Time-resolved fluorescence immunochromatographic assay for the detection of mycoplasma pneumoniae IgM and IgG / 中华核医学与分子影像杂志

Objective:To establish time-resolved fluorescence immunochromatographic assay (TFICA) for rapid and quantitative detection of mycoplasma pneumoniae (MP) immunoglobulin (Ig)M and IgG.Methods:Based on capillary effect and europium nanospheres, rapid TFICA for MP-IgM and IgG detections were developed with the optimized parameters (coupling rates of antigens or antibodies to microspheres, dilution of labeled nanospheres, fixture concentrations on test line and serum dilutions). The methodological performances were estimated such as sensitivity, specificity, stability. By testing 55 healthy control samples, the reference values of TFICA were obtained. The reliability was evaluated by Kappa test from detecting sera of 88 cases (33 patients and 55 healthy controls) using TFICA and commercial kits by chemiluminescence immunoassays (CLA). Results:After screening the assay conditions, the mass ratios of mouse anti-human IgG and MP antigen with nanospheres were 1∶20 and 1∶100 respectively; the work dilutions of nanobeads conjugated with anti-human IgG and MP antigen were 1∶200 and 1∶100 respectively; the spraying concentrations of MP antigen and goat anti-human IgM were 0.5 and 1.0 g/L on the test line respectively, and the working dilutions of serum sample were both 1∶300. In the MP-IgM and IgG detections, the linear working ranges were (0.78-70.00)×10 3 relative unit (RU)/L and (0.17-200.00)×10 3 RU/L, while the sensitivities of the assays were 0.78×10 3 and 0.17×10 3 RU/L, respectively. No cross reactions were found with antithyroid peroxidase antibody, anticardiolipin antibody or thyroglobulin antibody. In these MP-IgM and IgG assays, the relative standard deviations were 3.7%-14.8% and 2.9%-14.0%, the average reduction rates of fluorescence were 13.7% and 14.2% respectively after incubation at 37 ℃ for 5 d. The reference values of MP-IgM and IgG were 3.33×10 3 and 2.61×10 3 RU/L, while the Kappa values between TFICA and CLA were 0.79 and 0.76, respectively. Conclusion:TFICA is a simple, sensitive, specific and quantitative method for detecting MP-IgM and IgG antibodies, and may show great promise for future clinical use.

Analysis on Muscle Mechanical Properties of Stroke Patients’ Hand Grasping Training Based on AnyBody / 医用生物力学

Objective To establish the biomechanical model of skeletal muscle during hand grasping for reverse dynamics simulation, so as to obtain the maximum muscle force of each muscle involved in the process of hand grasping under different forces. Methods CT scanning was performed on a volunteer’s hand, and CT data of his hand were imported into Mimics software for 3D reconstruction, so as to obtain the bone models of each segment. After adjusting the model coordinates by Geomagic Studio, the model was imported in AnyBody software for establishing the kinematics model of the hand skeleton. The related muscles involved in the flexion of each finger were added, to establish the skeletal muscle model of the hand. The model was then used to simulate the reverse dynamics of hand grasping. ResultsThe maximum muscle force of each muscle in the whole process of finger movement was obtained after the 5-30 N external force was applied to each distal phalanx. With the increase of force, the maximum muscle force of each muscle showed a linear trend. For example, the maximum muscle force of flexor pollicis longus increased from 18.49 N to 110.93 N; when the external force was 5 N, the maximum muscle force of flexor pollicis brevis, flexor pollicis longus, adductor pollicis and flexor digiti minimi brevis during hand grasping was 7.70, 18.49, 9.49, 8.39 N, respectively. The muscle force of superficial and deep flexors was greater than that of other muscles in the process of finger movement, which played a major role in grasping the hand. Conclusions The maximum muscle force of the muscles involved in hand grasping under different resistance, and the relationship between muscle force of main muscles and joint angles, can provide guidance and references for the evaluation of hand rehabilitation effect of stroke patients, as well as certain theoretical basis for the manufacture of rehabilitation equipment.

Experimental Study of Stress Effects on the Degradation Rate in vitro of Novel Magnesium Alloy Bone Screws / 医用生物力学

Objective To study the effect of stress on the degradation rate in vitro of novel magnesium alloy bone screw. Methods A three-dimensional （3D） model of the tibia fracture was established using the reverse engineering method. Then, based on the FE model, the in vitro degradation experimental device for bone screws was designed. The stress distribution of the screw by finite element calculation was used as the in vitro experimental load, which effectively improved the accuracy and efficiency of the experiment. The experimental samples were divided into four groups. Group A was treated as control group without force application, while Groups B, C and D were subjected to 150, 250 and 350 N axial forces. The influence of different mechanical environment on the degradation rate in vitro of bone screws was investigated. Finally, combining the stress distributions with the degradation experiment results in vitro, the curve between the stress and the degradation rate in vitro of novel magnesium alloy bone screws was obtained. Results Degradation experiments in vitro showed that Group A had the lowest weight loss and hydrogen production, and the average degradation rate was (0.315±0.005) mm/a. While in the stress groups, the weight loss and hydrogen production increased gradually with the axial force increasing. The average degradation rates of Groups B, C and D were (0.379±0.006), (0.469±0.007) and (0.547±0.009) mm/a, respectively. Conclusions When the novel magnesium alloy bone screw was degraded in mechanical environment, the greater stress on the screw would cause the faster degradation rate in vitro. The obtained relationship between the maximum stress and the average degradation rate in vitro of the novel megnesium alloy bone screw provided data support and theoretical guidance for material selection, design and clinical application of magnesium alloy bone screws.

Surgical outcome evaluation and prediction analysis of laparoscopic left sided hepatectomy for hepatolithiasis / 中华外科杂志

Objective@#To explore the risk factors of long-term treatment outcomes and establish predicting model for laparoscopic left hepatectomy in hepatolithiasis.@*Methods@#Clinical data of 108 patients with hepatolithiasis who underwent laparoscopic left sided hepatectomy and with complete follow-up data were retrospectively collected from June 2011 to June 2016 at the Second Affiliated Hospital of Nanchang University. Twenty-six males and 82 females were enrolled. The age was (52.4±11.7) years (range:20-80 years) , and the median follow-up time was 36 months (range: 24-83 months) . Patients were randomly divided into training group (79 cases) and validation group (29 cases) with a ratio of about 3∶1. Twenty-five preoperative and intraoperative clinical factors were selected for potential factors that might affect long-term outcomes, and quality of life was used as an surrogate evaluation index. Univariate analysis and multivariate logistic regression analysis were used to investigate the potential risk factors, and to construct and validate the predictive nomogram for surgical outcomes.@*Results@#Among 108 patients, 10 patients (9.3%) had residual stones, 8 patients (7.4%) had recurrent stones, 12 patients (11.1%) had recurrent cholangitis and 3 patients (2.8%) died. Univariate analysis showed that history of hepatobiliary surgery, gender, activation of partial thromboplastin time, alkaline phosphatase, use of choledochoscopy, postoperative stone residual, serum creatinine, postoperative biliary drainage and operation time were risk factors that may affect long-term outcomes (all P<0.15) . Multivariate analysis showed that the history of previous hepatobiliary surgery (OR=2.305, 95% CI: 0.383-4.227, P=0.019) , postoperative biliary drainage (OR=2.043, 95% CI: 0.182-4.209, P=0.048) , operation time ≥262.5 minutes (OR=1.971, 95% CI: 0.154-4.023, P=0.045) were independent risk factor affecting long-term outcomes. Based on the above factors, the predictive nomogram model was constructed. Internal and external validations showed good discrimination (area under the curve of receiver operating curve>0.7) and calibration (Hosmer-Lemeshow test: P>0.05) performance, which indicated that the prediction effect was favorable.@*Conclusions@#History of previous hepatobiliary surgery, postoperative biliary drainage and operation time ≥262.5 minutes are independent risk factors for long-term outcome. The predictive nomogram model based on risk factors relates to surgical outcomes presented good clinical predictive effects, which might contribute to the prediction of the long-term outcomes of laparoscopic left sided hepatectomy for hepatolithiasis.

Repair of bile duct defection with decellularized vascular matrix scaffold in a porcine model / 中华普通外科杂志

Objective To explore the feasibility of repairing porcine bile duct defect with decellularized rabbit abdominal aorta matrix scaffold.Methods Sodium dodecyl sulfate and Sodium deoxycholate were used to remove the cells in the blood vessel,and the residual DNA and RNA fragments were removed by nuclease.The prepared scaffold was implanted to repair defect of bile duct in swine,which were sacrificed after 45 days of surgery for histological evaluation.Results HE,Masson and elastic fiber staining showed that the composition and structure of the scaffold maintained their native features after dcellularization treatment.DNA content in acelllular scaffold (0.12 ± 0.01) μg/mg dry weight) was significantly decreased as compared with the native ones (2.31 ± 0.03) μg/mg dry weight,P ＜ 0.05).Collagen content was increased from (152 ±22) μg/mg dry weight in intact aorta to (177 ±21) μg/mg dry weight.Adipose derived mesenchymal stem cells with typical morphology survived well in the decellularized vascular matrix.It was observed that seeded ASCs penetrated into the inner wall of the scaffold.After transplantation,there was no leakage in the anastomosis and collapse of acellular blood vessel matrix.After 45 days of transplantation,repaired bile duct was harvested for histological evaluation.HE and Masson staining revealed that there were a large number of cells distributed in the inner wall of the scaffold,and some suspected epithelial cells and glands were found.Conclusion Decellularized aorta matrix scaffold hold great potential in serving as scaffold repairing defect of bile duct.

The features of idiopathic membranous nephropathy in elderly patients and its correlation with the expression of the expression of phospholipase A2 receptor / 中华老年医学杂志

Objective To investigate the clinico-pathological features of idiopathic membranous nephropathy (IMN ) and the expression of phospholipase A2 receptor (PLA2R ) in elderly patients. Methods A total of 109 elderly patients with IMN confirmed by renal biopsy at Wuxi People's Hospital from July 2008 to February 2015 were included.Data were retrospectively collected. Results (1)Participating patients with IMN had a mean age of (67.3 ± 5.4)years ,and 67.9% of them had hypertension and 65.1% had nephrotic syndrome.Compared with non-elderly patients ,elderly patients had a higher proportion with hypertension (67.9% vs.25.2%)(P＝0.000) ,higher systolic pressure[(143.1 ± 15.2)mmHg vs. (127.3 ± 13.3)mmHg](P＝ 0.000) ,higher diastolic pressure [(88.4 ± 10.0)mmHg vs. (80.2 ± 8.4)mmHg](P＝ 0.000) ,more severe tubulointerstitial lesions [(3.1±1.9)points vs.(2.0±1.9)points](P＝0.000),and lower eGFR[(70.9±22.9)ml·min－1· 1.73 m －2vs. (90.6 ± 27.1 ) ml·min－1·1.73 m －2] ( P ＝ 0.000 ). (2 ) There were more severe tubulointerstitial lesions[(4.7 ± 1.8)points vs. (2.4 ± 1.7)points ,2.9 ± 1.6 points](P ＝ 0.000 , 0.000)and lower eGFR[(50.4 ± 17.4)ml·min－1·1.73 m －2vs. (80.3 ± 19.7)ml·min－1·1.73 m －2, (72.3 ± 21.4)ml·min－1·1.73 m －2](P＝0.000 ,0.000)in elderly patients of pathological stage Ⅱ, compared with patients of pathological stages Ⅰ and Ⅰ-Ⅱ. (3)The rate of positive PLA2R was 82.4%.Patients with positive PLA2R had higher proteinuria[(4.5 ± 2.3)g vs. (2.9 ± 1.1)g](P＝0.042) ,lower eGFR[(66.8 ± 21.8)ml·min－1·1.73 m －2vs. (97.7 ± 16.0)ml·min－1·1.73 m －2](P＝0.000) ,and more severe tubulointerstitial lesions [(3.1 ± 2.0)points vs. (1.7 ± 1.1)points](P＝0.037)than patients with negative PLA2R. (4)Multiple regression analysis showed that PLA2R positive rate(P＝0.008) ,tubulointerstitial lesion(P＝0.000) ,and level of cholesterol(P＝0.025)were negatively correlated with eGFR (R2＝0.572). Conclusions Compared with non-elderly patients , elderly patients with IMN have poorer prognosis as a result of higher blood pressure and more severe tubulointerstitial lesions.Elderly patients with IMN of advanced pathological stages and positive PLA2R have more severe kidney injury and tubulointerstitial lesions ,resulting in poor prognosis.

Establishment of Eu3+ / Sm3+ dual?label time?resolved fluoroimmunoassay for measurement of anti?bodies to hepatitis C virus and its clinical application / 中华核医学与分子影像杂志

Objective To establish a time-resolved fluoroimmunoassay (TRFIA) system for simul-taneous measurement of immunoglobulin (Ig)M and IgG antibodies to HCV. Methods Recombinant HCV antigen was fixed on microtiter plates to detect serum HCV antibodies. Eu3+-labeled anti-human IgM and Sm3+-labeled anti-human IgG were prepared. HCV-IgM and HCV-IgG TRFIA were established using indirect assay and further optimized and evaluated. The one-sided 95th-percentile was used to calculate the cut-off (CO) values. Results Defining 1 sample/ CO (S/ CO) as measuring unit, the detection limit was 0.06 S/ CO for HCV-IgM and 0.15 S/ CO for HCV-IgG. When diluted a strong-positive specimen from 1 :12.5 to 1 :51200, there was a good liner range within 1 :12.5 to 1 :12800 for HCV-IgM and 1 :25 to 1 :6400 for HCV-IgG. The average intra-assay CV of HCV-IgM and HCV-IgG were 3.37％ and 3.66％, respectively, and the aver-age inter-assay CV were 6.52％ and 6.75％, respectively. Compared with enzyme-linked immunosorbent as-say (ELISA) kits, the positive conformity rate, the negative conformity rate and total conformity rate were 100.0％(20/ 20), 90.0％(18/ 20), 95.0％(38/ 40) for HCV-IgM TRFIA, and were 100.0％ (20/ 20), 95. 0％(19/ 20), 97.5％(39/ 40) for HCV-IgG TRFIA, respectively. Additionally, the established HCV-IgM and HCV-IgG assay kits presented good stability with a decline in the value of fluorescence of 11.1％and 9.5％ respectively after being stored at 37 ℃ for 7 d. Conclusions The established HCV-IgM and HCV-IgG TRFIA could simultaneously measure HCV-IgM and HCV-IgG antibodies at one step. The method has wider detectable range and may be a more sensitive, stable, and reliable method for diagnosing HCV in-fection.

Establishment and application of light initiated chemiluminescence assay of high mobility group box 1 / 中华核医学与分子影像杂志

Objective To establish a fast and quantitative light initiated chemiluminescent assay (LICA) method for high mobility group box1 (HMGB1) determination.Methods Two strains of paired HMGB 1 monoclonal antibodies were used.One was used to coat receptor microspheres.The other was labeled with biotin first and then composed with chain mildew element of affinity donor microsphere to form LICA method for HMGB1.After optimizing the reaction system,the technical specifications of the method was evaluated.Serum HMGB1 levels of common pneumonia patients (CPP) and severe pneumonia patients (SPP) were measured and compared with that of health controls.Two-sample t test was used.Results The sensitivity of LICA was 0.1 μg/L,with linear measurement ranging from 0.1 to 1 000 μg/L.The precisions of intra-and inter-analysis were 1.74％-2.92％ and 1.93％-3.73％ respectively,both were lower than 5％.The recovery rate was 99.74％ (range:94.53％-106.37％).The correlation coefficient of LICA and enzyme-linked immunosorbent assay (ELISA) was 0.888 2.The LICA method had good specificity and no obvious cross reaction with HMGB2 and HMGB3.The serum HMGB1 level in CPP (n=35) and SPP (n=25) was significantly higher than that in health controls (n=35):(6.76±3.13),(19.69±+9.04) vs (1.49±+0.74) μg/L;t values:-5.447 and-5.186,both P＜0.01.The HMGB1 levels between CPP and SPP were also significantly different (t=-3.500,P＜0.01).Conclusions The established LICA method of HMGB1 has high sensitivity and specificity with reliable results.This method is also homogeneous,fast and cleaning-free,thus has a good prospect in clinical application.

Dual-antigen sandwich-type time-resolved fluoroimmunoassay for phospholipase A2 receptor antibody, a new marker of membranous nephropathy / 中华核医学与分子影像杂志

Objective To establish a time-resolved fluoroimmunoassay (TRFIA) method for detecting M-type phospholipase A2 receptor (PLA2R) antibody and to investigate the diagnostic value of serum PLA2R antibody for idiopathic membranous nephropathy (IMN).Methods The microplates coated with recombined PLA2R antigen and Eu3+-streptavidin labeled PLA2R antigen were used to establish a dual-antigen sandwich-type TRFIA for PLA2R antibody detection (anti-PLA2R-TRFIA).The serum concentrations of PLA2R antibody in 63 IMN patients (36 males,27 females,age 25-75 years) and 90 healthy volunteers (30 males,60 females,age 22-53 years) were quantitatively analyzed.Kruskal-Wallis H test and MannWhitney u test were used to analyze the data.Results The range of anti-PLA2R-TRFIA was 0-10 mg/L and the sensitivity was 5 μg/L,while ED20,ED50 and ED80 of the standard curve were (0.144±0.012),(0.707±0.029) and (3.466±0.098) mg/L,respectively.The CV of inter-and intra-assay were 4.7％ and 5.1％,respectively.The average concentration of serum PLA2R antibody in healthy volunteers was 0.455(0.320,0.593) mg/L,but in IMN patients it was 2.690(0.717,7.750) mg/L (z=-3.688,P＜0.05).Meanwhile,the serum levels of PLA2R antibody in IMN patients were significantly different between different stages and ages (x2 values:10.328,9.716,both P＜0.05).According to the receiver operating characteristic (ROC) curve,when the diagnostic cut-off was set at 0.80 mg/L for IMN detection,the sensitivity and specificity of anti-PLA2R-TRFIA were 73.0％ (46/63) and 95.6％ (86/90),respectively.Conclusions AntiPLA2R-TRFIA has been well established.This quick and easy-performance method could increase the diagnostic accuracy for IMN.

The features and correlation analysis of clinico-pathological and expression of PLA2R in idiopathic mem-branous nephropathy / 实用医学杂志

Objective To investigate the features and correlation analysis of clinico-pathological and expression of phospholipase A2 receptor(PLA2R)in idiopathic membranous nephropathy. Methods A number of 244 patients of IMN proved by renal biopsy were recruited in Wuxi People's Hospital from July 2008 to February 2015. Data were restrospectively collected. Results In the 244 IMN patients(mean age 54.07 ± 15.22 years,130 males and 114 females),44.3% had hypertension and 62.7% had nephrotic syndrome. Compared with female patients ,male patients had more severe proteinuria and lower eGFR(P 8 g/24 h had higher level of cholesterol and blood pressure ,lower eGFR and more severe tubulointerstitial lesions(P < 0.05). Pathological stage Ⅰ and Ⅱaccounted for 98%,and there were more severe tubulointerstitial lesions and lower eGFR in the patients of stage Ⅱthan that in the patients of stageⅠandⅠ~Ⅱ(P<0.05). The positive rate of PLA2R accounted for 84.3%. Lower eGFR and more severe tubulointerstitial lesions were found in PLA2R-positive patients than those in PLA2R-neg-ative patients(P < 0.05). Multiple regression analysis showed that tubulointerstitial lesions(B =-7.253),hyper-tension ratio(B=-10.726)and the level of cholesterol(B=-2.077)had negative correlations with eGFR(P<0.01, R2=0.470). Conclusions IMN patients of male gender,grave proteinuria,high pathological stage and positive PLA2R should be treated more actively , since severe tubulointerstitial lesions and kidney injury were more common in those patients.

Biomechanical properties of cervical traction under neck muscle force / 医用生物力学

Objective To establish a three-dimensional(3D) finite element model of cervical vertebrae (C1-7),and study its biomechanical properties under muscle force by cervical traction,so as to provide references for clinical treatment.Methods On the basis of nonlinear finite element model of normal cervical vertebrae and combined with clinical traction methods,cervical traction at the extension angle of 0°,10°,20°,30°,40° under the same traction weight,was simulated by finite element analysis (FEA) software to obtain and select the joint force and muscle force that were appropriate for FEA on the model.Results In the process of cervical extension by traction,under the muscle force,the average maximum equivalent stress of cervical vertebrae,intervertebral disc and uncovertebral joints increased by 4.86,1.79,0.69 MPa,respectively,and the average maximum relative displacement of cervical vertebrae in sagittal and vertical axis direction increased by 1 1.1,1.26 mm,respectively.The biomechanical properties of cervical traction were similar to the FEA results reported in the literature.Conclusions Neck muscles play an active role in promoting the stress and displacement of cervical vertebrae,intervertebral discs and uncovertebral joints and it should be taken into consideration when performing cervical traction in clinic.In addition,the traction angle should not be too large:0.-20. is generally recommended as a relatively safe angle range at the initial stage.

Numerical modeling of a temperature field on the surface of skin tissues exposed to high temperature / 中国组织工程研究

BACKGROUND:Researchers suggest that thermal damage happens when the temperature of biological tissues is over 44℃. Accurate analysis of burn range wil facilitate diagnosis and therapy of skin burns. OBJECTIVE: To establish the mathematical model of the skin tissues subjected to high temperature and to analyze heat transfer process, and then to predict the burn range of the skin. METHODS:A finite element method was used to simulate the temperature field when the skin tissue was burned. And the relevant animal experiment was conducted to identify the mathematical model. RESULTS AND CONCLUSION:The temperature field was obtained by using the finite element method and the variation tendency between the theoretical data and the experimental data were agreed, but they did not fuly coincide. The scope of skin burn can be predicted by the present finite element method, and help to research the process of heat transfer.

The establishment and application of a time-resolved fluoroimmunoassay in detection of HBV large surface protein / 中华核医学与分子影像杂志

Objective To establish a method of TRFIA with high sensitivity and broad detecting range for serum HBV large surface protein (HBV-LP).Methods The monoclonal antibody of HBV-LP was covered on the microwell plate and incubated with HBV-LP in blood sample,then Eu3+ labeled antibody of HBs was added.HBV-LP in standard substance,blood samples of 66 chronic hepatitis B patients and 30 healthy controls was detected by the TRFIA and ELISA.x2 test and linear correlation analysis were used for data analysis.Results The dose-response curve of standard substance with TRFIA had good linear correlation (r=0.999).Normal reference range was established at 0-1.36 mg/L based on the ELISA results of 30 healthy controls.The sensitivity was 0.10 mg/L.The specificity was 100％ (30/30).Correlation coefficient between the TRFIA and the ELISA was 0.800 9 (P＜0.001).The positive detecting rates of the 2 methods were significantly different (89.4％(59/66) vs 77.3％(51/66),x2 =6.13,P＜0.01).The recovery rate for HBV-LP was between 95.93％-107.62％.The effective detecting range(CV＜10％) of TRFIA was 1.35-2 764.00 mg/L,and that of ELISA was 10.8-691.0 mg/L.Conclusion The TRFIA was established for HBV-LP detection with higher sensitivity and wider detecting range compared to ELISA.It has potential value for HBV screening and monitoring of antiviral therapy.

Identification of Inulae Herba, Inulae Flos and Their Closely Related Species Based on ITS2 Sequence / 世界科学技术-中医药现代化

This study aimed to identify Inulae Herba, Inulae Flos and their closely related species using the ITS2 bar-code, and secure the quality and clinical curative effect of these medicinal materials. DNA was extracted from Inula linariifolia, I. japonica, I. britanica, which are the original species of Inulae Herba and Inulae Flos, together with their closely related species. The ITS2 sequence was amplified by PCR and sequenced bidirectionally. Sequence assembly and generation of consensus sequence were conducted by the CodonCode Aligner. The genetic distances were comput-ed using MEGA 5.0 in accordance with the Kimura-2-parameter (K2P) model, and the phylogenetic tree constructed by the neighbor-joining (NJ) method. The results showed that the ITS2 sequences of the various species have stable variable sites. The intraspecific genetic distances among Inulae Herba and Inulae Flos were obviously lower than the interspecific genetic distance among the above two medicinal materials and its adulterants. The NJ tree based on ITS2 sequences can clearly differ from Inulae Herba, Inulae Flos and their closely related species. It is concluded that ITS2 sequence can be used as DNA barcode to identify Inulae Herba, Inulae Flos and their closely related species.

Comparison and Analysis for the Content of Dendrobine in Dendrobium nobile from Different Seasons and Various Parts / 世界科学技术-中医药现代化

To determine the content of dendrobine in Dendrobium nobile from different harvest times and plant parts, to research the inherent rule about it. GC with internal standard was used to determine. The content of dendrobine had significant differences in different periods and parts. The dendrobine content is higher in four-year root than in three-year root. The dendrobine content in the upper segment of stem is the highest, secondly is in the middle seg-ment, and in the low segment is the lowest. This offered evidence to determine the most appropriate harvest time and fair use of different parts for D.nobil.

Using ITS2 Sequence to Identify Descurainiae, Lepidii Semen and Its Adulterants / 世界科学技术-中医药现代化

Descurainiae, Lepidii Semen and their it adulterants were identified by analysising their ITS2 sequences. The genomic DNA was extracted from 46 samples including Descurainiae and Lepidii Semen and their it adulterants. Their ITS2 sequences were amplified, and purified PCR products were sequenced. Sequence assembly and consensus sequence generation were performed using CodonCode Aligner v 4.25. The genetic distances, variable sites and the neighbor-joning (NJ) phylogenetic tree were computed by MEGA 6.0 in accordance with the Kimura 2-parameter (K2P) model. The results showed that the intra-specific genetic distances of Descurainia sophia and Lepidium apetalum were 0.021 and 0.010, which were smaller than inter-specific ones of D. sophia, L. apetalum and their adulterants. The NJ tree showed that both D. sophia and L. apetalum were clustered into one monophyletic branch, and clearly separated with their sibling species. Therefore ITS2 sequence was able to identify Descurainiae and Lep-idii Semen and its adulterants to ensure the quality of medicines and clinical efficacy.

Identification of Xanthii Fructus and Its Adulterants Based on ITS2 Sequence / 世界科学技术-中医药现代化

Objective: To identify Xanthii Fructus and secure its quality and safety in medication. Methods: Total ge-nomic DNA was extracted from Xanthii Fructus and its adulterants. ITS2 sequences were amplified, and purified PCR products were sequenced. Sequence assembly and consensus sequence generation were performed using the CodonCode Aligner V 4.2. The Kimura 2-Parameter (K2P) distances were calculated using MEGA 5.0. The neigh-bor-joining (NJ) phylogenetic trees were constructed. Results: The intraspecific genetic distances of Xanthii Fructus were 0. The interspecific genetic distances between Xanthii Fructus and its adulterants were ranged from 0.009 to 0.542. The NJ tree showed that Xanthii Fructus could differ from its adulterants obviously. Conclusion: ITS2 can be used to identify Xanthii Fructus from its adulterants effectively, and our study further confirmed the effectiveness of ITS2 to identify traditional Chinese medicinal materials.

Study on the phenotype and the function of CD8αα+TCRαβ+regulatory T cells / 中华微生物学和免疫学杂志

Objective To investigate the phenotype and the immunoregulatory function of CD8αα+TCRαβ+regulatory T cells in peripheral blood samples from mice.Methods The distribution profile and the phenotype of CD8αα+TCRαβ+regulatory T cells in C57BL/6 mice were detected by flow cytometry.The cytokines released by CD8αα+TCRαβ+regulatory T cells upon the stimulation with anti-CD3 antibody were analyzed by cytometric bead array.The in vitro immunosuppressive activity of CD8αα+TCRαβ+regulatory T cells on activated CD4+T cells was analyzed by using flow cytometry and carboxyfluorescein succinimidyl ester ( CFSE ) .An adoptive cell transfer assay was set up to evaluate the immunoprotective effects of CD8αα+TCRαβ+ regulatory T cells in a mouse model of experimental autoimmune encephalomyelitis ( EAE) .Results CD8αα+TCRαβ+regulatory T cells were detected in liver, spleen and peripheral blood samples collected from na?ve C57BL/6 mice.Compared with CD8αβ+TCRαβ+regulatory T cells, CD8αα+TCRαβ+regulatory T cells showed a memory-activated phenotype of CD25+CD122high CD44high CD62Llow CD69high NK1.1+DX5+.CD8αα+TCRαβ+regulatory T cells could produce IL-2 after 24 hours stimulation with anti-CD3 antibody, followed by producing IFN-γ, TNF-α, IL-4, IL-17A and traces of IL-6 and IL-10. In vitro, CD8αα+TCRαβ+regulatory T cells specifically suppressed the proliferation of activated CD4+T cells ( P<0.01 ).Moreover, they could delay the onset of EAE in mice and reduce clinical score (P<0.01).Conclusion CD8αα+TCRαβ+regulatory T cells were a unique population with immunoregula-tory function, which could be used as a potential therapeutic target in the treatment of autoimmune disease.

Conservation Status of Shennongjia Traditional Chinese Medicine Resources and Its Sustainable Utilization Strategy / 世界科学技术-中医药现代化

Shennongjia is well known as the national treasure of traditional Chinese medicine resources at home and abroad. In order to provide Shennongjia with better protection of traditional Chinese medicine and promote the sustainable utilization of its resources, based on the specific analysis of She nnongj ia′s conservation status and available resources, we put forward primary strategies and specific measures for the sustainable utilization framework of Shennongjia traditional Chinese medicine resources, in view of resources conservation, development, utilization, as well as the resources administration.

Identification of Toxic Hyoscyami Semen and Its Adulterants Using DNA Barcoding / 世界科学技术-中医药现代化

The misuse of toxic drugsisseriousone of the threats of public health. In this study, toxic Hyoscyami Semen and its adulterants were identified by DNA barcoding. The genomic DNA was extracted from 61 samples including Hyoscyami Semen and its adulterants by reagent kit method. Their ITS2 sequences were amplified, and purified PCR products were sequenced. Sequence assembly and consensus sequence generation were performed using CodonCode Aligner v 4.25. The genetic distances, variable sites and the neighbor-joning (NJ) phylogenetic tree were computed by MEGA 6.0 in accordance with the Kimura 2-parameter(K2P) model. The results showed that the intra-specific genetic distances of Hyoscyamusniger were 0.005 which were smaller than inter -specific ones (0.360) of H. niger and their adulterants. The NJ tree showed that H. niger was clustered into one monophyletic branch, and clearly separated with other species. Therefore, ITS2 sequence was able to identify Hyoscyami Semen and its adulterants to ensure the safty of medicines.