1.Preventive effect of carboxymethyl chitosan solution on postoperative peritoneal adhesions in a rat model
Juanjuan CUI ; Zhanqin FENG ; Shouqiang ZHANG ; Zengjuan ZHENG ; Weifen ZHANG
Chinese Journal of Tissue Engineering Research 2014;(8):1250-1256
BACKGROUND:The patients are suffering from peritoneal adhesions that are caused after abdominal operation. As so far, there is stil no effective drug or method that can completely prevent peritoneal adhesions. Carboxymethyl chitosan is a biocompatible and biodegradable biomedical material with anti-adhesion effects, which is an ideal biomaterial for prevention of peritoneal adhesion theoreticaly.
OBJECTIVE:To investigate the novel anti-adhesion properties of carboxymethyl chitosan anti-adhesion solution on the prevention of postsurgical adhesion in vivo in a rat model.
METHODS:Fifty-six adult male Wistar rats were randomly divided into four groups: 0.9% normal saline solution (group A), hyaluronic acid gels (group B), medical chitosan gels (group C) and carboxymethyl chitosan anti-adhesion solution (group D). The model of postoperative intestinal adhesion was established by making cecal scratches/abdominal wal defects. Al the rats were scarified after 2 or 3 weeks. Whole blood was colected by cardio-puncture, lung tissue and tissue adhesion were stripped. The incidence and degree of adhesions, histological effects, expression of transforming growth factor-β1 (TGF-β1), the amounts of hydroxyproline and white blood cels were observed.
RESULTS AND CONCLUSION:The formation of postsurgical adhesions in groups B, C and D was significantly decreased, which was lighter than that of group A (P < 0.05). Furthermore, the adhesion formation in group D was significantly decreased in comparison with group A (P < 0.01). At the same time, the levels of transforming growth factor-β1, hydroxyproline and white blood cels in group D were lighter than those of group A (P < 0.05), and the histopathological results indicated that a marked reduction in inflammatory cels and fibroblasts. Carboxymethyl chitosan anti-adhesion solution can effectively reduce the degree and incidence of postoperative adhesion, and it is becoming a promising drug delivery system in the context of postsurgical anti-adhesion.
2.Preparation and characterization of proanthocyanidins/bletilla striata polysaccharide/chitosan microsphere
Dawei WANG ; Baofang WEN ; Hongcai SHANG ; Zhanqin FENG ; Weifen ZHANG
Chinese Journal of Biochemical Pharmaceutics 2015;37(7):133-137
Objective To prepare proanthocyanidins/bletilla striata polysaccharide/chitosan microspheres ( PC/BSP/CTS ) and the physic-chemical characterizations were investigated.Methods The PC/BSP/CTS microspheres were prepared by spray drying method.The morphology of PC/BSP/CTS microspheres was observed by Scanning Electron Microscopy(SEM), and its physic-chemical characteristics such as diameters, release in vitro, moisture content, swelling ratio, solid density were studied.ResuIts The PC/BSP/CTS microspheres were successfully prepared by spray drying method, SEM showed that PC/BSP/CTS microspheres had the spherical shape with smooth surfaces.The diameters of microsphere A, B and C were 10~20, 2~15, 10~25μm.The in-vitro release showed that the cumulative release of three kinds of microspheres A, B, C was 25.07 %, 38.83 %and 60.00 % in 24 h, which had no burst release, while with time prolonged to 48 h, the cumulative release was 28.89%, 43.17% and 72.86%, respectively.The results of moisture content, swelling ratio, solid density were 15.35% ~23.51%, 46.50% ~105.80%, 0.375 ~0.496. ConcIusion The PC/BSP/CTS microspheres are successfully prepared by spray drying method which possess good characteristics and sustained-release effect, which would be as a good pulmonary drug delivery system.
3.Experimental study of astragaluspolysaccharides on the prevention of radio-pulmonary lesion in Rats
Haijing ZHANG ; Zhaoqiu HU ; Yuqing SUN ; Zhanqin FENG ; Hongcai SHANG ; Weifen ZHANG
Chinese Journal of Biochemical Pharmaceutics 2015;(9):8-11
Objective To investigate the effects of astragalus polysaccharides on radio-pulmonary lesion.Methods 48 Wistar rats were divided into normal group, model group, astragalus polysaccharides treated group and dexamethasone treated group, each had 12 rats.Models of radio-pulmonary lesion of rats in later three groups were established by whole-thorax irradiation 20 Gy, then normal group was pretending to irradiation.Rats in normal group and model group were given 0.9% NaCl, astragalus polysaccharides treated group and dexamethasone treated group were given astragalus polysaccharides(8mg/mL) and dexamethasone(0.05mg/mL), accordingly.All rats were with 8 weeks, at 2 w and 8 w post-injury, 6 rats in each group were selected randomly and anesthesia to death.The erythrocytes and leukocytes in blood and bronchoalveolar lavage fluid ( BALF) in each group were detected and histological examination of lung tissues were performed by Haematoxylin and Eosin staining to study general morphology.Results Radiation-induced lung injury rat model were successfully constructed, general observation showed that irradiated rats gradually appear listlessness, reduce the volume of activities, arched, hair removal and other symptoms, astragalus polysaccharide (APS) and dexamethasone treated rats had gradual improvement than model group.Compared with control group, erythrocytes and leukocytes counts in serum of model group at 2 w and 8 w post-injury were significantly decreased(P <0.05), the numbers of erythrocytes and leukocytes in BALF were significantly increased(P <0.05). Compared with model group group, the leukocytes, erythrocytes and neutrophils counts in serum of astragalus polysaccharides treated group and dexamethasone treated group were all increased(P <0.05), respectively, and the number of leukocytes and erythrocytes in BLAF were significantly decreased(P<0.05).All the indexes between two groups had no significant difference.The pathological changes of lung tissues showed that rats in model group had rupture alveolar wall, widened alveolar interval, pulmonary interstitial hyperplasia, and the alveolar space and interstitial lung stroma had a large number of inflammatory cells exudation.The symptoms in astragalus polysaccharides treated group and dexamethasone treated group were all alleviated, and there was no significant difference between the two groups.Conclusion Astragalus polysaccharides have therapeutic effect on radio-pulmonary lesion in rats, and the therapeutic effects of astragalus polysaccharides are roughly similar to dexamethasone.Further research is needed to elucidate the mechanism behind the effects of astragalus polysaccharides in order to develop appropriate treatment.
4.Construction of Salmonella Pullorum C79-13ΔcrpΔasd mutant balanced-lethal host-vector system and its biological features
Songbiao CHEN ; Jing LI ; Ke SHANG ; Chuan YU ; Chunjie ZHANG ; Xiangchao CHENG ; Yinju LI ; Zhanqin ZHAO
Chinese Journal of Immunology 2014;(8):1083-1087,1092
In order to develop a safer vaccine strain exploit Salmonella Pullorum vaccine strain as a live vaccine vector.Methods:AΔcrpΔasd mutant of S.pullorum C79-13 strain was constructed and it was developed E.coli donor strain mutant was generated through the two-step method introduced by χ7213 ( pREΔasd) was conjugated with the recipient of C 79-13Δcrp.The C79-13ΔcrpΔasd the transduction of recombinant suicide plasmid .Results:PCR and sequencing results showed that ΔsipBSL1344 was suc-cessfully constructed.The further study indicated that foreign DAP must be supplied for the ΔcrpΔasd mutant to grow,in addition,the asd gene was transmitted stably .Compared with C79-13 strain,the O antigens was identical to C79-13 strain,but the growth velocity was reduced significantly ,significantly reduced virulence .Conclusion: The ΔcrpΔasd mutant can accept asd+plasmid to construct host-vector balance lethal system , and this system can be used to express foreign gene efficiently and to develop potential oral multivalent vaccines.
5.Effect of curcumin on learning-memory ability and expression of HMGB1 and JNK in rat model of Alzheimer disease
Lisha YE ; Yuan HAN ; Qixing LIU ; Zhanqin ZHANG ; Hongxia MEI ; Hong CAO ; Qingquan LIAN ; Jun LI
Chinese Journal of Pathophysiology 2014;(6):1114-1118
AIM:To evaluate the effect of curcumin on impaired learning-memory ability and the expression of high mobility group box protein 1 ( HMGB1 ) and c-Jun N-terminal kinase ( JNK ) in a rat model of Alzheimer disease (AD).METHODS:Male Sprague-Dawley rats, weighing 250~270 g, were randomly divided into 4 groups (n=9):blank control group (group A), model group (group B), curcumin treatment group (group C, curcumin injected intraper-itoneally at 100 mg· kg-1· d-1 for 6 consecutive days) and solvent control group (group D).The rats of AD model were induced by injection of ibotenic acid into the nucleus basalis of Meynert ( NBM) bilaterally.All rats were trained in Morris maze to assess the ability of learning and memory .The expression of HMGB1 and JNK in the hippocampus was detected by the methods of immunohistochemistry and Western blotting .RESULTS:Compared with group A , the average escape laten-cy (AEL) in groups B and D were obviously longer (P<0.05), while AEL in group C in the 5th and 6th days were signif-icantly shorter (P<0.05).The releases of HMGB1 in the CA1 and CA3 areas in groups B and D from the nucleus were a-bundant.Compared with groups B and D , HMGB1 in hippocampal CA1 and CA3 areas in group C secreted out of the nu-cleus decreased obviously (P<0.05).No significant difference of the release of HMGB1 between group A and group C was observed (P>0.05).No significant difference in the expression of HMGB1 in the hippocampus among the 4 groups was found (P>0.05).However, compared with groups B and D , the expression of JNK in group C was decreased obvi-ously (P<0.05).CONCLUSION: Curcumin significantly improves the learning and memory ability of AD rats .The probable mechanisms may be related to inhibiting the release of HMGB 1 from the nucleus of hippocampal neurons and de-creasing the expression of JNK in the hippocampus .
6.Construction and characterization of host-vector balanced lethal system of attenuated salmonella choleraesuisΔcrpΔcyaΔasdC78-1 (pYA3493)
Ke SHANG ; Junfeng ZHANG ; Xiangchao CHENG ; Chunjie ZHANG ; Yinju LI ; Guihua CHEN ; Yunfei YAN ; Chongkai ZHAI ; Zhanqin ZHAO
Chinese Journal of Immunology 2015;(3):358-363
Objective:In order to develop an oral live vaccine vector of swine that can stably carry exogenous genes.Methods:Mutant ΔcrpΔcyaΔasdC78-1 was constructed by the method of suicide plasmid pREasd-mediated bacteria homologous recombination on the basis of attenuated Salmonella choleraesuisΔcrpΔcyaC78-1.Complementary plasmid pYA3493 with asd was electrotransformed into the mutant,and thenΔcrpΔcyaΔasdC78-1(pYA3493) host-vector balanced lethal system was constructed.Its biological characteristics were analyzed further.Results:The results of PCR and sequencing showed thatΔcrpΔcyaΔasdC78-1(pYA3493) was constructed suc-cessfully.Biological characteristics showed that the serotype of ΔcrpΔcyaΔasdC78-1(pYA3493) was identical to ΔcyaΔasdC78-1 and vaccine strain C500 and it can stably carry theΔasd gene in vitro;its growth speed was a little slower than ΔcrpΔcyaC78-1 strain,but both of their growth speeds were significantly slower than vaccine strain C500;the biochemical characteristics of ΔcrpΔcyaΔasdC78-1 ( pYA3493 ) were basically the same with ΔcrpΔcyaC78-1 strain.Oral virulence test in mice showed that the virulence ofΔcrpΔcyaΔasdC78-1 ( pYA3493 ) was similar with ΔcrpΔcyaC78-1, but its median lethal dose is 412 times of vaccine strain C500.Conclusion:These results demonstrated that attenuated Salmonella choleraesuisΔcrpΔcyaΔasdC78-1(pYA3493) strain had the potential to be used as an oral live vaccine vector for expressing foreign genes efficiently.
7.Construction and characterization of invasion protein B gene deleted mutant asd host-vector balanced lethal system
Chunjie ZHANG ; Songbiao CHEN ; Xiangchao CHENG ; Chengshui LIAO ; Jing LI ; Lei HE ; Mingliang ZHANG ; Chuan YU ; Zuhua YU ; Yanyan JIA ; Zhanqin ZHAO
Chinese Journal of Immunology 2016;(2):210-213
Objective: To develop an oral live vaccine vector which stably carries exogenous genes.Methods:SL1344ΔsipBΔasd host-vector balanced lethal system was constructed by the method of recombinant suicide plasmid-mediated allelic exchange on the basis of attenuated Salmonella typhinurium SL1344ΔsipB.Then,the biological characteristics of SL1344ΔsipBΔasd was analyzed.Results:The results showed that the mutant was stabile with the Δasd gene in vitro;the serotype and growth rate of SL1344ΔsipBΔasd strain was almost same as the parent SL1344ΔsipB and SL1344 strain.And the mutant strains remain swim ming zones.Virulence test in mice showed that the virulence of SL1344ΔsipBΔasd which carried complementary plasmid pYA3493 by electro-transformation decreased by 1.4%compared with SL1344.Conclusion: These results showed that the SL1344ΔsipBΔasd mutant was successfully constructed.It is likely that this mutant should be used as a live vector to express foreign genes.
8.Influence of fusion protein of IBDV VP2 and chicken interleukin-2 on immune response in chicken.
Chen WANG ; Zhanqin ZHAO ; Chunjie ZHANG ; Yichen LIU ; Ke DING ; Yinju LI ; Xiangchao CHENG ; Puyan CHEN
Chinese Journal of Biotechnology 2010;26(4):476-482
In order to research immunogenicity of the recombinant rVP2-IL-2 fusion protein, we obtained the rVP2-IL-2 fusion protein using Pichia pastoris expression system, and then evaluated its potential to induce immune responses in chicken. The effect was determined in the form of protective anti-IBDV VP2 titers, antibodies (IgG1 and IgG2a), lymphocyte proliferation, the levels of interferon-gamma and interleukin-4 cytokines, and challenge experiment. Antibody titers and proliferation lymphocyte level suggested that the fusion protein could elicit specific humoral immune and cellular immune responses, antibody sub-type results indicated that the rVP2-IL-2 fusion protein induced secretion both of IgG1 and IgG2a. The seem result elicited from cytokines ELISA test, secretion of both of Th1 (gamma-IFN) and Th2 (IL-4) were induced by the rVP2-IL-2 fusion protein. Challenge experiment result shown that chicken immunized the rVP2-IL-2 fusion protein obtained 85% protection. These results confirm that the fusion protein enhances the protection against IBDV through both humoral and cell-mediated immunity, and thus could serve as a candidate for the development of IBDV subunit vaccine.
Animals
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Antibodies, Viral
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biosynthesis
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blood
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Chickens
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immunology
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Immunoglobulin G
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blood
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Interleukin-2
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biosynthesis
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genetics
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Pichia
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genetics
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metabolism
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Recombinant Fusion Proteins
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biosynthesis
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genetics
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immunology
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Th1 Cells
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immunology
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Th2 Cells
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immunology
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Vaccines, Subunit
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immunology
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Viral Structural Proteins
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biosynthesis
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genetics
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Viral Vaccines
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immunology
9.Construction and characterization of invasion protein B gene deleted mutant of Salmonella typhimurium SL1344 strain
Songbiao CHEN ; Jing LI ; Chuan YU ; Lei HE ; Chunjie ZHANG ; Xiangchao CHENG ; Yinju LI ; Yunfei YAN ; Xiuzhe JIN ; Guihua CHEN ; Zhanqin ZHAO
Chinese Journal of Immunology 2015;(2):215-220
Objective:In order to develop a safer vaccine strain exploit Salmonella typhimurium vaccine strain .A ΔsipB mutant of Salmonella typhimurium SL 1344 strain was constructed.Methods: Firstly, the recombinant suicide plasmid containing the missing 585 bp sipB ( PREΔsipB ) was built by homologous recombination , and screenned by two-step method.Results: PCR and sequencing results showed that SL 1344ΔsipB was successfully constructed.It was no significant changes compared with SL 1344.But compared with the parent strains SL 1344 , the mutant strain had obvious change in its virulence , oral challenge of bacteria in mice revealed that LD50 of the mutant strain was 1.70 ×108 CFU,the toxicity reduced about 1.4%.The protection rate induced by the sipB mutant was 50%,and the serum antibody peaked 14 d post-immunization.Conclusion:The SL1344ΔsipB mutant was constructed suc-cessfully,and genetic stability ,significantly reduced virulence.The study provides a new approach for further study of the relationship between the gene and pathogenicity of Salmonella typhimurium.It is likely that the ΔsipB mutant could be adapted to develope attenuated Salmonella vaccine.
10.Characterization of attenuated Salmonella C500 strain with a delta asd mutant and use as an Asd+ balanced-lethal host-vector system.
Zhanqin ZHAO ; Yindi XU ; Bin WU ; Xiangchao CHENG ; Yinju LI ; Xibiao TANG ; Chunjie ZHANG ; Huanchun CHEN
Chinese Journal of Biotechnology 2009;25(1):29-36
Salmonella enterica serovar Choleraesuis strain C500 is a live, attenuated vaccine that has been used in China for over 40 years to prevent piglet paratyphoid. The objective of this study was to evaluate the potential of attenuated Salmonella enterica serovar Choleraesuis C500 strain with a delta asd mutant as an effective live vaccine vector by the Asd+ balanced-lethal host-vector system. Here, we compared the characteristics of S. enterica serovar Choleraesuis delta asdC500 strain with the parent C500 strain, including phenotype, growth rate, virulence, safety, and expression for heterologous antigen. The mean generation times of delta asdC500 mutant, the vector control delta asdC500 (pYA3493), and the parent avirulent C500 vaccine strain in Luria broth were 30.7, 28.1, and 27.9 min, respectively. The fermentation patterns of theses three strains on different carbohydrates, and the levels of production of H2S, were similar. The O and H antigens of delta asdC500 mutant, delta asdC500 (pYA3493) and delta asdC500 (pYA-F1P2) were 6,7:C:1,5, identical to the parent strain C500. By the method of Reed and Muench, groups of mice were challenged by the intraperitoneal route with different amounts of delta asdC500 (pYA3493) or the parent C500 strain, and the virulence of delta asdC500 (pYA3493) with LD50 of 1.1 x 10(7) CFU was a little lower than C500 with LD50 of 4.4 x 10(6) CFU. All piglets inoculated with delta asdC500 (pYA3493) or C500 survived, and no signs of disease were observed during the entire experimental period. No major differences were found in these two groups. In addition, the recombinant pYA-F1P2 plasmid was very stable in the recombinant delta asdC500 (pYA-F1P2) strain, which expressed secretorily a large amount of the recombinant filamentous hemagglutinin type I domain and pertactin region 2 domain antigen (rF1P2) of Bordetella bronchiseptica. In this study, we have shown that the delta asdC500 mutant had a series of biological characteristics similar to the parent vaccine strain C500. Furthermore, the strain could express secretorily a large amount of heterologous antigen. It is likely that this Salmonella expression and delivery system could be easily adapted to develop multivalent recombinant Salmonella vaccines against infectious agents using the Asd+ balanced-lethal host-vector system.
Amino Acid Oxidoreductases
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genetics
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Animals
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Bacterial Proteins
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genetics
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Gene Deletion
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Genetic Vectors
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Mice
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Mutation
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Salmonella Vaccines
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genetics
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immunology
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Salmonella enterica
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genetics
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immunology
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pathogenicity
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Swine
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Transduction, Genetic
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Vaccines, Attenuated
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genetics
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immunology
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Vaccines, Synthetic
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genetics
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immunology
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Virulence