Abstract: Objective　To collect extensively drug-resistant tuberculosis (XDR-TB) Mycobacterium tuberculosis strains isolated from Xi'an City between 2019 and 2020, and analyze the drug resistance patterns of XDR-TB strains to second-line anti-tuberculosis drugs and the occurrence of new defined extensively drug-resistant tuberculosis in Xi'an, in order to provide evidence for guiding clinical drug use of multidrug-resistant tuberculosis (MDR-TB) patients. Methods A total of 3 088 strains of Mycobacterium tuberculosis that underwent phenotypic drug susceptibility testing at Xi'an Chest Hospital from January 2019 to December 2020 were retrospectively selected to analyze the resistance of anti-tuberculosis drug. Among the stored MDR-TB strains, 114 strains of preserved multidrug-resistant Mycobacterium tuberculosis were randomly selected for bedaquiline and linezolid susceptibility testing. Combined with the results of previous second-line drug susceptibility testing, the incidence of newly defined extensive drug resistance was analyzed. Results Among the 3 088 Mycobacterium tuberculosis strains analyzed, 411 strains (14.3%) showed resistance to isoniazid, 347 strains (11.2%) showed resistance to rifampicin, 142 strains (4.6%) showed resistance to ethambutol, 550 strains (17.8%) showed resistance to streptomycin, and 237 strains (7.6%) exhibited multidrug resistance. Of 237 MDR-TB strains, the resistance rates of ethambutol, moxifloxacin, rifampicin, sodium para-aminosalicylate, prothioconazole, capreomycin, amikacin, and clofazimine were 44.3%, 26.6%, 33.3%, 24.1%, 5.1%, 4.2%, 3.0%, and 2.5%, respectively. Among the randomly selected 114 MDR-TB strains, none showed resistance to bedaquiline, three showed resistance to linezolid, and one strain met the new definition for extensively drug-resistant tuberculosis. Conclusion In Xi'an City, high rates of resistance among MDR-TB strains are observed for ethambutol, quinolone and sodium para-aminosalicylate, and the drug susceptibility tests should be obtained as much as possible when using these drugs. The incidence of new definition extensively drug-resistant tuberculosis is low, and bedaquiline and linezolid remain effective drugs for the treatment of multidrug-resistant tuberculosis even without drug susceptibility testing results.

Objective Analysis of the teaching effect of the micro-classroom in the experimental teaching of diagnostics.Methods In the 50 classes of undergraduate clinical specialty of 2014 level of guangdong medical mniversity,2 classes were selected as the control group (n=67) by random number table method,2 classes were selected as the experience group (n=65).Theoretical knowledge and clinical skills were calculated.Recognition of two groups of students on two kinds of teaching methods.The data of two groups were compared using t-test and chi-square test.Results The results of the theoretical knowledge and clinical skills of the experimental group were significantly higher than the control group (P< 0.05).There was a significant difference between the two groups (P< 0.05),which was found to be able to deepen the understanding of theoretical knowledge,to improve the ability of clinical skills operation,to cultivate clinical thinking and to mobilize the enthusiasm of independent learning.Conclusions Micro classroom teaching can significantly improve the quality of experimental training of diagnoses.

Objective:To construct and express the fusion protein between ZZ affinity peptide and alkaline phosphatase and examine its biological activities.Methods:The alkaline phosphatase gene was cloned into pEZZ 18 vector containing ZZ peptide gene resulted in the pEZZ-AP recombinant vector.Then the vector was transformed and expressed in E.coli DH5α.And HisTrap affinity chromatography was employed to separate and purify the target protein.After analyzed by Western blot , ZZ-AP fusion protein was applied to immunocytochemistry as an alterative second antibody.Results:The result of SDS-PAGE showed that the fusion protein with a relative molecular weight was consistent with the theoretical values (62 kD).The fusion protein has rabbit IgG-binding ability and en-zymatic activity of alkaline phosphatase ,those were validated in Western blot;and it produced a good signal that was comparable in its staining pattern to that generated with goat anti-rabbit IgG-AP in immunocytochemistry.Conclusion: The ZZ-AP fusion protein was constructed successfully ,it has rabbit IgG-binding ability and enzymatic activity of alkaline phosphatase.We anticipate that the ZZ-AP fusion protein has a potential application in immunoassay field.

BACKGROUND: Previous studies has explored the effects of nerve growth factor (NGF) on cultured rabbit Schwann cells as well as preparation of acellular nerve allografts, in addition, the neural complex was prepared in vitro.OBJECTIVE: To investigate the effect of acellular nerve allografts on the functional recovery and reconstruction of the nerve-muscle structure of the sciatic nerve defect in rabbits.DESIGN, TIME AND SETTING: The randomized controlled animal experiment was performed at the Center Laboratory of Third Hospital of Hebei Medical University from March 2006 to June 2007.MATERIALS: Totally 37 healthy, adult, New Zealand, white rabbits were selected. One rabbit was prepared for extracted nerve bridge, and the others were randomly divided into the experimental and control groups, with 18 animals in each group.METHODS: The double sides of sciatic nerve were removed, divested surrounding tissues, and cut into 3 cm segments, then placed in TdtonX-100 solution for 12 hours, washing with distilled water. The procedure was repeated, and TritonX-100 totally used for 96 hours. Then the extracted nerve bridge was conserved in Hank's solution at temperature of 4 ℃. The concentration of 2~(nd) passage Schwann cell was regulated to 1 ×10~(11)/L, and injected to extracted nerve bridge, followed by DMEM culture, to obtain neural complex. Rabbits in the experimental group were prepared a 20 m defects above knee joints, and the recellularized neural complex were transplanted into the defective sciatic nerve. Rabbits in the control group were prepared by transplanting extracted nerve bridge without Schwann cells.MAIN OUTCOME MEASURES: The elcosis and healing of feet were observed at weeks 4, 8, and 16 after operation, meantime,the recovery of axon of distal end peroneal nerves, myelin sheath, tibialis anterior, as well as motor end plate were detected by electromyogram, light microscope and transmission electron microscope.RESULTS: At weeks 4, 8 and 16 after operation, the rejection was not found in the operating field. The experimental group was better than the control group in healing of ulcers, the number of nerve fibres, the ultramicrostructure of medullary sheaths, the regeneration nerve, the structure of the motor end plate and muscle. At 4 weeks after operation, no nerve conduction was found in 2 groups, but at weeks 8 and 16 after operation, the experimental group was better than the control group in wet weight of tibialis anterior muscle and nerve conduction velocity (P < 0.05).CONCLUSION: The recellularized neural complex can significantly promote the reconstruction of the nerve-muscle structure and functional recovery of injured sciatic nerve.

Dioscin is the main raw material for the synthesis of steroid hormone drugs. Currently, the direct acid hydrolysis is the mainly industrial production method for dioscin. However, the use of strong acid can not only destroy the structure of dioscin resulting in very low yield, but also produce a large amount of waste water and residues, which seriously pollute the environment. So the clean production of dioscin is the urgent demand of water conservation and environmental protection. In the paper, the recent research pro-gresses in the clean production technology and process of dioscin were summarized, and the advantages and problems were discussed in order to provide reference for the improvement and application of the new technology and process.

Objective To establish a method for simultaneous determination for the contents of four flavones (ononin, calycosin, genistein and formononetin) of Hedysari Radix in Gansu Province with quantitative analysis of multi-components by single-marker (QAMS); To prove its feasibility and accuracy. Methods Calycosin was taken as internal standard substance. Relative correction factors (RCF) of ononin, genistein and formononetin to calycosin were established. The contents of ononin, calycosin, genistein and formononetin were determined to realize QAMS. Results RCF was with good repeatability. The results of QAMS were consistent with the results of the external standard method. Conclusion The method that determines the contents of ononin, genistein and formononetin with calycosin as internal standard substance, can be used for quantitative analysis of Hedysari Radix.

AIM To establish an ultra-performance liquid-chromatography tandem mass spectrometry (UPLC-MS/MS) method for the simultaneous content determination of five constituents in Jiangzhi Huoxue Tablets (Polygoni multiflori Radix,Astragali Radix,Chuanxiong Rhizoma,etc.).METHODS The 50％ methanol extract of this drug was performed on a 40 ℃ thermostatic Restek UItra BiPh column (100 mm × 2.1 mm,5 μm),with the mobile phase comprising of acetonitrile (containing 0.1％ formic acid)-0.1％ formic acid flowing at 0.4 mL/min in a gradient elution manner.RESULTS Stilbene glycoside,tanshinone Ⅱ A,emodin,ferulic acid and puerarin showed good linear relationships within the ranges of 4.01-1 027,0.7-187,1.48-380,3.98-1 020 and4.285-1 097 ng/mL (r ＞0.994 0),whose average recoveries were 98.57％-101.0％ with the RSDs (n =6) of less than 4.79％.CONCLUSION This specific and sensitive method can be used for the quality control of Jiangzhi Huoxue Tablets.

ObjectiveTo investigate the methods and clinical significance of detecting PLAC4 and COL6A1 gene on fetal chromosome 21 from maternal peripheral blood. Methods From Oct. 2008 to Nov. 2009 30 normal pregnancies in Weifang People's Hospital were selected as pregnant group, and 9 nonpregnant women were selected as control group. Quantitative real-time PCR was used to determine transcript levels of the target genes ( PLAC4 and COL6A1 ) in blood samples. Correlation between the expression level and gestational age was analyzed. Results ( 1 ) PLAG4 mRNA was detected in peripheral blood of all pregnant women. Its maximum level was 12. 760 × 103 copies/ml, whereas the minimum was 2. 105 × 103 copies/ml, and the average value is 6. 612 × 103 copies/ml. In control group the PLAC4 mRNA could not be detected. There was statistically significant difference ( P ＜ 0. 01 ) between the two groups. ( 2 ) COL6A1 mRNA is detected in pregnant group and control group, and the concentration was 6. 847 × 103 copies/ml and 7. 322 × 103 copies/ml respectively, with no statistically significant difference ( P ＞ 0. 05 ). ( 3 ) Correlation analysis: there was no relationship between the level of PLAC4, COL6A1 mRNA and the gestational age, the correlation coefficients (r) were 0. 29 and 0. 31, and the P values were 0. 121 and 0. 168 respectively. Conclusions COL6A1 mRNA can be detected in both pregnant group and control group, so it is not specific for pregnancy. PLAC4 mRNA can be detected only in pregnant women, so it has specificity in pregnancy and can be a discriminative marker gene for prenatal dignosis of trisomy 21 fetuses.

Objective:To prepare the site-specific biotinylation of enhanced green fluorescence protein with double biotin molecules using Avi-tag technology.Methods:The EGFP gene was prepared by PCR and cloned into pdi-Avitag resulting the vector pEGFP-( Avitag) 2.The fusion protein EGFP-( Avitag ) 2 was expressed in E.coli DH5αand purified by employing IMAC.The site-specific biotinylation was implemented by BirA enzyme in vitro, and then was identified by competitive ELISA and Western blot.Results:The recombinant prokaryotic expression vector pEGFP-(Avitag)2 was correctly constructed,and EGFP-(Avitag)2 fusion was successfully expressed in E.coli DH5α.The results of competitive ELISA and Western blot showed that the EGFP-( Avitag) 2 could be site-specific biotinylation with double biotin molecules based on Avi-tag technology.Conclusion: The site-specific biotinylation of EGFP with double biotin molecules is successfully prepared,and we anticipate that can be used for BAS to improve the sensitivity and specificity of immunosensors.

Objective: Lymph node metastasis (LNM) often occurs in cN0 papillary thyroid microcarcinoma (PTMC). The risk factors for lymph node metastasis, especially for high-volume metastasis, were investigated in this study. Methods: The medical records of 1,268 consecutive PTMC patients admitted in the Peking Union Medical College Hospital from 2013 to 2014 were reviewed. Their clinical and pathological features were collected. Univariate and multivariate analyses were performed to identify the risk factors for LNM/highvolume LNM. Results: Of the 1,268 patients, 416 patients (32.8％) and 43 (3.4％) had LNM and high-volume LNM, respectively. According to the univariate analysis results for the risk factors of LNM, male (42.22％ vs. 30.26％, P<0.01), <40 years (<40 years, 48.39％; 40-59 years, 27.62％; ≥60 years 22.45％, P<0.03), multifocality (41.00％ vs. 29.03％, P<0.01), without chronic thyroiditis (36.44％ vs. 20.62％,P<0.01), tumor size >0.5 cm (35.77％ vs. 23.05％, P<0.01) were associated with LNM. Meanwhile, according to the multivariate analysis results, male, multifocality, and tumor size >0.5 cm are independent risk factors for LNM (OR=1.516, 1.743, and 1.788, respectively, all P<0.05). The protective factors for LNM are 40-59 years, ≥60 years, and chronic thyroiditis (OR 0.388, 0.301, and 0.472, respectively,all P<0.05). In the univariate analysis of risk factors for high-volume LNM, the results indicated that being male (6.30％ vs. 2.61％, P= 0.005), <40 years (<40 years, 7.62％; 40-59 years, 2.05％; ≥60 years 0, P<0.001), and tumor size >0.5 cm (4.01％ vs. 1.36％, P=0.027) are associated with high-volume LNM. In multivariate analysis, the results suggest that being male is an independent risk factor for LNM (OR=2.383, P=0.002), whereas age of 40-59 years is a protective factor for LNM (OR=0.270, P<0.001). Conclusion: Lymph node metastasis often ocucrs in cN0 PTMC, whereas high-volume LNM is rare. Being male and <40 years old are risk factors for both LNM and highvolume LNM.