Objective:To identify enolase,47 kD allergen,from Litopenaeus vannamei by Mass spectrometry. Methods: The proteins were extracted from Litopenaeus vannamei tissue with acetone precipitation method. The protein components were analyzed by SDS-PAGE and Western blot. By using Matrix-Assisted Laser Desorption/Ionization time of flight mass spectrometry ( MALDI-TOF/TOF-MS) ,the 47 kD allergen from Litopenaeus vannamei was identified as enolase. Results:By SDS-PAGE,we proved that the native protein components from Litopenaeus vannamei were completely. According to the Western blot result more than 14 components could react with the positive serum. MALDI-TOF/TOF analysis results showed that the suspected proteins were enolase. A sensitization frequency was 55%. Conclusion:Enolase was identified as a new allergen of Litopenaeus vannamei.

Objective To observe the changes of inflammatory factors in acute lung injury (ALI) in mice induced by lipopolysaccharide (LPS), and to explore the influence of different doses of LPS on ALI onset and progress at different time points. Methods Intratracheally, LPS at the dosages of 2.5, 5.0, 7.5 and 10.0 mg/kg were administered to a total of 210 C57BL/6 mice, and according to the difference in dosage, they were divided into four groups. The ALI model was replicated by intratracheally dropping of LPS. And a normal control group and a normal saline control group were established (each, n=10). The changes of index of pathological lung tissue and lung tissue wet/dry (W/D) ratio were observed at 1, 2, 4, and 8 hours after injury, and simultaneously, the levels of norepinephrine (NE), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and protein in serum and bronchoalveolar lavage fluid (BALF) were detected. Results ①The degree of lung injury induced by LPS was dose-and time-dependent.②With the increase of LPS dosage and prolongation of time, in LPS group, the lung W/D ratio and the index of pathological lung tissue were increased;additionally, the levels of NE, TNF-α, IL-6 and protein in serum or BALF were also significantly increased. The critical occurrence point of acute respiratory distress syndrome (ARDS) with specific characteristics was at 5.0 mg/kg of LPS acting for 4 hours [lung W/D ratio: 4.97±0.41, index of pathological changes of lung tissue (score): 5.60±1.52; serum NE (ng/L): 379.99±27.65, TNF-α (ng/L): 159.15±20.62, IL-6 (ng/L): 177.15±29.13;BALF NE (mg/kg):105.85±13.66, TNF-α(mg/kg):227.22±48.01, IL-6 (mg/kg):251.55±54.08, total protein (g/L):1.59±0.37]. The injury induced by LPS acting for 8 hours in the dosage group 10.0 mg/kg was the most significant in comparisons with other groups of dosages at the same time points [lung W/D ratio:5.10±0.18 vs. 5.01±0.43, 5.01±0.19, 4.91±0.30; index of pathological changes of lung tissue (score): 9.20±1.48 vs. 8.00±1.00, 6.00±1.22, 4.40±0.89;serum NE (ng/L): 447.43±34.63 vs. 419.23±30.62, 391.16±54.91, 372.59±51.52; TNF-α(ng/L): 205.99±31.31 vs. 181.01±25.11, 161.01±13.98, 138.83±28.95; IL-6 (ng/L): 233.76±34.84 vs. 206.21±26.68, 186.58±26.54, 156.99±28.83;BALF NE (mg/kg):190.82±41.75 vs. 153.30±35.42, 122.64±25.15, 80.23±13.69;TNF-α(mg/kg):305.24±72.99 vs. 292.77±38.07, 249.60±35.20, 193.63±10.83; IL-6 (mg/kg): 354.81±67.79 vs. 303.02±54.24, 272.43±32.34, 197.64±12.35;total protein (g/L):2.31±0.30 vs. 2.02±0.26, 1.62±0.19, 1.10±0.24, P<0.05 or P<0.01]. Conclusions The severity of ALI induced by LPS in mice was positively correlated to LPS dosage and duration of its action. After administration of LPS 5 mg/kg for 4 hours, remarkable characteristic manifestations of ARDS occur in mice, reaching the critical point.

Objective To construct adenovirus vector containing firefly luciferase reporter gene (AdLuc) and infect bone marrow mesenchymal stem cells (BMSC),then to take bioluminescence imaging in vitro and in vivo for identification.Methods The luciferase gene was amplified with PCR from psiCHECK-2 plasmid and cloned into the adenoviral shuttle vector (pShuttle-CMV).It was confirmed by Nhe Ⅰ/Xba Ⅰ digestion and sequencing.PShuttle-CMV-Luc and backbone vector (pAdeno) were homologous recombined.Then the recombinant plasmid was packaged in HEK293 cells and the virus titer was detected.The BMSC were infected by the recombinant adenovirus.The bioluminescence imaging in vitro was performed to determine the best multiplicity of infection (MOI),and the relationship between bioluminescence intensity and MOI was analyzed by curve fitting regression analysis.Viability was evaluated via Trypan blue staining.The transfected BMSC (l× 106) were implanted into the muscles of forelimb of SD rats,and then tracked by bioluminescence imaging in vivo.Cell viability was compared using two-way repeated measures analysis of variance between groups.Results Enzyme digestion and sequence analysis indicated that Ad-Luc was successfully constructed.The virus titer was 1 × 1010 plaque forming unit (PFU)/ml.The bioluminescence detection in vitro showed that Ad-Luc could infect BMSC high efficiently to express luciferase and the best MOI was 50.The bioluminescence intensity enhanced with increase of MOI (R2 =0.98).No statistically significant difference was found in cell viability between transfected and untransfected BMSC at 1,3,5,7 d.The cell survival rates were (92.5±2.3)％ vs (94.1±1.8)％,(91.4±0.9)％ vs (92.7±2.0)％,(92.1±1.6)％ vs (93.3± 2.4) ％,(91.9 ± 1.5) ％ vs (93.0 ± 3.1) ％,respectively (F =4.38,P ＞ 0.05).The bioluminescence imaging in vivo showed that BMSC survived 1,3,7 d after implantation.However,bioluminescence signal decreased gradually over time.Conclusion It is feasible to apply the optical reporter gene imaging for tracing transplanted stem cells in vitro and in vivo due to the effective transformation of luciferase reporter gene into BMSC by adenovirus vector.

Objective:To establish a GC fingerprint analysis method for identification of volatile oil in ultramicro-powder of Houttuynia cordata from different habitats, then to control the quality sensitively.Methods: GC was used to analyze the volatile constituents of ultramicro-powder of Houttuynia cordata from 12 different habitats;SPDTM-1CapillaryColumn(30m?0.32mm?0.25?m),temperature programming and FID detector were applied.Results: The mutual mode of GC fingerprints was set up and the similar degrees to the volatile oil from of different habitats were compared.Conclusion:The GC fingerprints of volatile oil in ultramicro-powder can be used to identify the Houttuynia cordata from different habitats and evaluate its quality.

AIM: To establish the quality standard for Tongfeng Huadu Tincture (Radix Gentianae Macrophyllae, Radix Angelicae Pubescentis, Rhizoma Chuanxiong, Herba Asari, etc.). METHODS: The Radix Gentianae Macrophyllae, Radix Angelicae Pubescentis, Rhizoma Chuanxiong, Herba Asari of Tongfeng Huadu Tincture were identified by TLC. Strychnine of Tincture was determined by HPLC. RESULTS: The average recovery was 97.07% and RSD and 2.1% (n=5). CONCLUSION: The method is reliable, accurate and specific. It can be used for quality control of Tongfeng Huadu Tincture.

The study found that the physician-patient trust crisis results from overreliance on technology trust instead of interpersonal trust and institutional trust. The alleged “Paternalistic government innovation”in healthcare service has caused wastes of healthcare resources and gap below public expectancy due to its incompetence in resolving social problems,further eroding institutional legality and intensifying such crisis.This research aimed to identify government accountabilities in building such trust from three aspects.

[Objective] Molecular distillation technique was applied for the separation and purification of Atractylodes Lancea oil (ALO) to increase the yield of atractylodin. [Methods] Atractylodes Lancea oil prepared with supercritical extraction was refined by molecular distillation technique. The optimum refining conditions were selected by uniform design containing two factors of temperature and vacuum and five levels. The content of atractylodin from refined Atractylodes Lancea oil was measured and the remainder after isolation was detected by high performance liquid chromatography (HPLC). [Results] A yield rate of atraotylodin from Atractylodes Lancea oil was 52.17% at the temperature of 105℃ and under the vacuum of 100 Pa. [Conclusion] The application of molecular distillation technique makes it easy to collect the volatile atractylodin from Atractylodes Lancea oil at lower temperature and under higher vacuum; this method ensures the yield rate of atractylodin over 50% and has no problem of environmental pollution.

Objective:To analyze the prognostic factors of robot-assisted radical cystectomy (RARC).Methods:The clinical data of 224 patients underwent RARC from December 2014 to December 2018 in Nanjing Drum Hospital were reviewed. There were 193 males and 31 females, aged 36-92 years, with mean of 68 years. There were 7 patients(3.1%)undergoing neoadjuvant chemotherapy, the ASA scores of 125 patients (55.8%) were more than 2, and the mean body mass index was 23.4(15.4-35.5)kg/m 2. All patients were treated with RARC, with 72(32.1%) patients undergoing intraoperative blood transfusion. Kaplan-Meier method was used to analyze recurrence-free survival rate (RFS), cancer-specific survival rate (CSS) and overall survival rate (OS). Cox multivariate risk ratio model was used to evaluate the correlation between survival outcome and perioperative and pathological factors in patients treated with RARC. Results:For pathological status, there were 82 of ≤T 1, 64 of T 2, 57 of T 3 and 21 of T 4. Of all the patients, 49(21.9%) had lymph node metastasis, 12(5.4%) had positive surgical margin, 82(36.6%) had lymphovascular invasion(LVI), and 41(18.3%) underwent adjuvant chemotherapy. Follow-up time was between 11-60 months, and the median follow-up time was 24 months. The 5-year cumulative OS, RFS and CSS were 57.15%, 48.84% and 59.60%, respectively. Univariate Cox regression analysis showed that T stage( HR=5.764, 95% CI 1.926-17.249, P=0.002; HR=4.086, 95% CI 1.611-10.364, P=0.003; HR=9.391, 95% CI 2.118-41.637, P=0.003), N stage( HR=6.446, 95% CI 3.438-12.087, P<0.001; HR=5.661, 95% CI 3.086-10.385, P<0.001; HR=5.980, 95% CI 2.982-11.992, P<0.001), LVI( HR=3.319, 95% CI 2.008-5.486, P<0.001; HR=2.894, 95% CI 1.782-4.701, P<0.001; HR=3.471, 95% CI 2.017-5.974, P<0.001), American Society of Anesthesia (ASA)score( HR=2.888, 95% CI 1.619-5.150, P<0.001; HR=1.765, 95% CI 1.060-2.940, P=0.029; HR=2.612, 95% CI 1.424-4.792, P=0.002), body mass index( HR=0.886, 95% CI 0.819-0.957, P=0.002; HR=0.885, 95% CI 0.819-0.955, P=0.002; HR=0.862, 95% CI 0.792-0.938, P=0.001), age( HR=1.580, 95% CI 1.250-1.997, P<0.001; HR=1.362, 95% CI 1.088-1.705, P=0.007; HR=1.530, 95% CI 1.190-1.968, P=0.001) and intraoperative blood transfusion( HR=1.899, 95% CI 1.160-3.108, P=0.011; HR=2.218, 95% CI 1.371-3.587, P=0.001; HR=2.227, 95% CI 1.312-3.782, P=0.003) were significantly related to survival outcome. Multivariate Cox regression analysis showed that T stage( HR=4.506, 95% CI 1.433-14.175, P=0.01; HR=3.159, 95% CI 1.180-8.454, P=0.022; HR=7.810, 95% CI 1.674-36.444, P=0.009), N stage( HR=6.096, 95% CI 2.981-12.467, P<0.001; HR=5.368, 95% CI 2.683-10.740, P<0.001; HR=5.539, 95% CI 2.497-12.288, P<0.001) and ASA score( HR=6.180, 95% CI 2.371-16.110, P<0.001; HR=2.702, 95% CI 1.175-6.215, P=0.019; HR=6.471, 95% CI 2.290-18.286, P<0.001) were independent predictors of RFS, CSS and OS, and adjuvant chemotherapy( R=0.434, 95% CI 0.202-0.930, P=0.032) could only predict OS. Conclusion:T stage, N stage and ASA were main independent predictors of postoperative survival outcomes, and adjuvant chemotherapy was independent predictor of OS.

Objective:To understand the status of outpatients′medical care and their impact on the doctor -pa-tient trust in Beijing .Methods:Three tertiary hospitals were selected due to most concentrated high -quality medi-cal resources , the largest number of patient visits , universalism and particularism trust could be compared , taking field observations , personal interviews and questionnaires methods .Results:The main factors affecting clinical doctor-patient trust are time, the doctor skills, medical ethics, medical and prescription drug division , etc.are appropriate and effective .Conclusion:Play the role of pyramidmedical system′s overall effectiveness , and es-tablish a patient-centered system for outpatient care as soon as possible a comprehensive grasp of the overall con -struction team doctor , continue to strengthen the standardized management of medication .