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1.
Article in Chinese | WPRIM | ID: wpr-906524

ABSTRACT

Objective:To analyze the chemical constituents in Euodiae Fructus by ultra-high performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS/MS). Method:The chromatographic separation was performed on an ACQUITY UPLC BEH C<sub>18</sub> column (2.1 mm×100 mm, 1.7 μm) with acetonitrile (A)-0.1% formic acid aqueous solution (B) as mobile phase (0-3 min, 6%A; 3-4 min, 6%-10%A; 4-7 min, 10%-12%A; 7-8 min, 12%-14%A; 8-13 min, 14%-15%A; 13-15 min, 15%-20%A; 15-18 min, 20%-30%A; 18-21 min, 30%-49%A; 21-25 min, 49%-51%A; 25-27 min, 51%-73%A; 27-30 min, 73%-80%A; 30-31 min, 80%-100%A; 31-32 min, 100%A) for gradient elution. The column temperature was 35 ℃, and the flow rate was 0.4 mL·min<sup>-1</sup>. Mass spectrometry was performed using an electrospray ionization and data were collected in positive and negative ion modes, and the detection range was <italic>m</italic>/<italic>z</italic> 100-1 200. The chemical constituents in Euodiae Fructus were identified rapidly and comprehensively based on the accurate relative molecular mass and combined with literature data and reference substances. Result:A total of 92 chemical constituents were speculatively identified from the 70% methanol extract of Euodiae Fructus, including 39 alkaloids, 19 flavonoids, 12 limonoids, 20 phenolic acids and 2 organic acids. Among them, 26 compounds were confirmed by the reference substances. Conclusion:The compound types of Euodiae Fructus are multiple and quite different in polarity. The chemical compositions of Euodiae Fructus from different regions and species are similar. The established method is rapid and accurate, with which the chemical compositions of Euodiae Fructus have been identified comprehensively. Therefore, this study provides an experimental reference for further clarifying active and toxic constituents of Euodiae Fructus.

2.
Chinese Medical Journal ; (24): 1457-1464, 2021.
Article in English | WPRIM | ID: wpr-878178

ABSTRACT

BACKGROUND@#Clinical observational studies revealed that 99Tc-methylene diphosphonate (99Tc-MDP) could reduce joint pain and swollenness in rheumatoid arthritis (RA) patients. This multicenter, randomized, double-blind, double-dummy study aimed to evaluate the effects of 99Tc-MDP plus methotrexate (MTX) vs. MTX alone or 99Tc-MDP alone on disease activity and structural damage in MTX-naïve Chinese patients with moderate to severe RA.@*METHODS@#Eligible patients with moderate to severely active RA were randomized to receive 99Tc-MDP plus MTX (n = 59) vs. MTX (n = 59) alone or 99Tc-MDP (n = 59) alone for 48 weeks from six study sites across four provinces in China. The primary outcomes were the American College of Rheumatology 20% improvement (ACR20) response rates at week 24 and changes in modified total Sharp score at week 48.@*RESULTS@#At week 24, the proportion of participants achieving ACR20 was significantly higher in the MTX + 99Tc-MDP combination group (69.5%) than that in the MTX group (50.8%) or 99Tc-MDP group (47.5%) (P = 0.03 for MTX + 99Tc-MDP vs. MTX, and MTX + 99Tc-MDP vs.99Tc-MDP, respectively). The participants in the MTX + 99Tc-MDP group and the 99Tc-MDP group had significantly less important radiographic progression than the participants in the MTX group over the 48 weeks (MTX + 99Tc-MDP vs. MTX: P = 0.03, 99Tc-MDP vs. MTX: P = 0.03, respectively). There was no significant difference in terms of adverse events (AEs) among the groups. No serious AEs were observed.@*CONCLUSIONS@#This study demonstrated that the combination of 99Tc-MDP with MTX inhibited structural damage and improved disease activity in RA patients compared with MTX and 99Tc-MDP monotherapies, without increasing the rate of AEs. Additional clinical studies of 99Tc-MDP therapy in patients with RA are warranted.@*TRIAL REGISTRATION@#Chictr.org, ChiCTR-IPR-14005684; http://www.chictr.org.cn/showproj.aspx?proj=10088.


Subject(s)
Antirheumatic Agents/therapeutic use , Arthritis, Rheumatoid/drug therapy , China , Diphosphonates , Double-Blind Method , Drug Therapy, Combination , Humans , Methotrexate/therapeutic use , Technetium/therapeutic use , Treatment Outcome
3.
Article in Chinese | WPRIM | ID: wpr-849643

ABSTRACT

Objective To analyze the status of misdiagnosis of exertional heat stroke (EHS), its causes and influence on prognosis. Methods The clinical data of patients with EHS in 9 military hospitals from January 2012 to December 2018 were analyzed retrospectively. According to the time of diagnosis (from onset to the establishment of preliminary or suspected diagnosis), the patients were divided into ≤0.5 h, 0.5-1 h, 1-3 h, 3-6 h and >6 h groups. The number of organs involved and the clinical outcome (death or survival) of patients in each group were recorded, and the relationship between delayed diagnosis and prognostic indexes was analyzed. Through the analysis of misdiagnosis-related medical records, the distribution characteristics and possible causes of misdiagnosis were found. Results Among 122 EHS patients, 23 died, with a total fatality of 18.9%. The diagnosis time showed a skewed distribution, and the median (quartile interval) was 1.5(2.63) h. The correlation analysis between the time of diagnosis and the time of initiation of cooling showed a positive correlation (r=0.871, P<0.05). The number of involved organs increased with the delay of diagnosis, and it was significantly higher in patients diagnosed more than 6 h than that in patients diagnosed in the early stage (within 0.5 h, P<0.05). The risk of death also increased significantly with the delay of diagnosis, and the fatality rate of patients diagnosed more than 6 h was significantly higher than that of patients diagnosed in the early stage (within 0.5 h, P<0.05). The composition ratio of misdiagnosis varied with time. The misdiagnosis rate within 0.5 hours of onset was 87.7%. The diagnosis was mainly based on symptomatic description (64.5%), followed by misdiagnosis as nervous system disease (24.3%). The main causes for the delay in diagnosis were the lack of typical clinical manifestations in the early stage of the disease and the lack of understanding of the disease in the on-site emergency medical personnel. Conclusions The misdiagnosis rate would be high in the early stage of EHS, and it may be significantly related to organ injury and prognosis. The main cause of misdiagnosis in the early stage of EHS might be the lack of understanding of the disease in the on-site emergency medical personnel, which suggests an urgent need to improve the EHS recognition by on-site emergency medical personnel.

4.
Chinese Medical Journal ; (24): 2899-2904, 2019.
Article in English | WPRIM | ID: wpr-781747

ABSTRACT

BACKGROUND@#Clinical outcomes of undifferentiated arthritis (UA) are diverse, and only 40% of patients with UA develop rheumatoid arthritis (RA) after 3 years. Discovering predictive markers at disease onset for further intervention is critical. Therefore, our objective was to analyze the clinical outcomes of UA and ascertain the predictors for RA development.@*METHODS@#We performed a prospective, multi-center study from January 2013 to October 2016 among Chinese patients diagnosed with UA in 22 tertiary-care hospitals. Clinical and serological parameters were obtained at recruitment. Follow-up was undertaken in all patients every 12 weeks for 2 years. Predictive factors of disease progression were identified using multivariate Cox proportional hazards regression.@*RESULTS@#A total of 234 patients were recruited in this study, and 17 (7.3%) patients failed to follow up during the study. Among the 217 patients who completed the study, 83 (38.2%) patients went into remission. UA patients who developed RA had a higher rheumatoid factor (RF)-positivity (42.9% vs. 16.8%, χ = 8.228, P = 0.008), anti-cyclic citrullinated peptide (CCP) antibody-positivity (66.7% vs. 10.7%, χ = 43.897, P < 0.001), and double-positivity rate of RF and anti-CCP antibody (38.1% vs. 4.1%, χ = 32.131, P < 0.001) than those who did not. Anti-CCP antibody but not RF was an independent predictor for RA development (hazard ratio 18.017, 95% confidence interval: 5.803-55.938; P < 0.001).@*CONCLUSION@#As an independent predictor of RA, anti-CCP antibody should be tested at disease onset in all patients with UA.

5.
Article in Chinese | WPRIM | ID: wpr-701140

ABSTRACT

AIM:To investigate the effect and mechanism of osthole on increasing the cytotoxicity of doxorubi -cin(DOX)to prostate cancer cells.METHODS:MTT assay was performed to evaluate the viability of LNCaP cells trea-ted with osthole and DOX.The protein expression of silent information regulator 1(SIRT1),p53,acetylated p53 and Pu-ma,as well as release of cytochrome C and activation of caspase-9 and caspase-3 in the LNCaP cells treated with osthole and DOX were determined by Western blot.The apoptosis of the LNCaP cells treated with osthole and DOX was analyzed by flow cytometry.RESULTS:Osthole significantly increased the cytotoxicity of DOX against p 53-wildtype prostate cancer cell line LNCaP.Osthole significantly inhibited the expression of SIRT 1 in the LNCaP cells.Transfection with SIRT1 plas-mid decreased the cytotoxicity of osthole and DOX co-treatment against LNCaP cells.Combination with osthole and DOX significantly induced the over-expression and acetylation of p53.Transfection with p53 siRNA significantly decreased the synergistic effect of osthole on cytotoxicity of DOX-treated LNCaP cells.Combination with osthole and DOX significantly in-duced the release of cytochrome C into the cytoplasm from mitochondria,followed by activation of caspase-9 and its down-stream molecule caspase-3,thus leading to cell apoptosis in the LNCaP cells.CONCLUSION:Osthole promotes the p53-dependent apoptosis in DOX-treated prostate cancer LNCaP cells by down-regulating the expression of SIRT1.

6.
Chinese Medical Journal ; (24): 2683-2692, 2018.
Article in English | WPRIM | ID: wpr-775034

ABSTRACT

Background@#Tofacitinib is an oral Janus kinase inhibitor for the treatment of rheumatoid arthritis (RA). This study assessed the efficacy and safety of tofacitinib in Chinese patients with RA enrolled in Phase 3 and long-term extension (LTE) studies.@*Methods@#ORAL Sync was a 1-year, randomized, placebo-controlled, Phase 3 trial. Patients received tofacitinib 5 or 10 mg twice daily (BID) or placebo advanced to tofacitinib 5 or 10 mg BID at 3 or 6 months. All patients remained on ≥1 background conventional synthetic disease-modifying antirheumatic drug. ORAL Sequel is an open-label LTE study (data-cut: March 2015; data collection and analyses were ongoing, and study database was not locked at the time of analysis; study was closed in 2017). Efficacy outcomes: American College of Rheumatology (ACR) 20/50/70 response rates and Disease Activity Score in 28 joints using erythrocyte sedimentation rate (DAS28-4 [ESR]). Patient- and physician-reported outcomes: Health Assessment Questionnaire-Disability Index (HAQ-DI), Patient and Physician Global Assessment of Arthritis, and pain (visual analog scale). Safety was assessed throughout.@*Results@#ORAL Sync included 218 patients; 192 were subsequently enrolled into ORAL Sequel. In ORAL Sync, more patients achieved ACR20 (tofacitinib 5 mg BID, 67.4%; 10 mg BID, 70.6%; placebo, 34.1%) and DAS28-4 (ESR) <2.6 (tofacitinib 5 mg BID, 7.1%; 10 mg BID, 13.1%; placebo, 2.3%) with tofacitinib versus placebo at Month 6. Mean changes from baseline in HAQ-DI were greater with tofacitinib versus placebo at Month 6. In ORAL Sequel, efficacy was consistent to Month 48. Incidence rates for adverse events of special interest in tofacitinib-treated patients were similar to the global population.@*Conclusions@#Tofacitinib significantly reduced signs/symptoms and improved physical function and quality of life in Chinese patients with moderate-to-severely active RA up to Month 48. The safety profile was consistent with the global population.@*Clinical Trial Identifier@#NCT00856544 and NCT00413699.


Subject(s)
Administration, Oral , Adult , Aged , Arthritis, Rheumatoid , Drug Therapy , Asians , Female , Humans , Male , Middle Aged , Piperidines , Therapeutic Uses , Protein Kinase Inhibitors , Therapeutic Uses , Pyrimidines , Therapeutic Uses , Pyrroles , Therapeutic Uses , Surveys and Questionnaires , Young Adult
7.
Acta Pharmaceutica Sinica ; (12): 1157-1164, 2017.
Article in Chinese | WPRIM | ID: wpr-779708

ABSTRACT

An ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry(UHPLC-Q-TOF/MS)method has been established to analyze the bioactive components of the aqueous extract of Euodiae Fructusand detect the metabolites in rats after oral administration of the aqueous extract. Compounds were detected by analyzing their retention times, high resolution mass data, mass spectra and comparing with the reference substances. As a result, 27 compounds were characterized from the aqueous extract of Euodiae Fructus, and 25 compounds were identified on the basis of the reference substances. 16 parent compounds and 35 metabolites were detected in rats' plasma, urine and feces samples, and all of the parent compounds were determined unambiguously through comparison with the reference substances. Four classes of compounds were identified from the aqueous extract of Euodiae Fructus, including phenolic acids, limonins, flavonoid glycosides and alkaloids. Phenolic acids, flavonoid glycosides and alkaloids were detected in rats' plasma, urine and feces, while limonins were detected in the urine and feces. The main metabolism pathways of these compounds in rats might include hydroxyl, hydrogenation, methylation, sulfating, glucuronidation, and so on. The results of this study provide references in the material basis of Euodiae Fructus in vivo.

8.
Parenteral & Enteral Nutrition ; (6): 346-350, 2017.
Article in Chinese | WPRIM | ID: wpr-665458

ABSTRACT

Objective:TThe aim of this study was to observe the changes of the volume and distribution of body fluid after abdominal surgery,and further to explore its characteristics and influence factors.Methods:Sixtyone patients were included between March and June in 2016.The volume of intracellular water (ICW),extracellular water (ECW) and total body water (TBW) was estimated by InbodyS 10 on preoperative day 1 (PreD 1) and postoperative day (POD) 1,3,5 and 7.Furthermore,the patients were grouped according to the age,sex,type of operation,operation time and the daily liquid input,and the influence factors of postoperative fluid volume were analyzed in each subgroup.Results:Comparing to that of PreD1,the ICW,ECW,and TBW,mainly ECW,were increased significantly on POD1 (P< 0.05),and the level of postoperative fluid volume was decreased to that of PreD1 between POD3 and POD7.It showed that there was difference in the net increasing of body fluid on POD1 between different type of the patients.Obviously,the net increasing of fluid volume in woman on POD 1 was more than that in man.The net increasing of fluid volume on POD1 was correlated with the operation time and net liquid input.Conclusion:The fluid retention was found in the early stage of postoperative patients,and mainly exists in ECW.The main influence factors resulting postoperative fluid retention were prolonged operation time and increased net fluid input.

9.
Article in Chinese | WPRIM | ID: wpr-607690

ABSTRACT

BACKGROUND: Currently, a variety of new dressings have been on the market, which are diversified and exhibit multifunctional trends. However, ideal wound dressings are still in exploration.OBJECTIVE: To introduce the basic physiological function and polymer advantage of dextran as well as its effects to promote wound healing in combination with other macromolecule materials, in order to impel the development of dextran as a new medical dressing .METHODS: A computer-based search of CNKI, PubMed, WanFang, VIP databases was performed to retrieve reviews or research articles addressing dextran and medical dressings published from January 2000 to December 2016. The keywords were dressing, dextran, wound healing in Chinese and English, respectively.Finally, 31 articles were included in result analysis.RESULTS AND CONCLUSION: Dextran has the physiological functions of promoting wound healing and immune function. These physiological functions are the basis of dextran as a medical dressing. Additionally, dextran has some polymer advantages, such as water absorption, biodegradability and non-toxicity. It is noteworthy that dextran can be combined with macromolecules to produce new polymer materials that can promote wound healing in animal experiments. Taken together, dextran, as a medical dressing, has a broad clinical prospect in wound healing.

10.
Article in Chinese | WPRIM | ID: wpr-255151

ABSTRACT

Ubiquitin-specific protease(USP), which belongs to cysteine protease, is an important member of the deubiquitinating enzyme family(DUB). USP plays an important role in the immune response against viral infections, in which it can regulate the production of type I interferon through various ways to initiate or weaken the antiviral immune response. USP2b, USP3, USP18, USP25, UL36USP and HAUSP play a role of antivirus; while USP4, USP13, USP15 and USP17 negatively regulate antiviral immune response. In this article we review the recent progress on roles of USP family in antiviral immune response.


Subject(s)
Humans , Interferon Type I , Allergy and Immunology , Ubiquitin-Specific Proteases , Allergy and Immunology , Virus Diseases , Allergy and Immunology
11.
Chinese Journal of Biotechnology ; (12): 595-604, 2014.
Article in Chinese | WPRIM | ID: wpr-279480

ABSTRACT

Secretory anti-gp96 scFv fragment was expressed in Pichia pastoris to obtain a small molecule antibody that specifically recognizes heat shock protein gp96. The gp96-scFv fragment gene was synthesized and cloned to Pichia pastoris expression plasmid pPICZa-A. Pichia pastoris X33 was electroporated with the linearized recombinant expression vector, and expression of gp96-scFv fragment was induced by methanol. The His-tagged recombinant protein was then purified by affinity chromatography and analyzed with SDS-PAGE and Western blotting assays. The biological activities of recombinant gp96-scFv fragment were determined by Western blotting, Immunofluorescence, ELISA and FACS assays. The gp96-scFv fragment was expressed successfully in Pichia pastoris. About 50 mg of recombinant protein could be purified from 1 liter of the Pichia pastoris culture supernatant. Its molecular weight was about 15 kDa. The gp96-scFv fragment could specifically bind to gp96 protein by Western blotting, immunofluorescence, ELISA and FACS analyses. Pichia pastoris-expressed gp96-scFv fragment specifically recognizes gp96 protein, which could be used for Western blotting, Immunofluorescence, ELISA and FACS analyses.


Subject(s)
Blotting, Western , Chromatography, Affinity , Electrophoresis, Polyacrylamide Gel , Enzyme-Linked Immunosorbent Assay , Membrane Glycoproteins , Allergy and Immunology , Pichia , Metabolism , Plasmids , Recombinant Proteins , Single-Chain Antibodies
12.
Article in Chinese | WPRIM | ID: wpr-452638

ABSTRACT

Objective To analyze the effects of adenohypophysis function after treating with thyrotropin releasing hormone and its clinical significance in patients with brain trauma. Methods There were 22 cases with traumatic brain injuries from July 2010 to September 2012 in Chinese people's Liberation Army nine eight hospital,after injuried within 4 to 12 hours,then 1 1 cases who were given thyrotropin-releasing hormone(TRH)were selected as experimental group,while 1 1 cases who were given the same amount of isotonic saline were selected as control group,then the score of GCS, ICS,RLS85 and the improvement of adenohypophysis function were observed. Results After treatment,the score of glasgow coma scale (GCS ), innsbruck coma scale (ICS),and the reaction level scale (RLS85)between two groups were significantly increased in three days compared with before treatment,and within three days after injury situation,the improvements of ICS and RLS85 in experimental group were better than control group(P<0.05 ). Compared with control group,the levels of each gland pituitary hormone in experimental group were significantly increased(P<0.05 ),and on the third day,the growth hormone (GH)was reduced significantly,finally 50%of that in control group. Conclusion Patients with brain injury treated with thyrotropin releasing hormone,has no significant adverse reactions,with the characteristics of safe and effective.

13.
Protein & Cell ; (12): 317-327, 2014.
Article in English | WPRIM | ID: wpr-757497

ABSTRACT

Cytotoxic T cells (CTLs) play a key role in the control of Hepatitis B virus (HBV) infection and viral clearance. However, most of identified CTL epitopes are derived from HBV of genotypes A and D, and few have been defined in virus of genotypes B and C which are more prevalent in Asia. As HBV core protein (HBc) is the most conservative and immunogenic component, in this study we used an overlapping 9-mer peptide pool covering HBc to screen and identify specific CTL epitopes. An unconventional HLA-A2-restricted epitope HBc141-149 was discovered and structurally characterized by crystallization analysis. The immunogenicity and anti-HBV activity were further determined in HBV and HLA-A2 transgenic mice. Finally, we show that mutations in HBc141-149 epitope are associated with viral parameters and disease progression in HBV infected patients. Our data therefore provide insights into the structure characteristics of this unconventional epitope binding to MHC-I molecules, as well as epitope specific CTL activity that orchestrate T cell response and immune evasion in HBV infected patients.


Subject(s)
Adult , Amino Acid Sequence , Animals , Binding Sites , Epitopes , Chemistry , Allergy and Immunology , Metabolism , Female , Genotype , HEK293 Cells , HLA-A2 Antigen , Metabolism , Hepatitis B Core Antigens , Chemistry , Allergy and Immunology , Metabolism , Hepatitis B virus , Genetics , Metabolism , Humans , Hydrogen Bonding , Male , Mice , Mice, Inbred BALB C , Mice, Transgenic , Middle Aged , Molecular Dynamics Simulation , Mutation , Protein Binding , Protein Structure, Tertiary , T-Lymphocytes, Cytotoxic , Allergy and Immunology , Metabolism
14.
Protein & Cell ; (12): 348-356, 2014.
Article in English | WPRIM | ID: wpr-757487

ABSTRACT

During virus infection, viral RNAs and mRNAs function as blueprints for viral protein synthesis and possibly as pathogen-associated molecular patterns (PAMPs) in innate immunity. Here, considering recent research progress in microRNAs (miRNAs) and competitive endogenous RNAs (ceRNAs), we speculate that viral RNAs act as sponges and can sequester endogenous miRNAs within infected cells, thus cross-regulating the stability and translational efficiency of host mRNAs with shared miRNA response elements. This cross-talk and these reciprocal interactions between viral RNAs and host mRNAs are termed "competitive viral and host RNAs" (cvhRNAs). We further provide recent experimental evidence for the existence of cvhRNAs networks in hepatitis B virus (HBV), as well as Herpesvirus saimiri (HVS), lytic murine cytomegalovirus (MCMV) and human cytomegalovirus (HCMV) infections. In addition, the cvhRNA hypothesis also predicts possible cross-regulation between host and other viruses, such as hepatitis C virus (HCV), HIV, influenza virus, human papillomaviruses (HPV). Since the interaction between miRNAs and viral RNAs also inevitably leads to repression of viral RNA function, we speculate that virus may evolve either to employ cvhRNA networks or to avoid miRNA targeting for optimal fitness within the host. CvhRNA networks may therefore play a fundamental role in the regulation of viral replication, infection establishment, and viral pathogenesis.


Subject(s)
Animals , DNA Viruses , Genetics , Physiology , Host-Pathogen Interactions , Physiology , Humans , MicroRNAs , Metabolism , RNA Viruses , Genetics , Physiology , RNA, Messenger , Metabolism , RNA, Viral , Metabolism , Virus Diseases , Allergy and Immunology , Virology , Virus Replication
15.
Article in English | WPRIM | ID: wpr-819692

ABSTRACT

OBJECTIVE@#To evaluate of the curative effect of human umbilical cord mesenchymal stem cells (hUC-MSCs) on rat acute radiation pneumonitis.@*METHODS@#Fourty rats were randomly divided into control group, radiation group, stem cell prevention group, stem cell treatment group and prednisone treatment group. All rats except those in the control group were radiated with X ray to establish the acute radiation pneumonitis damage model. The hUC-MSCs cultured in vitro was administrated to the rats of the prevention group via tail vein (1×10(6) cells/kg BW) 24 h before the radiation, while the same administration was performed in the rats of the treatment group 24 h after the radiation. After 24 h post the radiation, the rats in the radiation group were given 0.4 mL physiological saline, and those in the prednisone group were given 1 mg/kg prednisone. All rats were observed and executed 72 h after the radiation to detect lung histological changes.@*RESULTS@#After the administration of hUC-MSCs, the survival status of the rats in the prevention group and treatment group was obviously better than that in the control group. As shown by the histological staining, the morphology, proliferation activity and bronchial state of lung tissues were better in the prevention group and treatment group than in the control group.@*CONCLUSIONS@#The hUC-MSCs have definite therapeutic effects on acute radiation pneumonitis in rats.


Subject(s)
Animals , Antigens, CD , Metabolism , Disease Models, Animal , Humans , Lung , Pathology , Male , Mesenchymal Stem Cell Transplantation , Methods , Mesenchymal Stem Cells , Cell Biology , Radiation Pneumonitis , Pathology , General Surgery , Rats , Rats, Wistar
16.
Acta Pharmaceutica Sinica ; (12): 1618-1623, 2013.
Article in Chinese | WPRIM | ID: wpr-298035

ABSTRACT

Tanshinones are the bioactive components of the Chinese medicinal herb Salvia miltiorrhiza, while its biosynthetic pathway remains to be characterized. Rapid identification and characterization of the genes correlated to tanshinones biosynthesis is very important. As one of the intermediates of tanshinones biosynthesis, the ferruginol content is relative low in both root and engineered bacteria. It is urgent to construct an efficient system for conversion of miltiradiene to ferruginol to obtain large amount of ferruginol as the substrates for further identifying other downstream genes involved in tanshinones biosynthesis. In this study, we constructed the whole-cell yeast biocatalysts co-expressing miltiradiene oxidase CYP76AH1 and cytochrome P450 reductases (SmCPR1) from Salvia miltiorrhiza, and then characterized it with RT-PCR. After permeabilization, the yeast whole-cell could catalyze turnover of miltiradiene to ferruginol efficiently through single-step biotransformation with a conversion efficiency up to 69.9%. The yeast whole-cell biocatalyst described here not only provide an efficient platform for producing ferruginol in recombinant yeast but also an alternative strategy for identifying other CYP genes involved in tanshinones biosynthesis.


Subject(s)
Biosynthetic Pathways , Biotransformation , Cytochrome P-450 Enzyme System , Genetics , Metabolism , Diterpenes , Metabolism , Abietanes , Chemistry , Electrophoresis, Agar Gel , Gene Amplification , NADPH-Ferrihemoprotein Reductase , Genetics , Metabolism , Open Reading Frames , Plasmids , Saccharomyces cerevisiae , Genetics , Metabolism , Salvia miltiorrhiza , Chemistry
17.
Chinese Journal of Biotechnology ; (12): 1765-1775, 2013.
Article in English | WPRIM | ID: wpr-242455

ABSTRACT

Although DNA vaccination is now a promising strategy against hepatitis B virus (HBV) infection, this approach has relatively modest antiviral effect, indicating that immunosuppressive mechanisms may occur in the long-term established infection. In this study, we studied the immunogenicity and anti-HBV efficiency of a combination of HBV surface (HBsAg) and core (HBcAg) DNA vaccine, enhanced by heat shock protein (HSP) gp96 or HSP70 and mediated by in vivo electroporation. Immunization with gp96 adjuvanted HBsAg/HBcAg DNA formulation induced potent T cell and antibody immunity against HBsAg and HBcAg. Notably, treatment with gp96 or HSP70 as adjuvant resulted in reduction of Treg populations by around 20%. Moreover, compared with nonimmunized control mice, immunization with gp96 or HSP70 adjuvanted DNA vaccine dramatically decreased serum HBsAg and viral DNA levels, and HBcAg expression in liver. These results may therefore provide an effective strategy for designing gp96-based DNA vaccine for immunotherapy of chronic HBV infection.


Subject(s)
Adjuvants, Immunologic , Animals , Electroporation , HSP70 Heat-Shock Proteins , Allergy and Immunology , Hepatitis B Core Antigens , Allergy and Immunology , Hepatitis B Surface Antigens , Allergy and Immunology , Hepatitis B Vaccines , Allergy and Immunology , Hepatitis B, Chronic , Immunization , Membrane Glycoproteins , Allergy and Immunology , Mice , Mice, Transgenic , Vaccines, DNA , Allergy and Immunology
18.
Article in English | WPRIM | ID: wpr-820522

ABSTRACT

OBJECTIVE@#To investigate the expression characteristics of TNF-α in myocardium and hepatic tissue of rats with compound stress of hyperthermia and lipopolysaccharide (LPS).@*METHODS@#Male SPF Wistar rats were randomly divided into room temperature+physiological saline group (Group C), hyperthermia+physiological saline group (Group H), room temperature+LPS group (Group L) and hyperthermia+LPS group (Group HL). The rats were put in simulated climate cabin. Group HL and Group H were exposed in the environment at a dry bulb temperature (TDB) of (35.0±0.5) °C, while Group L and Group C were exposed in the environment at a TDB of (26.0±0.5) °C. The rats in Group HL and Group L were given tail intravenous injection of LPS 10 mg/kg, while the rats in Group H and Group C were given tail intravenous injection of 9 g/L NaCl 10 mL/kg. After the stress, immunohistochemical SABC staining method was used to detect the expression characteristics of TNF-α in myocardium and hepatic tissue of rats, and those rats were given routine pathological examinations.@*RESULTS@#The expression of TNF-α in myocardium and hepatic tissue in Group HL was enhanced remarkably, and the tissue damages of Group HL were severest.@*CONCLUSIONS@#The cardiotoxicity and hepatotoxicity caused by compound stress of hyperthermia and LPS is closely related to the expression of TNF-α.


Subject(s)
Animals , Arterial Pressure , Cardiomyopathies , Fever , Metabolism , Immunohistochemistry , Lipopolysaccharides , Liver , Metabolism , Liver Diseases , Male , Myocardium , Metabolism , Random Allocation , Rats , Rats, Wistar , Stress, Physiological , Physiology , Systemic Inflammatory Response Syndrome , Metabolism , Tumor Necrosis Factor-alpha , Metabolism
19.
Article in English | WPRIM | ID: wpr-819748

ABSTRACT

OBJECTIVE@#To test the curative effect of human umbilical cord-derived mesenchymal stem cells on rat acute radioactive enteritis and thus to provide clinical therapeutic basis for radiation sickness.@*METHODS@#Human umbilical cord-derived mesenchymal stem cells were cultivated in vitro and the model of acute radioactive enteritis of rats was established. Then, the umbilical cord mesenchymal stem cells were injected into the rats via tail vein. Visual and histopathological changes of the experimental rats were observed.@*RESULTS@#After the injection, the rats in the prevention group and treatment group had remarkably better survival status than those in the control group. The histological observations revealed that the former also had better intestinal mucosa structure, more regenerative cells and stronger proliferation activity than the latter.@*CONCLUSIONS@#Human umbilical cord-derived mesenchymal stem cells have a definite therapeutic effect on acute radioactive enteritis in rats.


Subject(s)
Animals , Cell- and Tissue-Based Therapy , Cells, Cultured , Disease Models, Animal , Enteritis , Therapeutics , Humans , Male , Mesenchymal Stem Cell Transplantation , Radiation Injuries, Experimental , Therapeutics , Rats , Rats, Wistar , Umbilical Cord , Cell Biology
20.
Chinese Journal of Biotechnology ; (12): 1598-1605, 2011.
Article in Chinese | WPRIM | ID: wpr-304541

ABSTRACT

Heat shock protein gp96 isolated from tumor tissues holds great promise for tumor immunotherapy. However, at present only very limited amount of gp96 protein can be isolated from tumor tissues. Here, we reconstituted the yeast-expressed gp96 (recombinant gp96, rgp96) with B16.F10 melanoma antigens in vitro to prepare new gp96 tumor vaccine on large-scale, and analyzed its induction of specific anti-tumor immunoresponses by ELISPOT, IFN-gamma intracellular staining and cytotoxicity assays. Immunization with rgp96-tumor antigen complexes significantly inhibited B16 tumor growth compared with either rgp96 or tumor antigens alone and led to enhancement of tumor-specific T-cell activities, which was found similar to that of tumor tissue derived gp96. Our results therefore may provide bases for large-scale preparation of the new generation of gp96 tumor vaccines.


Subject(s)
Animals , Cancer Vaccines , Genetics , Allergy and Immunology , Therapeutic Uses , Female , Heat-Shock Proteins , Genetics , Allergy and Immunology , Therapeutic Uses , Melanoma, Experimental , Therapeutics , Mice , Mice, Inbred C57BL , Neoplasm Transplantation , Recombinant Fusion Proteins , Genetics , Allergy and Immunology , Therapeutic Uses , Skin Neoplasms , Therapeutics , Yeasts , Genetics , Metabolism
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