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Objective @#To evaluate the effect of gingival stem cells-derived exosomes (GMSC-Exos) treatment on the expression of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in rats with periodontitis, so as to provide the evidence for periodontitis treatment.@*Methods@#Forty specific pathogen-free (SPF) rats at ages of 8 weeks were randomly divided into 4 groups, including the blank group, periodontitis group, GMSC-Exos group and PBS group. Rats in the periodontitis group, GMSC-Exos group and PBS group were modeled for periodontitis using the ligature method. Rats in the blank group and periodontitis group were given no treatment, while rats in the GMSC-Exos group and PBS group were given 20 μL GMSC-Exos and PBS by injection, respectively. The periodontal index was measured in all rats 4 weeks post-treatment, and the TNF-α and IL-6 levels were measured in rat serum samples using enzyme-linked immunosorbent assay (ELISA). The TNF-α and IL-6 gene expression was quantified using the polymerase chain reaction (PCR) assay in the gingival tissues of the rat left upper maxillary area, and the periodontal tissues in the left upper maxillary areas were sampled for pathological examinations. Periodontal clinical indexes, IL-6 and TNF-α levels were compared in each group.@*Results@#The gingival sulcus bleeding index, gingival index, probing depth, and plaque index in the GMSC-Exos group (1.87±0.41, 1.03±0.19, 1.91±0.09 and 1.11±0.17) were higher than those in the blank group (0.96±0.31, 0.83±0.31, 1.09±0.05 and 1.01±0.38), but lower than those in the periodontitis group (2.65±0.50, 1.36±0.22, 2.61±0.07 and 1.51±0.26) and PBS group (2.44±0.50, 1.23±0.20, 2.49±0.10 and 1.39±0.28) (all P<0.05). The serum IL-6 and TNF-α levels in the GMSC-Exos group [(205.97±11.47) and (90.11±8.57) pg/mL] were higher than those in the blank group [(143.10±4.87) and (80.07±5.13) pg/mL], but lower than those in the periodontitis group [(367.33±13.89) and (158.29±13.10) pg/mL] and PBS group [(364.23±13.62) and (140.60±11.73) pg/mL] (all P<0.05). The IL-6 and TNF-α mRNA expression in the rat gingival tissues in the GMSC-Exos group (1.09±0.14 and 1.61±0.29) was higher than that in the blank group (0.99±0.10 and 1.06±0.14), but lower than that in the periodontitis group (1.63±0.09 and 3.63±0.26) and PBS group (1.58±0.11 and 3.79±0.32) (all P<0.05). Pathological examinations showed alleviation of periodontal tissue destruction, inflammatory cell infiltration and alveolar bone resorption, and no obvious root dental root regeneration in the junctional combined epithelium in the GMSC-Exos group relative to the periodontitis group and the PBS group. @*Conclusion@#Administration of GMSC-Exos may reduce periodontal inflammation and alveolar bone resorption by inhibiting IL-6 and TNF-α expression in rats with periodontitis.
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Objective: To intensively study the chemical constituents from the seed cake of Camellia oleifera and its pharmacological activities,in order to provide scientific basic for its further development and utilization. Method: All kinds of column chromatography and spectral methods were employed to isolate and identify the monomeric compounds from its ethyl acetate portion of ethanol extract. The in vitro anti-inflammatory effects were evaluated by LPS-induced inflammatory model in RAW264.7 macrophages. Result: Eight phenolic acids and two flavonoids were isolated from the ethyl acetate soluble portion and identified as p-hydroxybenzoic acid(1),protocatechuic acid(2),gallic acid(3),methyl gallate(4),ethyl gallate(5),isovanillic acid(6),ethyl 3,4-dihydroxylbenzoate(7),2-(3',4'-dihydroxyphenyl)-1,3-benzodioxole-5-aldehyde(8),quercetin(9),rutoside(10). Among them, compounds 4-8 were first isolated from this plant. These compounds had good anti-inflammatory activities against NO production in LPS-stimulated RAW264.7 macrophages in an obvious dose-dependent manner. Among them, compound 8 showed a strongest activity. Conclusion: The above results show that the phenolic acids and flavonoids from seed cake of C. oleifera have good prospects for the development and application of anti-inflammatory drugs.
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Objective To explore the use of Diagnosis Related Groups(DRGs)evaluation index in performance management system of hospitals.Methods The performance evaluation system was built based on medical business volume index,efficiency indicators,cost control indexes,drug control indexes, medical quality and medical safety indexes,by means of extracting the home page of hospital discharge records from 2009 to 2013 and grouping automatically with the“BJ-DRGs”group-maker.Results The operation evaluation indexes of the hospital have seen great progress since advent of the DRGs evaluation indexes.Conclusion Introduction of DRGs has scored great success in the performance appraisal system of the hospital.
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Objective To evaluate the efifcacy and safety of hemostatics of injected gelatin matrix (HIGM) under the guidance of contrast-enhanced ultrasound (CEUS) for treating splenic trauma in canine model. Methods A total of 24 commercial hybrid dogs underwent celiotomy with creation of uniformly blunt splenic trauma lesion of 4.0 cm×4.0 cm×2.5 cm (length, width and depth, respectively) by hemostatic clamp. Subjects were prospectively randomized into two groups. The treatment group was treated with HIGM under the guidance of CEUS and the positive control group received thrombin solution. Conventional ultrasound and CEUS were performed to record the ascites and the splenic lesion areas at 1st, 3rd, 7th, 14th and 21st day. The ifne needle biopsy and splenectomy were performed for histopathologic examination. The weight, free intraperitoneal lfuid and injury site were compared with t test between HIGM and postive group. Results All animals in two groups survived. All dogs stopped hemorrhage after injection of HIGM under CEUS guidance. The area of injury site was (12.91±0.89) cm2, (4.45±0.75) cm2 and (1.38±0.23) cm2 at 1st, 3rd and 7th day and splenic lesions were not found at 14th and 21st day in all dogs (n=12) of HIGM group. The splenic lesion was (16.74±0.91) cm2, (11.26±0.99) cm2, (8.02±0.82) cm2 and (1.58±0.36) cm2 in the postive group at 1st, 3rd, 7th and 14th day and splenic lesions were not found at 21st day in all dogs (n=12). At 7th and 14th day post-injection, lesion areas were statistically significant between two groups (t=27.162, P=0.008;t=15.129, P=0.001). Free intraperitoneal lfuid was (0.91±0.05) cm at 1st day detected by conventional ultrasound and free intraperitoneal fluid was not found at 3rd, 7th, 14th and 21st day in all dogs (n=12) of HIGM group. The free intraperitoneal fluid in thepositive group was (1.96±0.17) cm, (1.30±0.11) cm and (0.81±0.12) cm at 1st, 3rd and 7th day and free intraperitoneal lfuid was not found at 14th and 21st day in all dogs (n=12). At 1st, 3rd and 7th day post-injection, free intraperatitoneal lfuid was statistically significant between two groups (t=20.934, P=0.003; t=41.310, P=0.000; t=22.520, P=0.000). Histopathological examination showed that there was no foreign body and foreign body granuloma and the structure of red pulp was recovered at 7th, 14th and 21st day. Gross anatomy showed that the splenic injury site was recovered completely without complications. Conclusion This study explored the value of HIGM for splenic trauma and provided a preliminary experimental evidence for clinical treatment.
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<p><b>OBJECTIVE</b>To explore the efficacy of homemade hemostatics of injected gelatin matrix (HIGM) for immediately treating blunt hepatic trauma in canine model without additional pressure.</p><p><b>METHODS</b>A total of 27 commercial hybrid dogs underwent celiotomy to establish hepatic trauma model after general anesthesia. The dogs were prospectively randomized into 3 groups: the treatment group (n=9, with the direct application of homemade hemostat), the positive control group (n=9, with thrombin solution), and the negative control group (n=9, with 0.9% normal saline). Time to hemostasis and intra-abdominal blood loss were recorded, and heart rate (HR), mean arterial pressure (MAP), and hematological parameters were compared among these three groups. Gross examinations were performed 30 minutes after surgery.</p><p><b>RESULTS</b>Significantly shorter time to hemostasis [(1.20±0.33) min] and less blood loss [(47.22±8.61) ml] were observed in the treatment group than in control groups (P 0.05). No cases of bleeding occurred in any animals in the treatment group, and no signs of infection and adhesion formation were evident due to exposure to HIGM. Two cases in the positive control group (22.22%) were found to have rebleeding. All animals in the negative control group experienced visible bleeding.</p><p><b>CONCLUSION</b>HIGM is effective for controlling bleeding after hepatic trauma without the additional compression, and therefore may be valuable in field surgery.</p>
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Animals , Dogs , Disease Models, Animal , Gelatin , Hemostatics , Injections , Liver , Wounds and InjuriesABSTRACT
This study investigated the expression of hemeoxygenase-1 (HO-1) in rats with acute lung rejection and its implication.A valid rat orthotopic left lung transplantation model (SD rat→Wistar rat) was established by using an improved three-cuff anastomosis technique.The rats were divided into control group,CoPP (HO-1 inducer)-treated group and ZnPP (HO-1 inhibi-tor)-treated group.The severity of acute rejection was graded on the basis of the morphologic changes of the lung samples stained with HE.The expression of HO-1 protein in lung tissue was de-tected by using immunohistochemistry and Western blot,and HO-1 mRNA activity was assayed by RT-PCR.The results showed that the expression of HO-1 protein was significantly increased with the acute rejection grading in rats (P<0.01).As compared with control and ZnPP-treated groups,the se-verity of acute rejection was not alleviated and the grade not reduced significantly in CoPP-treated group (P>0.05).It was concluded that HO-1 protein might be involved in the pathological process of post-graft acute rejection.The expression of HO-1 protein was increased gradually with aggravation of acute rejection,and HO-1 protein might be used as an index to monitor acute rejection after lung transplantation.
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<p><b>OBJECTIVE</b>To screen proteins interacting with HCV NS4A protein in leukocytes by yeast-double hybridization.</p><p><b>METHODS</b>The bait plasmid pGBKT7-NS4A was transformed into yeast AH109 was transformed, and the expressing of the fusion protein was identified by SDS-page. The transformed yeast was mated with yeast Y187 containing leukocytes cDNA library plasmid in 2xYPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing x-alpha-gal for selecting two times and screening. After extracting and sequencing of plasmid from blue colonies, analysis was conducted by bioinformatics. And, the gene encoding the interesting protein was cloned, and back-cross was performed.</p><p><b>RESULTS</b>Forty-five colonies were sequenced, among them, 29 colonies were human calcium modulating cyclophilin ligand (CAML). The gene encoding CAML was cloned, and the interaction between NS4A and CAML was ensured.</p><p><b>CONCLUSION</b>Seven kinds of proteins interacting with NS4A in leukocytes were successfully screened and the results brought some new clues for studying the pathogenesis of HCV.</p>
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Humans , Adaptor Proteins, Signal Transducing , Genetics , Metabolism , Carrier Proteins , Genetics , Metabolism , Cloning, Molecular , Gene Library , Leukocytes , Cell Biology , Metabolism , Protein Binding , Transformation, Genetic , Two-Hybrid System Techniques , Viral Nonstructural Proteins , Viral Proteins , Genetics , MetabolismABSTRACT
A lipase from Aspergillus niger F044 was purified to homogeneity using ammonium sulfate precipitation, dialysis, DEAE-Sepharose Fast Flow anion exchange chromatography and Sephadex G-75 gel filtration chromatography. This purification protocol resulted in a 73.71-fold purification of lipase with 33.99% final yield, and the relative molecular weight of the enzyme was determined to be approximately 35-40kD using SDS-PAGE. The optimum pH and temperature for lipolytic activity of the lipase was 7.0 and 45 degrees C , respectively. It was extremely stable at 60 degrees C and retained 98.70% of its original activity for 30min. The stability declined rapidly as soon as the temperature rose over 65 degrees C . The lipase was highly stable in the pH range from 2.0 to 9.0 for 4h. Ca2+ and Mg2+ ions stimulated lipolytic activity, whereas Mn2+ , Fe2+ and Zn2+ ions caused inhibition. The values of Km and Vmax calculated from the Lineweaver-Burk plot using pNPP as hydrolysis substrate were 7.37mmol/L and 25.91 micromol/(min x mg), respectively. The N-terminal sequence of the lipase was Ser/Glu/His-Val-Ser-Thr-Ser-Thr-Leu-Asp-Glu-Leu-Gln-Leu-Phe-Ala-Gln, which is highly homogeneity with that of lipase, as reported by Torossian.
Subject(s)
Amino Acid Sequence , Aspergillus niger , Chromatography, Gel , Chromatography, Ion Exchange , Electrophoresis, Polyacrylamide Gel , Enzyme Stability , Fungal Proteins , Chemistry , Metabolism , Hydrogen-Ion Concentration , Lipase , Chemistry , Metabolism , Molecular Sequence Data , Molecular Weight , Sequence Analysis, Protein , TemperatureABSTRACT
The fermentation conditions of alkaline lipase producing by Pseudomonas cepacia PCL-3 were optimized.Based on the analysis of single factorial experiments,dextrin was the most suitable carbon source,peptone and urea were the suitable compound nitrogen sources among the examined materials.Three significant factors(urea,inoculum and initial pH) were selected from the eight factors related to lipase production by Plaekett-Burman method,and were further optimized with response surface analysis.And then,steepest ascent procedures were applied to define the optimal response region of the three factors.The obtained optimal conditions were urea 0.15%,inoculum 3.05% and initial pH 8.38,under which conditions,the enzyme activity was improved from 25.37 U/ml to 48.88 U/ml,enhanced 1.93 folds.Starting from the flask conditions,the highest lipase activity of 47.69U/ml was achieved by batch fermentation in a 10 L fermentor after 52 h of the cultivation.
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Bioimprinting is a new developed technique to improve the characteristics of enzymes.Bioimprinting by lauric acid was conducted to improve the esterification activity of lipase PS in sol-gel immobilization process with methyltrimethoxysila(MTMS) and tetramethoxysila(TMOS) as the precursors.Results generated by checking the esterification activity and scanning electron microscope showed that bioimprinting can enhance the specific activity and thermal stability of lipase PS.The bioimprinting system was optimized by orthogonal experiment,and the optimal condition for lipase bioimprinting is water/silane molar ration(R) 12,polyethylene glycol(PEG) 120?l,and lauric acid 0.15 mmol.Compared with the free enzyme and the non-imprinted enzymes,the specific activity of imprinted enzymes has been improved 44.3 fold and 2.4 fold,respectively.Imprinted lipase show better thermal stability,and the relative activity is 58% after incubated in 80 ℃ for 0.5 h,while no activity was detected for the free enzyme.
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The fermentation conditions of lipase production by Geotrichum candidum Y162 were optimized. Initially, the most suitable carbon olive oil, nitrogen source soybean flour and NH4Cl, salt BaCl2 and MgCl2 were selected according to single factorial experiments respectively. Based on the result, screening methodology Plackett-Burman design was used to evaluate the effects of twelve factors related to lipase production and three statistically significant factors olive oil, BaCl2 and NH4Cl were selected. The path of steepest ascent was used to approach the optimal region of lipase production subsequently. Then, the optimal combined concentration for maximum enzyme activity were further optimized by response surface methodology and determined as follows: olive oil 2.35%, BaCl2 0.36%,and NH4Cl 4.69%.The optimization of culture conditions of G.candidum Y162 led to a 2.25-fold increase in lipase production relative to initial result 14.16 U/ml, which indicate that single factor in combination with response surface methodology is an effective method for optimization of lipase production conditions by G.candidum Y162.
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From the N-terminal amino acid sequence of the Aspergillus niger F044 lipase,a potential homologous gene A84689 to the anl(the gene encoding the Aspergillus niger lipase)was found by means of bioinformatics.Based on the nucleotide sequence of the A84689,primers were designed to amplify anl.Nucleotide sequencing of the genomic anl gene revealed an open reading frame of 1 044 nucleotides,containing three introns(54,45 and 51 nucleotides).The deduced amino acid sequence of the anl gene corresponds to 297 amino acid residues including a signal sequence of 27 amino acid residues.The cloned cDNA coding for mature Anl(the protein of the Aspergillus niger lipase)was overexpressed in Escherichia coli BL21(De3),and the recombinant Anl was purified.The denatured recombinant Anl by 8mol/L urea was refolded in vitro by dilution and DEAE Sepharose Fast Flow chromatography.