Objective To analyze the genetic diversity of eight species of Caragana Fabr. in Ordos Plateau. Methods To study the genetic diversity of the eight species with ISSR method. Results The total percentage of polymorphic bands (PPB) of the eight species was up to 100%, which was extremely high and showed an abundant genetic diversity. However, the genetic diversity of different species was not the same. The genetic diversity of C. tibetica, C. korshinskii, C. stenophylla, C. roborovskyi, and C. intermedia was higher, while that of C. opulens and C. purdomii was lower, and that of C. brachypoda was the lowest. The analytic results of the Nei′s gene diversities and the Shannon′s information index were similar. Different species could be differentiated through UPGMA method and characteristic banding pattern of ISSR. Conclusion ISSR Molecular marker could be used in molecular authentication and hereditary background study of the species of Caragana Fabr. These results provide the scientific basis of developing effective protection measures for these species.

Objective To select the strains which can produce tanshinone ⅡA like its host plant Salvia miltiorrhiza bung. Methods A total of 50 strains of endophytic fungi were isolated from healthy, living and symptomless tissues of Salvia miltiorrhiza bung, among which 29 strains were obtained from the root, 14 from the stem, 3 from the leaf, 3 from the flower and 1 from the seed. Their antimicrobial activities against nine different bacteria, including both Gram-negative and Gram-positive bacteria, were measured by Oxford plate agar diffusion bioassay. Results Our data showed that all but four strains had significant antibacterial activities on at least one indicator bacterium to some extent, and five strains (DR1, DR4, DR16, DR18 and DF2) manifested quite prominent antibacterial activities against certain pathogenic bacteria. In some degree, it might indicate that this endophytic fungus isolated from the tissues of Salvia miltiorrhiza bung has a potential value as a natural antibacterial medicine as well. Thin layer chromatography (TLC) and high-performance liquid chromatography (HPLC) were carried out to test selected strains, both inside and outside of the cell to see if any strain can produce tanshinone ⅡA. The result showed that extracts from three strains, labeled as DR12 (outside cell), DR21 (inside cell) and DF3 (inside cell), had a component with the same Rf value in TLC assay as that of authentic tanshinone ⅡA. The extract from DR12 (outside cell) and DR21 (inside cell) had a peak at retention time identical to that of authentic tanshinone ⅡA in HPLC. Conclusion The fungi appear to produce the bioactive compound tanshinone ⅡA, and they could be used to produce tanshinone ⅡA by fermentation. It provides a new way to synthesize this natural medicine.

Objective To investigate the replantation and postoperative rehabilitation methods for simultaneous amputation of 10 fingers and both forearms.Methods A case of replantation for simultaneous amputation of both forearms and 10 fingers was carried out with microsurgery method in September,2014.The replantation involved 3 teams over 11 hours and 55 minutes to consequently conduct alternate anterograde and retrograde replantation and accurate blood vessels,nerve anastomosis and further followed with physical therapy and occupational therapy treatment in 2 weeks of the surgery.Results All the amputated arms and fingers revived after the surgery.After 14 months follow-up,function of wrist flexion and expansion was normal,superficial and deep sensory functions on hands were good,function of thumb and finger grip,pinch and opposition had partially recovered,the two-point discrimination was 8-10 mm,and all of above rated good according to the temporary criteria of the upper limb functionality set forth by Hand Surgery Branch of China Medical Association.Conclusion For the case of simultaneous amputation of both forearms and 10 fingers,it is very likely to carry out successful replantation as well as achieve satisfactory function restoration with excellent teamwork and accurate vessel and nerve anastomosis under microsurgery as well as rehabilitation treatment afterwards.

Objective To discuss the clinical effect of thumb and fingers reconstruction with vascular anastomosis transplantation from toes.Methods From April 2009 to April 2013,166 cases of thumb and finger defect were treated,including 46 cases Ⅰ °-Ⅲ° thumb defect and 120 cases Ⅰ °-Ⅴ° finger defects.Sixty-two cases were emergency reconstructed by vascular anastomosis transplantation from toes,the other 104 cases were subemergency reconstructed.Early functional rehabilitation was carried out postoperative.Results All 208 thumb and fingers in 166 cases with these procedures were survived.Patients were followed up from 4 to 24 months,averaged of 1 l months.The reconstructed thumb and fingers were all with abundant blood supply,having similar shape to the normal thumb and fingers,good pain and temperature sensation,with two-point discrimination of 6-10 mm,with normal range of joint activity,flexible function of finger to finger and finger to palm.Most patients were satisfied with the thumb and finger shape,regained life and work ability as before.The donor sites had no obvious discomfort,and walking and weight-bearing function remained normal.Conclusion For patients with thumbs Ⅰ °-Ⅲ° and fingers Ⅰ °-Ⅴ ° degree of traumatic defect,emergency and subemergency reconstruction of fhumb and fingers by vascular anastomosis transplantation from toes have good clinical effect and less damage to the donor site.

OBJECTIVE:To investigate the compatible stability of Levetiracetam(Lev)injection with 3 injections. METHODS:Each Lev injection 1000 mg mixed with 0.9% Sodium chloride injection 100 mL,5% Glucose injection 100 mL or Sodium lactate Ringer's injection 100 mL respectively. Under the light condition,at 25 ℃,the color and clarification degree of mixtures were ob-served at different time points within 24 h after mixing;pH value and the number of insoluble particles were determined. The contents of related impurities(impurity A,B,C,D,2-hydroxypyridine)and Lev in mixtures were determined by HPLC. RESULTS:Under above condition,all mixtures were colorless clear liquid within 24 h;pH value had no significant change (RSD<1%,n=7);the number of insoluble particles was no more than the range stated in Chinese Pharmacopeia(2015 edition). Impurity B and C were not detected;the contents of other impurities were in line with the requirements of foreign pharmacopeia. No marked change was noted for relative content of Lev(RSD<1%,n=7). CONCLUSIONS:After mixing with 0.9% Sodium chloride injection,5% Glucose injec-tion or Sodium lactate Ringer's injection,Lev injection keep stable at 25℃within 24 h under the light condition.

Hepatitis B surface antigen (HBsAg) carrying preS sequences could be an ideal candidate for a new hepatitis B virus (HBV) vaccine with higher efficacy. Here we report the success in achieving efficient and stable expression of hepatitis B virus S antigen and preS1 epitope fusion protein (S/preS1) in CHO cells. The HMRCHEF53u/Neo-S/preS1 expression vector carrying S/preS1 gene was constructed and transfected into CHO-S cells. A stable and high-expression CHO cell line, named 10G6, was selected by ELISA and limiting dilution analysis. Western blotting analysis showed S/preS1 expressed from 10G6 cells possessed both S and preS1 antigenicity. 10G6 cells displayed characters of favorable growth and stable S/preS1 expression in repeated batch cultures as evaluated by viable cell density, viability and S/preS1 concentration. And cultivation of 10G6 cells in fed-batch mode resulted in S/preS1 production at 17-20 mg/L with viable cell density at 7 x 10(6)-10 x 10(6) cells/mL.
Animals ; CHO Cells ; Cricetulus ; Epitopes ; biosynthesis ; genetics ; Hepatitis B Surface Antigens ; biosynthesis ; genetics ; immunology ; Hepatitis B Vaccines ; biosynthesis ; genetics ; Hepatitis B virus ; Protein Precursors ; biosynthesis ; genetics ; immunology ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; Transfection

Objective To investigate the feasibility of non-invasive quantitative evaluation of portal pressure (Pp) by contrast enhanced ultrasonography (CEUS).Methods 18 portal hypertension patients (PTH group) were performed with CEUS within one week before splenectomy and pericardial devascularization,and 20 healthy volunteers as controls were recruited in this study (control group).Hepatic-right kidney sectionwas chosen to calculate the area under curve of portal vein/hepatic artery (Qp/Qa) and the perfusion intensity of portal vein/hepatic artery (Ip/Ia) through time intensity curves (TIC) of liver parenchyma generated from CEUS images.Pp was measured by intra-operative mesenteric vein catheter,and the correlation betweenPp and Qp/Qa,Ip/Ia were analyzed by Pearson correlation test.Results The levels of Qp/Qa and Ip/Ia in the PTH group were 2.28 ± 0.66 and 0.35 ± 0.14 respectively,which were both significantly declined than that in the controlgroup (5.72 ± 3.69 and 1.97 ± 0.17).In the PTH group,the correlation coefficient were-0.747 and-0.617,and the linear regression equations were Y =-83 X + 5.013 andY =-15X + 0.837,which indicated that Qp/Qa and Ip/Ia had significant correlation with Pp.Conclusions CEUS parameters,including Qp/Qa and Ip/Ia,are significantly correlated to Pp in portal hypertension patients,which indicate that CEUS could be a new non-invasive clinical method for evaluating Pp.

Objective To evaluate the BALB/c murine infective effects in different concentrations and different aerosol challenge times by Bordetella pertussis.Methods Four experiment groups according to different concentrations and different aerosol challenge times were designed.BALB/c murines were challenged by aerosol way.Group 1: 1010cfu/mL Bordetella pertussis challenge 15 min, group 2: 1010cfu/mL challenge 30 min, group 3: 109cfu/mL challenge 30 min, group 4: 1011cfu/mL challenge 30 min, using the normal saline challenge 30 min as control.At 0d,3d,7d,14d and 21d after challenge, the WBCs of all groups were measured and lung tissues were homogenized to calculate the bordetella pertussis clone in lung.Results After 3 days of challenge, WBCs in all groups were slightly increased.The WBCs of group 1, group 2, group 3 and group 4 were significantly increased after 7 days, with the average numbers of 8.52×109 per/L, 1.74×1010per/L, 1.15×1010per/L and 5×1010per/L, respectively.After 14 days, they were 1.77×1010per/L, 1.67×1010per/L, 1.27×1010per/L and 3.84×1010per/L respectively.WBCs in all groups were dramatically declined after 21 days.The WBC of negative control group had no obvious change during the whole process with the stable number of 3.4~7.0×109per/L.Bordetella pertussis were detected in lung of all experimental groups in each sampling point.The CFU in lung wase at peak at 7d or 14d after challenge, which was obviously decreased at 21d.Conclusion This aerosol challenge method can establish a bordetella pertussis infection mouse model successfully.

Due to the insufficient supply of embryonated chicken eggs,the preparation of large quantities of inactivated influenza vaccines will require an alternative virus culture system after the emergence or reemergence of a pandemic influenza virus.The Vero cell is one of the ideal options since it was used for producing many kinds of human vaccines.However,most of the influenza viruses can not grow well in Vero cells.To develop a new influenza vaccine with Vero cells as a substrate,the virus needs to adapt to this cell substrate to maintain high growth characteristics.By serial passages in Vero cells,the B/Yunnan/2/2005va(B)strain was successfully adapted to Vero cells,with the hemagglutination titer(HAT)of the virus reaching 1:512.The high growth characteristic of this strain is stable up to 21 passages.The strain was identified by hemagglutination inhibition (HAI)test and sequencing respectively;the HA;gene sequence of the virus was cloned and analyzed.The screening and establishment of high growth B virus provides an important tool for influenza vaccine production in Vero cells.

Objective To study the correlation between ultrasonographic features of thyroid nodule and BRAFV600Emutation. Methods A total of 179 patients with 194 suspicious throid nodules were included in this study. They underwent ultrasound, biopsy, pathology and BRAFV600Emutation examination between October 2015 and February 2016 at Chinese PLA General Hospital. The size of nodules were (1.1±0.8) cm. The size, echo, boundary, shape aspect ratio, calcification and capsular invasion of nodules were investigated. The correlation between ultrasonographic features of thyroid nodule and BRAFV600Emutation analyzed by chis-square test and Logistic Regression analysis using statistical data as independent variable, BRAFV600Emutation as dependent variable. Results There were significant different in nodule′s ratio, boundary, capsular invasion characteristic between the BRAFV600Epositive group and the BRAFV600Enegative group(χ2=11.174,45.517,11.046,all P < 0.05),and these signs are possibly associated with BRAFV600Emutation by logistic regression model analysis(OR=2.276,95%CI:1.117-4.638, P < 0.05; OR=8.412, 95%CI: 3.836-18.448,P < 0.001; OR=2.582, 95%CI: 1.138-5.860,P < 0.05). Conclusions The ratio, boundary, capsular invasion characteristic of thyroid nodules are possibly associated with BRAFV600Emutation. These signs can be used to predict BRAFV600Emutation and facilitate subsequent treatment for such nodules.