Objective:To investigate the levels of diphtheria-specific binding antibodies and neutralizing antibodies in mice immunized with pneumococcal polysaccharide conjugate vaccine using diphtheria toxoid as a carrier.Methods:NIH mice were immunized with one batch of diphtheria, tetanus and acellular pertussis combined vaccine, absorbed (DTaP-1) or three different batches of 13-valent pneumococcal polysaccharide conjugate vaccine (PCV13, containing diphtheria toxoid vector) at three dilutions (5-, 10- and 20-fold dilution). Serum samples were collected to test for diphtheria-specific antibody titers and diphtheria potencies of the vaccines. Another three batches of DTaP vaccine and three batches of tetanus toxoid, reduced diphtheria toxoid and acellular pertussis combined vaccine (Tdap) were used to immunize NIH mice. Serum samples were collected and the diphtheria potencies were detected. Statistical analysis was performed using one-way analysis of variance.Results:At the 5-fold and 10-fold dilutions, the titers of diphtheria-specific antibodies induced by three batches of PCV13 vaccine were all lower than those by DTaP-1 vaccine (all P<0.001), while there was no statistically significant difference at the 20-fold dilution ( P>0.05). The diphtheria potencies of the DTaP-1 vaccine and the three batches of PCV13 vaccine were 100.5, 76.2, 64.5, and 62.0 IU/ml, respectively. The diphtheria potencies of another three batches of DTaP vaccine were 82.5, 83.6, and 79.9 IU/ml, respectively, and those of three batches of Tdap vaccine were 10.3, 12.2, and 12.9 IU/ml, respectively. There was no statistically significant difference in diphtheria potency between DTaP vaccine and PCV13 vaccine( P>0.05), while there was a statistically significant difference between Tdap vaccine and the PCV13 vaccine ( P<0.001). Conclusions:The pneumococcal polysaccharide conjugate vaccine with diphtheria toxoid has good diphtheria immunogenicity and can induce the production of higher levels of diphtheria-specific binding antibodies and protective neutralizing antibodies in vivo. Pneumococcal capsular polysaccharide exerts an immune enhancement effect on diphtheria toxoid. The relevant results provide valuable guidance for determining carrier protein dosage in bacterial polysaccharide conjugate vaccines, planning vaccine co-administration, and selecting the dosage of diphtheria toxoid antigen in the research and development of combined vaccines.

To the present study aimed to investigate the protective effects of Erlong Zuoci Pills on oxidative stress induced by hydrogen peroxide (H2O2) in House Ear Institute-Organ of Corti 1 (HEI-OC1) and to explore the mechanism by cellular metabolomics. There were 6 groups in the experiment: the control group, model group, three dose groups of ELZC (low, medium, and high), and positive control ascorbic acid group. The oxidative stress injury model was established in the HEI-OC1 by inducing 0.9 mmol/L H2O2 for 12 h. The proliferation of HEI-OC1 cells was observed by CCK-8 assay; the contents and activity of lactate hydrogenase (LDH), reactive oxygen species (ROS), and superoxide dismutase (SOD) in HEI-OC1 cells were detected by corresponding kits. Finally, the endogenous substances of cells were analyzed from the perspective of metabolomics. Compared with the model group, ELZC groups could significantly increase the cell proliferation rate after administration. Moreover, they could also ameliorate the increase of ROS and LDH content and the decrease of antioxidant enzyme SOD caused by H2O2. Metabolomic results revealed significant differences among multiple groups in the scores of partial least squares discriminant analysis. The ELZC group could relocate the model group back to the control group. The metabolic regulation of ELZC on oxidative stress in HEI-OC1 cells mainly affects nucleotide metabolism and amino acid metabolism. In summary, the results indicate that ELZC exhibits protective effects on H2O2-induced oxidative stress in HEI-OC1 cells. Additionally, this protective effect may be produced by increasing the content of amino acids such as uridine and phenylalanine, thereby regulating pathways such as pyrimidine metabolism, phenylalanine metabolism, biosynthesis of phenylalanine, tyrosine, and tryptophan, and histidine metabolism.

Objective:To investigate the levels of diphtheria-specific binding antibodies and neutralizing antibodies in mice immunized with pneumococcal polysaccharide conjugate vaccine using diphtheria toxoid as a carrier.Methods:NIH mice were immunized with one batch of diphtheria, tetanus and acellular pertussis combined vaccine, absorbed (DTaP-1) or three different batches of 13-valent pneumococcal polysaccharide conjugate vaccine (PCV13, containing diphtheria toxoid vector) at three dilutions (5-, 10- and 20-fold dilution). Serum samples were collected to test for diphtheria-specific antibody titers and diphtheria potencies of the vaccines. Another three batches of DTaP vaccine and three batches of tetanus toxoid, reduced diphtheria toxoid and acellular pertussis combined vaccine (Tdap) were used to immunize NIH mice. Serum samples were collected and the diphtheria potencies were detected. Statistical analysis was performed using one-way analysis of variance.Results:At the 5-fold and 10-fold dilutions, the titers of diphtheria-specific antibodies induced by three batches of PCV13 vaccine were all lower than those by DTaP-1 vaccine (all P<0.001), while there was no statistically significant difference at the 20-fold dilution ( P>0.05). The diphtheria potencies of the DTaP-1 vaccine and the three batches of PCV13 vaccine were 100.5, 76.2, 64.5, and 62.0 IU/ml, respectively. The diphtheria potencies of another three batches of DTaP vaccine were 82.5, 83.6, and 79.9 IU/ml, respectively, and those of three batches of Tdap vaccine were 10.3, 12.2, and 12.9 IU/ml, respectively. There was no statistically significant difference in diphtheria potency between DTaP vaccine and PCV13 vaccine( P>0.05), while there was a statistically significant difference between Tdap vaccine and the PCV13 vaccine ( P<0.001). Conclusions:The pneumococcal polysaccharide conjugate vaccine with diphtheria toxoid has good diphtheria immunogenicity and can induce the production of higher levels of diphtheria-specific binding antibodies and protective neutralizing antibodies in vivo. Pneumococcal capsular polysaccharide exerts an immune enhancement effect on diphtheria toxoid. The relevant results provide valuable guidance for determining carrier protein dosage in bacterial polysaccharide conjugate vaccines, planning vaccine co-administration, and selecting the dosage of diphtheria toxoid antigen in the research and development of combined vaccines.

Objective: To assess the effect of viral macrophage inflammatory protein (vMIP) -Ⅱ on the expression of apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G (APOBEC3G) , and to explore the mechanisms. Methods: A recombinant plasmid pEGFP-N3-K4 (vMIP-Ⅱ plasmid group) and an empty plasmid pEGFP-N3 (empty plasmid group) were separately transfected into 293T cells, and quantitative PCR and Western blot analysis were performed to evaluate the effect of transfection with vMIP-Ⅱ gene on the APOBEC3G expression in 293T cells. Some 293T cells in the empty plasmid group and vMIP-Ⅱ plasmid group were treated with 1 000 IU/ml interferon (IFN) -α for 36 hours, and then Western blot analysis was conducted to determine the APOBEC3G expression in the empty plasmid group and vMIP-Ⅱ plasmid group with or without IFN-α treatment. Some 293T cells transfected with vMIP-Ⅱ plasmids were treated with 75 μmol/L AG490 (a JAK/STAT signaling pathway inhibitor) and 20 μmol/L U0126 (an ERK signaling pathway inhibitor) separately; after 24 hours, total protein was extracted from 293T cells, and Western blot analysis was conducted to determine the expression of APOBEC3G. A recombinant plasmid containing APOBEC3G promoter was constructed by using a luciferase reporter gene, and the promoter fragment included the full-length promoter sequence (POS) of APOBEC3G, sequences with the lengths of 1 560, 960, 720, 480, 420, 360, 330 and 240 bp, and the regulatory element-free region (NEG) of APOBEC3G, separately. Some 293T cells were co-transfected with the recombinant plasmid carrying luciferase reporter gene and vMIP-Ⅱ plasmid (experimental group), or the recombinant plasmid and empty plasmid (control group). Subsequently, the activity of the APOBEC3G promoter was evaluated, and the key promoter region through which the transcriptional activity of APOBEC3G was regulated by vMIP-Ⅱ was analyzed. Statistical analysis was carried out by using t test, one-way analysis of variance and least significant difference (LSD) -t test. Results: The mRNA and protein expression of APOBEC3G was significantly higher in the vMIP-Ⅱ plasmid group (2.500 ± 0.013, 1.472 ± 0.013 respectively) than in the control group (1, 0.364 ± 0.030 respectively; t = 6.22, 6.54 respectively, both P < 0.05) . The APOBEC3G expression significantly differed among the empty plasmid group, vMIP-Ⅱ plasmid group, empty plasmid + IFN-α group and vMIP-Ⅱ plasmid + IFN-α group (1, 2.030 ± 0.108, 2.700 ± 0.081 and 2.600 ± 0.099 respectively; F = 67.026, P < 0.001) , but there was no significant difference between the vMIP-Ⅱ plasmid group and empty plasmid + IFN-α group (t = 3.46, P > 0.05) . The APOBEC3G expression also significantly differed among the vMIP-Ⅱ plasmid group, vMIP-Ⅱ plasmid + AG490 group and vMIP-Ⅱ plasmid + U0126 group (0.617 ± 0.025, 0.179 ± 0.061, 0.359 ± 0.012 respectively; F = 70.019, P < 0.001) , and was significantly lower in the vMIP-Ⅱ plasmid + AG490 group and vMIP-Ⅱ plasmid + U0126 group than in the vMIP-Ⅱ plasmid group (t = 9.66, 11.836 respectively, both P < 0.01) . Luciferase activity assay showed that the promoter activity significantly differed among the vMIP-Ⅱ plasmid groups transfected with POS, 1 560-, 960-, 720-, 480-, 420-, 360-, 330-, 240-bp or NEG sequences (F = 81.092, P < 0.001) , and the APOBEC3G promoter activity decreased greatly from the 720-bp group to 480-bp group. Conclusion vMIP-Ⅱ upregulates the expression of APOBEC3G, likely through the JAK/STAT signaling pathway or the key promoter region regulating the transcriptional activity of APOBEC3G.

The ablated aerosols of biological matrix sample were studied using 213 nm nanosecond laser ablation system.The stable signal intensity and high sensitivity were obtained when the laser energy was 25%, the spot size was 200 μm, the scan rate was 20 μm/s, the frequency was 20 Hz and the carrier gas was 700 mL He + 700 mL Ar.Relative fractionation index of 56 elements were investigated and 31P as the internal standard element was selected under the optimized laser ablation conditions.The results showed that particle size of the biological sample was 3 μm, which was larger compared with NIST 610 sample.Element fractionation in biological sample was smaller than in glass sample, and relative fractionation index of most elements attained 1.0 ± 0.1.Element fractionation mechanism of biological sample was discussed.The possible reason why the relative fractionation index in biological sample with large particle size did not significantly increase compared to the glass sample is that the 3-μm particles entered into ICP can be atomized.On the other hand, enrichment effect for large ablation particles was relatively small.Further study of the influence factors of fractionation effect indicated that, the fractionation effect had relations with laser ablation energy, laser frequency and scan rate, negatively relation with the oxide boiling point, and positively relation with oxide bond energy and ionization energy.

TheLutetium-YttriumOrthosilicate(LYSO)isakindofscintillatingcrystalmaterialwiththebest comprehensive properties. In this work, the trace elements Ca, Fe, K, Li, Mg and Na in LYSO were determined by solution-cathode glow discharge-atomic emission spectrometry ( SCGS-AES ) . The optimal conditions included 0. 1 mol/L HNO3 sample solutions and operation at a voltage at 1080 V with a flow rate of 2. 0 mL/min. The LYSO matrix concentration tolerance of the SCGD source was determined to be 10 g/L. Sample solutions dissolving from several LYSO samples with HF, HNO3 and HClO4 were examined by SCGD-AES. For LYSO samples, the values obtained by SCGD-AES agree well with those obtained by axial inductively coupled plasma-atomic emission spectroscopy ( ICP-AES ) and inductively coupled plasma-mass spectroscopy (ICP-MS). The emissions of Lu and Y were not observed, so that the determination of trace elements in LYSO matrices could be conducted with little interference. The detection limits of Ca, Fe, K, Li, Mg and Na in LYSO were 1. 0, 3. 0, 0. 02, 0. 01, 0. 02 and 1. 0 mg/kg, respectively.

To design and manufacture a new bone matrix material (NBM) composed of both organic and inorganic materials for bone tissue engineering, the osteogenic potential of NBM combined with osteoblasts was studied in vitro and in vivo. Osteoblasts from bone marrow stromal cells of New Zealand rabbit were cultured with NBM in vitro , then the materials combined with osteoblasts were implanted into the skeletal muscles of rabbits. The osteogenic potential of NBM was evaluated using contrast microscope, scan electromicroscope and histological examination. Osteoblasts could attach and proliferate well in the NBM, secrecting lots of extracellular matrix, while in vivo experiment of the NBM in osteoblasts group showed that a large number of lymphacytes and phagocytes invaded into the inner of the material in the rabbit skeletal muscle beds after 4 weeks of implantation, no new bone formation was observed after 8 weeks. The different osteogenic potential expressed by NBM between in vitro and in vivo may be due to the immunogenity of NBM which causes cellular immunoreaction to destroy the osteogenic environment. More attention should be paid to the immunoreaction problem in tissue engineering between the host and organic inorganic composite materials.