Animals ; Endotoxemia ; metabolism ; pathology ; Female ; Liver ; metabolism ; pathology ; Male ; Mitochondria, Liver ; metabolism ; Rats ; Reactive Oxygen Species ; metabolism

The glycosylation heterogeneity of recombinant human pro-urokinase (pro-UK) was assessed using ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). Firstly, the source of heterogeneity was determined by measuring the M_r of intact protein before and after N-deglycosylation. Glycosylation sites and the proportion of O-glycopeptides then were determined at the peptide level. Finally, the N-glycans were confirmed and quantified using the N-glycan profile. Results show that the structural heterogeneity of pro-UK is mainly caused by glycosylation. All T₁₈ were fucosylated, and 6.4% of S_138/139 was O-glycosylated with two kinds of oligosaccharides with a ratio of 6.0% and 0.4% respectively. All N₃₀₂ positions were N-glycosylated by more than ten types of glycans, among which A2F and A3F accounted for 80% of the total. The assessment of glycosylation heterogeneity of pro-UK will provide a reference for quality standardization.

To develop an oral drug,ITF gene encoding ITF proein,was expressed in a live delivery vehicle lactococcus lactis.First,the ITF gene was cloned into the prokaryotic expressive vector pNICE:sec.Second,the recombinant vector pNICE:sec-ITF was transformated into Lactococcus lactis strain NZ9000 to express ITF protein.Then the recombinant ITF was induced to express and was identified by SDS-PAGE and Western blot.Rabbits are divided into blank control group,preparation group and therapeutic group which are respectively administrated wih PBS and pNICE:sec-ITF Lactococcus lactis.By grades of ulcer test whether administrated pNICE:sec-ITF Lactococcus lactis protects against HCl-induced gastric injury in rabbits.The results were described as follows.The ITF was amplified and cloned in the vector pNICE:sec successfully.The fusion protein(5.9kDa) was expressed in L.lactis by the induction of the nisin.The quantity of expression accounted for 5% of the total bacterial protein.Western bolt analysis confirmed that fusion protein could be recognized specially by Monoclonal Anti-human TFF3 Antibody.Preparation groups and therapeutic groups do good than control group.Prove that administrated pNICE:sec-ITF Lactococcus lactis is biologically active in an HCl-induced rabbit gastric mucosal injury model.

OBJECTIVETo observe the effect of Wenyang Huazhuo Tongluo Recipe (WYHZTLR) on the proportion of T helper 17 cells (Thl7)/regulatory T cells (Treg), and serum levels of IL-17 and IL-10 in peripheral blood of systemic sclerosis (SSc) patients with yang qi insufficiency and turbidity induced collaterals blockage syndrome (YQITICBS).

METHODSTotally 82 SSc patients were randomly assigned to the Western medicine group (as the control group) and the integrated Chinese and Western medicine group (as the treatment group), 41 cases in each group. All patients took methotrexate (MTX) tablet and prednisone tablet. Patients in the treatment group additionally took WYHZTLR. The treatment course for all was six consecutive months. Besides, another 70 healthy volunteers were recruited as a healthy control group (as the healthy group). Percentages of Th17 and Treg in peripheral blood were detected by flow cytometry. Serum levels of IL-17, IL-10, von Willebrand factor (vWF), aminoterminal propeptide of type l procollagen (PIIINP), and cross-linked carboxyterminal telopeptide of type I collagen ( I CTP) were measured by double antibody sandwich ELISA. The correlations between Th17/Treg and levels of vWF, PIIINP, I CTP, skin score, and disease activity index were observed by Pearson correlation analysis.

RESULTSThe percentage of Th17 in peripheral blood, ratios of Th17/Treg, and the serum level of IL-17 were significantly higher, but the percentage of Treg and the serum level of IL-10 were significantly lower in SSc patients, when compared with those of the healthy group (P <0. 05, P <0. 01). Compared with the same group before treatment, the percentage of Thl7, ratios of Thl7/Treg, and levels of IL-17, vWF, and PIIINP all decreased in the two groups after treatment (P <0.05, P <0.01), but the percentage of Treg and the IL-10 level increased (P <0. 05, P <0. 01). Meanwhile,the level of I CTP was higher in the treatment group after treatment (P <0. 05). The improvement of all indices except the percentage of Th17 was more obvious in the treatment group than in the control group (P <0. 05, P <0. 01). The ratio of Th17/Treg was positively correlated with levels of vWF, PIIINP, skin score, and disease activity index before and after treatment respectively (P <0. 01), but with no obvious correlation with the level of I CTP (P >0. 05).

CONCLUSIONWYHZTLR could achieve its therapeutic effect on SSc patients by regulating Th17/Treg imbalance, lowering levels of vWF and PIIINP, and elevating the level of I CTP.

Drugs, Chinese Herbal ; therapeutic use ; Enzyme-Linked Immunosorbent Assay ; Flow Cytometry ; Humans ; Interleukin-10 ; Interleukin-17 ; Methotrexate ; Peptide Fragments ; Procollagen ; Scleroderma, Systemic ; drug therapy ; T-Lymphocytes, Regulatory ; Th17 Cells

Objective To investigate the effect of early insulin therapy on NF-KB pathway and inflammatory cytokine responses in fiver of diabetic rat.Methods NF-KB p65 DNA binding was assayed with ELISA-based assay kit,cytokine gene expressions were quantified with real-time PCR and phosphoenolpyruvate carboxykinase(PEPCK),NF-KB and inhibitor KB(IKBα)protein levels wlere assayed with Westem blot.Results Compared with control,hepatic PEPCK protein level in the untreated diabetic rat increased by 40%.Early insulin and gliclazide treatment normalized PEPCK protein level.The abundance of IKBα protein was significantly decreased and nuclear NF-KB p65 DNA binding activity was incteased in untreated diabetic rats.IKBot protein content increased and NF-KB p65 DNA binding decreased during early intervention treatment.mRNAs encoding IL-1β and TNFα were increased,which were reduced to normal levels after insulin and gliclazide treatment.Conclusions It is suggested that early insulin treatment inhibits NF-KB activity and inflammatory cytokine responses in fiver that are involved in the aniefioration of insulin resistance in diabetic rats.Such results misht be due to indirect antiinflammatory effects of insulin thus relieving glucotoxicity and lipotoxicity in pefipherM tissues.

To study still further the activity of CNTF mutant designed by computer molecular modling,the methods of dissociated cultures of chick dorsal root ganglion、TF-1 prolification and the normal mice'weight loss tests weve used.The results indicated that the mutant protein promoted the survival of dorsal root ganglion、induced TF-1 prolification and made the normal mice lose weight,decrease appetite and reduce fat index.The weight loss effect was dependant with its administration dosage,ED50 was 150.986?g/kg/d.To TF-1,the specific activity reached 2.0?106U/mg against international reference reagent.In a word,CNTF mutant had excel bioactivity.So it provided clues for its development and application.

In order to investigate the biological effects of the VP22?-mE6?/mE7 built-in adjuvant fusion protein vaccine on the tumor associated with HPV-16 infection.HSV-1 VP22? and HPV-16 mE6?/mE7 genes were cloned,and the pET28a-VP22?-mE6?/mE7 recombinat prokaryotic expression vector was constructed.Vector was transformed into Rosetta(DE3)E.coli string and expressed under the induction of IPTG.The re-naturalized protein was then purified via Ni2+ affinity adsorbent column and identified by SDS-PAGE and Western blot.Purified protein was immunized BalB/C and C57BL/6 mice to evaluate the immunogenicity and anti-tumor activity.The expressed recombinant protein formed as inclusion body with a prediction MW about 34kDa and contained approximately 45% of total somatic protein.The VP22?-mE6?/mE7 immunization induced higher titer of specific IgG against HPV,higher level of lymphocyte proliferation and better effect on suppressing HPV16 positive TC-1 tumor growth than the mice immunized with mE6?/mE7 alone.The results showed that the recombined built-in adjuvant vaccine could induce specific cellular immune response in vitro and inhibit the TC-1 tumor proliferation in vivo,that would be a foundation for further studies and developments of inner adjuvant vaccines.

The medical CT scanner is rapidly evolving from the fan-beam mode to the cone-beam geometry mode. In this paper, a new cone-beam pseudo Lambda tomography was proposed based on the Noo's fan beam super-short scan formula and FDK framework. The proposed pseudo-LT algorithm, which avoids the computation of any PI line and any differential operation, has a significant practical implementation, thus leading to the images with quality improvement and reduced artifacts. The results in the simulation studies confirm the observation that the new algorithm can improve the image resolution over the traditional algorithms with noise projection data.
Algorithms ; Artifacts ; Cone-Beam Computed Tomography ; methods ; Humans ; Imaging, Three-Dimensional ; Models, Theoretical ; Phantoms, Imaging ; Radiographic Image Enhancement ; methods

OBJECTIVETo assess the clinical effect and safety of the Chinese medicine Longbishu Capsule combined with mesylate doxazosin in the treatment of benign prostatic hyperplasia (BPH) of the kidney deficiency and blood stagnation type.

METHODSThis was a randomized, double-blind, double-simulation control study. We equally assigned 60 men diagnosed with BPH of the kidney deficiency and blood stagnation type to an experimental and a control group, the former treated with mesylate doxazosin plus Longbishu Capsule and the latter with mesylate doxazosin plus placebo. We compared the International Prostate Symptom Score (IPSS), quality of life (QOL), Chinese symptom score (CSS), maximal urinary flow rate (Qmax), and prostate volume between the two groups of patients before and after 6 months of medication.

RESULTSAfter treatment, there were 5 cured cases, 13 markedly effective cases, 9 effective cases, 1 ineffective case, and 2 eliminated cases in the experimental group, as compared with 2 cured cases, 8 markedly effective cases, 10 effective cases, 7 ineffective cases, and 3 eliminated cases in the control group. The total effectiveness rate was obviously higher in the former (96.4%) than in the latter (74.1%). IPSS, Qmax, and CSS were improved in both of the groups after medication, even more significantly in the experimental than in the control group (IPSS: 15.22 ± 2.98 vs 18.15 ± 5.88, P <0.05; Qmax: [13.56 ± 2.26] ml/s vs [11.78 ± 2.97] ml/s, P <0.05; CSS: 6.18 ± 2.13 vs 9.52 ± 3.15, P <0.05). Because of the difference in the QOL score between the two groups at the baseline (P = 0.038 <0.05), no more comparison was made in this aspect after treatment.

CONCLUSIONThe combination of Longbishu Capsule with mesylate doxazosin is safe and effective for the treatment of BPH.

Adrenergic alpha-Antagonists ; therapeutic use ; Capsules ; Double-Blind Method ; Doxazosin ; therapeutic use ; Drug Therapy, Combination ; Drugs, Chinese Herbal ; therapeutic use ; Humans ; Male ; Prostatic Hyperplasia ; drug therapy ; physiopathology ; Quality of Life ; Treatment Outcome ; Urination

To establish a detection method of oncolytic adenovirus/p53 and standard of quality control, human telomerase reverse transcriptase (hTERT) promoter, CMV fusion promoter containing hypoxia reaction element (HRE) and p53 gene were identified by vector DNA restriction enzyme digestion and PCR analysis. The result conformed that all modified regions were in consistent with theoretical ones. Particle number was 2.0 x 10(11) mL(-1) determined by UV (A260). Infectious titer was 5.0 x 10(10) IU mL(-1) analyzed by TCID50. In vitro p53 gene expression in human lung cancer cell H1299 was determined by ELISA, and A450 ratio of nucleoprotein in virus infection group to control group was 5.2. Antitumor potency was evaluated by cytotoxicity assay using human lung cancer cell A549, and the MOI(IC50) of this gene therapy preparation was 1.0. The tumor cells targeted replication ability of recombinant virus was determined by TCID50 titer ratio of filial generation virus between human lung cancer cell A549 and human diploid epidermal fibrolast BJ cells after infected by virus with same MOI. TCID50 titer ratio of tumor cell infection group to normal cell infection control group was 398. The IE-HPLC purity of virus was 99.5%. There was less than 1 copy of wild type adenovirus within 1 x 10(7) VP recombinant virus. Other quality control items were complied with corresponding requirements in the guidance for human somatic cell therapy and gene therapy and Chinese pharmacopeia volume III. The detection method of oncolytic adenovirus/p53 was successfully established for quality control standard. The study also provided reference for quality control of other oncolytic viral vector products.
Adenoviridae ; genetics ; metabolism ; physiology ; Cell Line, Tumor ; Gene Expression Regulation, Neoplastic ; Genes, p53 ; Genetic Therapy ; Genetic Vectors ; Humans ; Neoplasms ; metabolism ; pathology ; virology ; Oncolytic Viruses ; genetics ; metabolism ; physiology ; Quality Control ; Recombinant Fusion Proteins ; genetics ; metabolism ; Transfection ; Virus Replication