalbuminemia,hypoglycemia,disor-der of blood coagulation mechanism,variety of amino acids increase,mainly citrulline rose. Mainly long - chain acyl carnitine increased among acyl of carnitine. Abnormal increase of urine 4 - hydroxy benzene acetic acid,4 - hydroxy benzene lactic acid and 4 - hydroxy benzene pyruvic acid. Six mutations were detected in SLC25A13 gene analysis,and L477R,G639S of them were novel mutations,851del4,1638ins23,IVS6 + 5G ﹥ A were hot mutation. All the patients were eased in jaundice before they were 1 year old. Conclusions The early clinical criterion of the patients is disor-der. Hyperlipidemia has been detected in the early course of the disease,and L477R,G639S are the novel mutations.

Objective To observe the clinical efficacy of Linda mixture self made by combination of disease with syndrome pattern for treatment of neonatal intrahepatic cholestasis disease caused by Citrin defects (NICCD) and to provide an experience in early diagnosis and treatment of NICCD.Methods The clinical features of 20 patients with NICCD was confirmed by NICCD common genetic loci detection during hospitalization admitted to the Department of Integrated Traditional and Western Medicine in Wuhan Children Hospital from June 2012 to June 2015 were retrospectively analysed.The clinical efficacy of Chinese medicine Linda mixture for treatment of NICCD,before and after treatment,the index changes of liver function and coagulation function were observed and Color Doppler ultrasound was used to detect the situation of hepatosplenomegaly in the patients.Results There were 20 cases all their pathological jaundice occurring within 3 months old,after treatment,15 cases of them obtained clinical recovery and 5 cases were getting better,the average length of stay in hospital was (14.67 ± 1.56) days.After the treatment,the levels of serum total bilirubin (TBil),direct bilirubin (DBil),alanine aminotransferase (ALT),total bile acid (TBA) were markedly lower compared with those before treatment [TBil (μmol/L):64.0 ± 39.5 vs.173.5 ± 54.1,DBil (μmol/L):37.7±24.8 vs.80.9±46.4,ALT (U/L):42.1±25.8 vs.55.0±32.2,TBA (μnol/L):67.5±20.3 vs.195.0±61.3,all P ＜ 0.05],albumin (Alb) and total albumin (GLB) were significantly higher compared with those before treatment [Alb (U/L):37.9 ± 4.7 vs.33.1 ± 4.7,GLB (g/L):17.3 ± 4.0 vs.14.6 ± 2.8,both P ＜ 0.05],activated partial thromboplastin time (AP'TT) of coagulation function was significantly reduced (s:38.2± 8.3 vs.63.1±24.0,P ＜ 0.05).Color Doppler ultrasound examination showed that hepatosplenomegaly was shrunk after treatment [hepatomegaly (cm):2.12 ± 0.70 vs.3.04 ± 0.25,splenomegaly (cm):0.25 ± 0.03 vs.0.58 ± 0.32,both P ＜ 0.05].After 1 month of follow up,the infants' body weights were almost normal,the average increase in weight being (1.01±0.32) kg,the color and frequency of stool were distinctly better compared with those before treatment,the stool color in 15 cases was golden,pale yellow in 5 cases and no one,pottery clay in color.The stool frequencies of 18 cases were kept under 5 times,and fatty diarrhea situation was ameliorated.Conclusion Using Chinese medicine for treatment of NICCD can obtain significant clinical effects.