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1.
细胞因子对乙肝病毒基因疫苗诱导产生抗体的影响   总被引:11,自引:1,他引:11  
将构建的三套乙肝病毒基因疫苗(分别编码S蛋白、preS1+preS2+S蛋白及preS2+S蛋白)注射于C57BL/6小鼠胫前肌内。基因疫苗接种3天后,以同样部位注射rhIL-2、rhIL-4、rhIL-6、rhIFN-γ。运用ELISA检测不同时间小鼠血清中乙肝表面抗原(HBsAg)及抗-HBs。结果显示:注射基因疫苗3天内,小鼠体内即可表达HBsAg;两周后,血清中可测到抗-HBs,两月后抗体水平达到高峰,并保持高水平至少两月以上;肌内注射的几种细胞因子均可提高血清中抗-HBs水平,与对照组相比差异有极显著性(P<0.01),其中,rhIL-4、rhIL-6、rhIFN-γ较rhIL-2作用强。提示,细胞因子可作为基因疫苗的佐剂。该研究为增强基因疫苗免疫效果提供了新途径。  相似文献   

2.
目的:构建乙型肝炎病毒(HBV)变异s基因真核表达载体,检测其诱导小鼠产生特异性体液免疫应答。方法:利用限制性内切酶定位克隆构建s基因nt587 G→A的真核表达载体pcMV-S2.S 145R(PR).用其转染人肝癌细胞系Hep G2后,用EIA、EILISA及免疫细胞化学法,观察其抗原性。以重组变异型s基因真核表达载体(PR)和载体pcDNA3.0分别免疫C57BL/6小鼠各5只。每只小鼠各肌肉注射纯化质粒100μg.用ELISA法检测血清抗-HBs及抗-HBs2抗体的效价。结果:体外实验证实,变异型HBs矩可与抗-HBs结合;PR免疫小鼠可诱导其产生抗-HBs抗体及抗.HBs2抗体,但抗-HBs2抗体的出现早于抗-HBs抗体约1~2wk。结论:HBV变异s基因(nt587G→A)的真核表达载体的表达产物具有良好的抗原性,能够诱导C57BL/6小鼠产生体液免疫应答。  相似文献   

3.
目的研究GM-CSF(粒-单核巨噬细胞集落刺激因子)与抗原化抗体联合基因免疫时,GM-CSF对TH细胞应答的调节作用。方法在编码免疫球蛋白重链的质粒中分别克隆黏蛋白1(MUC1)中特异性PDTRP抗原表位和GM-CSF编码基因,构建含PDTRP和GM-CSF编码基因的抗原化抗体表达重组体。经脾免疫和肌肉加强的方式免疫BALB/c小鼠。采用ELISA方法检测小鼠血清中特异性抗体的水平及其亚类并动态观察;RT-PCR方法检测TH细胞分化中细胞因子和转录因子的表达水平。结果抗原化抗体基因免疫能诱导机体产生免疫应答。GM-CSF增强抗原化抗体基因免疫诱生的抗PDTRP特异性IgG抗体水平并且伴随IgG1/IgG2a显著性升高,同时可增强淋巴细胞TH2型细胞因子及转录因子(IL-4、GATA-3)mRNA的表达水平。结论GM-CSF在增强抗原化抗体基因免疫诱导的免疫应答的同时使得免疫应答向TH2方向偏移。  相似文献   

4.
目的:探讨家蚕蛹表达的重组乙型肝炎表面抗原中蛋白[rHBsAg(PreS2 s)]口服免疫原性。方法:选择4周龄的SD大鼠,将重组病毒感染后的家蚕蛹,制成口服剂型灌胃大鼠,ELISA方法检测抗-HBs抗体及抗-PreS2抗体。结果:在灌胃重组蛋白组中有40%鼠体血清内检测到了特异性抗-HBs抗体及抗-PreS2抗体,抗体应答反应持续2—3周;口服免疫后抗体转阳的SD鼠加强免疫后能产生较强的二次免疫应答。进一步研究表明,口服家蚕蛹表达的rHBsAg(PreS2 s)具有加强免疫的效果。对酵母重组疫苗单剂腹腔注射过的SD鼠,用家蚕蛹表达的rHBsAg(PreS2 S)灌胃加强免疫后,能诱导鼠体迅速产生较强的抗-HBs抗体。结论:家蚕/BmNPV表达载体系统将有望开发成为新型的口服乙型肝炎疫苗表达系统  相似文献   

5.
目的 探讨10 μg和20 μg乙肝疫苗与HBIG联合免疫阻断HBV母婴传播的效果.方法 124例HBsAg阳性孕妇所生的婴儿随机分为两组,即10 μg乙肝疫苗组和20 μg乙肝疫苗组.婴儿于出生6h内及30 d分别注射200 IU HBIG,同时分别于出生24 h内、1个月及6个月注射3次10 μg或20 μg重组酵母乙肝疫苗.检测婴儿出生时以及1岁时血清HBV标志物.结果 两组新生儿血清HBsAg、HBeAg及抗-HBe阳性率与滴度之间差别均无统计学意义(P>0.05).所有新生儿血清HBV DNA水平均小于检测下限(500 U/ml).出生12个月时,所有124例婴儿血清HBsAg和HBeAg检测结果均为阴性;血清HBV DNA水平均在检测下限以下;10 μg和20 μg乙肝疫苗组血清抗-HBs阳性率分别为90.3%和96.8%,差异无统计学意义(P>0.05);抗-HBs水平分别为325.5±342.2 mIU/ml和463.7±353.3 mIU/ml,后者显著高于前者(P=0.01).而且,20 μg乙肝疫苗组产生高应答抗-HBs(> 100 mIU/ml)的比例显著高于10μg乙肝疫苗组(P =0.035).结论 20 μg乙肝疫苗联合HBIG方案阻断HBV母婴传播的效果优于10 μg乙肝疫苗联合HBIG方案.  相似文献   

6.
接种Aβ42全肽疫苗恒河猴的特异性体液免疫应答   总被引:2,自引:2,他引:2  
目的 观察恒河猴接种Aβ42肽疫苗后的特异性抗体的产生. 方法 将5只雄性恒河猴分别在0、 2、 6、 10、 14、 18、 22 wk肌内注射Aβ42肽疫苗; 用ELISA法检测恒河猴血清抗Aβ42抗体水平及IgG亚类; 用Western blot检测血清抗Aβ42抗体的特异性; 免疫组化染色法观察抗血清对Tg2576转基因小鼠脑组织中Aβ斑的识别. 结果 疫苗接种后第8周, 恒河猴血清中出现明显的抗Aβ42抗体, 抗体水平随着接种次数的增加而升高, 第24周达1∶ 4 320, 以后抗体水平开始下降.产生的抗Aβ42抗体以IgG1和IgG2为主(IgG2/IgG1>1).血清抗Aβ42抗体具有高度特异性, 可识别Tg2576转基因小鼠脑组织中的Aβ斑. 结论 Aβ42肽疫苗可有效地诱导恒河猴产生特异性体液免疫应答.  相似文献   

7.
HBV preS2S-rhGM-CSF融合基因表达质粒的构建和表达   总被引:3,自引:1,他引:2  
目的:研究GM-CSF和preS2对乙肝DNA疫苗的免疫增强作用。方法:采用PCR方法,扩增HBV preS2 S基因约846 bp的片段和rhGM-CSF(包括甘氨酸接头)基因450bp的片段。通过T-A克隆技术和基因定向克隆,构建融合基因的真核表达质粒pcDNA3.1-S2S-rhGM-CSF,并在HepG2细胞中表达。结果:经酶切、PCR及DNA测序鉴定,融合基因表达质粒HBV preS2S-rhGM-CSF成功地构建。将其转染HepG2细胞后,目的基因的转录通过RT-PCR得到证实,而且表达的融合蛋白能与抗-HBs、抗-preS2和抗-GM-CSF单克隆抗体(mAb)均产生特异性反应。结论:融合基因表达载体pcDNA3.1-S2S-rhGM-CSF的成功构建并表达,为进一步研究乙肝DNA疫苗奠定了实验基础。  相似文献   

8.
目的:探讨了支气管哮喘患儿血清巨噬细胞集落刺激因子(GM-CSF)和降钙素基因相关肽(CGRP)水平的变化及意义.方法:应用放射免疫分析对33例支气管哮喘患儿进行了血清GM-CSF和CGRP测定,并与35名正常健康儿作比较.结果:支气管哮喘患儿在治疗前血清GM-CSF水平非常显著地高于正常儿组(P<0.01),血清CGRP水平则显著地低于正常儿组(P<0.01),经治疗3个月后则与正常儿组比较无显著性差异(P>0.05).结论:血清GM-CSF水平的升高和CGRP水平的降低是支气管哮喘患儿发病的病理因素之一,有重要临床价值.  相似文献   

9.
乙型肝炎表面抗体(抗-HBs)是反映机体对乙型肝炎表面抗原(HBsAg)免疫能力的指标。我们以基因工程技术获得表达人源性抗—HBs Fab片段的工程化大肠杆菌,发酵表达该抗体的可溶性Fab片段,用固相放射免疫分析(SPRIA)定量测定Fab纯化晶的抗-HBs活性,结果如下。 材料和方法 一、人源性抗-HBs Fab抗体的制备:工程化人源性抗-HBs Fab片段的制备参见文献报道,并用抗-HBs阳性病员血清同法纯化作为对照。  相似文献   

10.
目的 研究含有登革病毒Ⅱ型NS1基因的重组质粒肌内注射小鼠后在其体内诱导的细胞和体液免疫。方法 用含有登革病毒NS1基因的真核表达质粒pCNX2 NS1于小鼠胫前肌注射并加强免疫 2次。然后定期处死 ,采集血液标本以及小鼠脾细胞 ,检测小鼠的体液和细胞免疫。结果 在末次免疫后 4周检测到小鼠抗NS1抗体 ,并且检测到小鼠CD4 、CD8 亚群的变化。结论 含有登革病毒NS1基因的真核表达质粒pCNX2-NS1免疫小鼠后 ,可以诱导小鼠产生针对NS1的稳定特异性体液、细胞免疫  相似文献   

11.
DNA-mediated immunization has been recognized as a new approach for prevention and treatment of hepatitis B virus (HBV) infection. However, the side effects of this approach have not been well described. Here we report that DNA-mediated immunization by intramuscular injection of plasmid DNA encoding HBV surface antigen (HBsAg) induced long-term persistence of HBsAg and HBsAg-specific antibody (anti-HBs) in the sera of the immunized BALB/c mice and resulted in liver and kidney lesions. The lesions persisted for 6 months after injection. Lesions were also found in normal mice injected with the sera from immunized mice, and in HBV-transgenic mice injected with anti-HBs antibody, or sera from immunized mice. Furthermore, lesions were accompanied by deposition of circulating immune complex (CIC) of HBsAg and anti-HBs antibody in the damaged organs. These results indicate that long-term persistence of HBsAg and anti-HBs in the immunized mice can result in deposited CIC in liver and kidney, and in development of lesions. The use of DNA containing mammalian replication origins, such as the plasmids used in this study, is not appropriate for human vaccines due to safety concerns relating to persistence of DNA; nevertheless, the safety of DNA-mediated immunization protocols still needs to be carefully evaluated before practical application.  相似文献   

12.
Jerne~([1])指出,机体对抗原的免疫反应可以通过抗体的个体型(Id)和抗个体型抗体(抗-Id)的相互作用而得到调节。这一理论陆续被许多学者的研究结果所证实。  相似文献   

13.
目的:HBV preS2S基因疫苗接种不同时相应用佐剂peDNA3.1IL-2/Fc,观察其对诱导免疫反应效果的影响。方法:采用HBV preS2S DNA基因疫苗作为基础免疫,重组质粒peDNA3.1IL-2/Fc作为佐剂加强免疫,设计实验组与HBVpreS2S基因疫苗同时及在注射BALB/c小鼠后第3天加用佐剂,采用0、2、4周的方案接种,检测各次接种后抗体水平、免疫脾细胞的杀伤活性、增殖活性和细胞因子分泌水平。结果:HBV preS2S注射3天后应用佐剂的免疫小鼠,其抗体滴度、免疫脾细胞杀伤活性、增殖活性和Th1型细胞因子分泌水平均比同时免疫组、单独应用HBV preS2S组及空载体对照组明显增强。结论:预先接种疫苗后加用IL-2/Fc佐剂,能明显增强疫苗免疫效应。  相似文献   

14.
目的 探讨IL-2/Fc融合表达后对HBVpreS2S基因疫苗诱导免疫反应的佐剂效应.方法 采用HBV preS2S DNA疫苗作为基础免疫,重组质粒pcDNA3.1IL-2/Fc作为佐剂加强免疫BALB/c小鼠,采用0、2、4周的方案接种,检测各次接种后抗体水平.初次免疫后7周,测定免疫脾细胞的杀伤活性、增殖活性和细胞因子的分泌水平.结果 pcDNA3.1IL-2/Fc作为佐剂在HBV preS2S注射3 d后免疫组小鼠抗体滴度、免疫脾细胞的杀伤活性和增殖活性、TH1型细胞因子的分泌水平,均比各对照组明显增强.结论 IL-2/Fc是有效的HBV preS2S DNA疫苗佐剂之一.  相似文献   

15.
Efficacy of HBV vaccine in long term prevention of HBV infection was evaluated at 3 years after vaccination in 38 children and 61 adults. All vaccinees were negative for all HBV markers (HBsAg, anti-HBs and anti-HBc) before vaccination. Vaccines (Hevac B) were given for 3 doses, one month apart, to 38 children aged 1 month - 14 years and 61 adults aged 15-45 years. After 3 years of vaccination, blood specimens were collected for the determination of HBsAg, anti-HBs and anti-HBc. The results revealed that no HBsAg antigenemia was found in all 99 vaccinees. Anti-HBs could not be detected in 4 children and 11 adults and this occurred only in the group of subjects who had initial anti-HBs less than 100 mlU/ml at 2 months after the last dose of vaccination. At three years after the first course of vaccination, 89.4 percent of children and 83.4 percent of adults still have anti-HBs above protective level (more than 10 mlU/ml) with geometric mean titers of 101 and 35 mlU/ml in children and in adult groups, respectively. The anti-HBc was detected in 2 out of 38 children and 10 out of 61 adults, but none of them became chronic hepatitis B carriers or developed clinical disease. It is recommended that everyone with anti-HBs values below 100 mlU/ml two months after the last dose of vaccine should be revaccinated with a booster dose within 6 months. Those with anti-HBs levels higher than 100 mlU/ml, should be checked up at 3 years; if the anti-HBs is less than 10 mlU/ml, they should be revaccinated.  相似文献   

16.
Vaccines containing hepatitis B surface antigen (HBsAg) induce antibody to HBsAg (anti-HBs) in most normal individuals and protects them from hepatitis B virus (HBV) infection. However, these vaccines are not efficient at inducing anti-HBs in immunosuppressed individuals, especially in immunosuppressed HBV carriers. The aim of this study was to prepare and to assess the efficacy of a dendritic cell (DC)-based vaccine in an immunosuppressed HBV transgenic mouse (HBV-Tg), an animal model of the HBV carrier state. In order to prepare immunosuppressed HBV-Tg, HBV-Tg were injected with FK-506, an immunosuppressive agent, once daily, intraperitoneally for 15 days. Spleen cells of immunosuppressed HBV-Tg expressed very little mRNAs for interleukin-2 and interferon-gamma. DCs were isolated from the spleen of immunosuppressed HBV-Tg and cultured with HBsAg (100 microg) for 48 h to prepare HBsAg-pulsed DCs. Immunosuppressed HBV-Tg expressing HBsAg in the sera were administered with HBsAg-pulsed DCs or unpulsed DCs or HBsAg in adjuvant for different durations. Immunosuppressed HBV-Tg (n = 8) twice administered with HBsAg-pulsed DCs expressed anti-HBs in the sera within 6 weeks of first injection. Seven of eight immunosuppressed HBV-Tg remained positive for anti-HBs in the sera for the next 12 weeks of observation in spite of receiving daily injection of FK-506 for the entire duration. However, immunosuppressed HBV-Tg administered with unpulsed DCs or HBsAg in adjuvant did not express anti-HBs in the sera. The data show that DCs from immunosuppressed HBV-Tg can be loaded with HBsAg to prepare immunogenic HBsAg-pulsed DCs. HBsAg-pulsed DCs induced anti-HBs in immunosuppressed HBV-Tg. This approach may be of use to induce and maintain anti-HBs in immunosuppressed human HBV carriers.  相似文献   

17.
Hepatitis B virus (HBV) surface antigen (HBsAg) and antibody to HBsAg (anti-HBs) are excellent markers for HBV infection and its immunity. The significance of isolated antibody to HBV core antigen (anti-HBc) seropositivity is not certain. To elucidate this, sera from 638 Chinese adult subjects, aged 18-52 years, seronegative for both HBsAg and anti-HBs, were tested for anti-HBc. Fifty-one (8%) were found to have an isolated anti-HBc seropositivity by ELISA, and all were negative for IgM-anti-HBc. The anti-HBc persisted in all subjects who attended follow-up for hepatitis B vaccination (n = 48) for a period of 8 months. These 48 subjects received 3 doses of hepatitis B vaccine (HB-VAX, 10 micrograms or 20 micrograms) at 0, 1, and 6 months: 72.9% developed a primary anti-HBs response (suggestive of a false-positive anti-HBc seropositivity), 4.2% developed an anamnestic or secondary anti-HBs response, and 22.9% did not develop an anti-HBs response. Increasing the cutoff point of the ELISA or reconfirmation with radioimmunoassay (RIA) reduced only a minor half of the false positives. This low specificity of anti-HBc ELISA/RIA, together with the high rate of anti-HBs response to hepatitis B vaccine, indicates that subjects with isolated anti-HBc seropositivity should be included in vaccination programs.  相似文献   

18.
目的:研究携带HBsAg基因的载体质粒pcDHBs诱导小鼠CTL应答效果。方法:将HBsAg基因连接到真核表达载体pcDNA3.1上,构建成载体质粒pcDHBs。将纯化后的质粒pcDRBs和pcDNA3.1肌肉注射免疫小鼠,眼眶采血检测血清中抗体水平。用质粒pcDHBs转染P815细胞制备乙肝疫苗诱导BALB/C小鼠CTL活性检测的靶细胞。免疫后,取脾细胞,按效靶比为10:1、25:1、50:1进行CIL杀伤检测。结果:免疫pcDHBs疫苗后,检测到小鼠血清中的RBsAb。用pcDHBs进行转染的P815细胞能够检测到HBsAg基因片段和蛋白质抗原的表达。用pcDHBs免疫组小鼠的CTL杀伤率均明显高于pcDNA3.1免疫组。结论:质粒pcDHBs作为核酸疫苗能够诱导小鼠体液免疫应答和CTL免疫应答。  相似文献   

19.
K Kurose  S M Akbar  K Yamamoto  M Onji 《Immunology》1997,92(4):494-500
The inability of hepatitis B virus (HBV) transgenic mice, which express abundant hepatitis B surface antigen (HBsAg) in sera from the neonatal period onwards, to produce antibody to HBsAg (anti-HBs) is considered to be due to defective function of lymphocytes. The defective function is thought to result from neonatal tolerance because antigenic challenge during the neonatal period is considered to be a tolerogenic event rather than an immunogenic one. However, a series of mixed culture experiments in vitro showed that lymphocytes taken from transgenic mice that had been injected with HBsAg in complete Freund''s adjuvant (CFA) constitutively produced anti-HBs when cultured with dendritic cells from age-, sex- and major histocompatibility complex (MHC)-matched normal mice, but not when cultured with dendritic cells from transgenic mice. The expression of major histocompatibility complex (MHC) class II and B 7.2 (CD86) antigens on dendritic cells was significantly lower in transgenic mice compared with the same from the normal mice (P < 0.05). Treatment of transgenic mice with interferon-gamma (IFN-gamma) resulted in up-regulation of MHC class II on dendritic cells, and lymphocytes from HBsAg-injected transgenic mice produced anti-HBs in vitro when cultured with dendritic cells from IFN-gamma-treated transgenic mice, but not when cultured with the dendritic cells from untreated transgenic mice. These experiments have shown that defective function of antigen-presenting cells (APC), not immunogenic tolerance, is responsible for the inability of murine HBV-carriers to produce anti-HBs. Production of anti-HBs by lymphocytes from HBsAg-injected transgenic mice in the presence of dendritic cells that express higher levels of MHC class II and CD86 antigens has inspired optimism that a more effective vaccine therapy can be developed for chronic HBV-carriers, injecting vaccine containing HBsAg with modulator(s) of APC function of dendritic cells.  相似文献   

20.
Six hundred forty-three children, negative for markers of hepatitis B virus (HBV) infections, were given three X 2-micrograms doses of Merck, Sharp and Dohme (MSD) plasma derived hepatitis B vaccine (H-B-Vax) at monthly intervals. Twelve months after the first dose of vaccine, antibody to hepatitis B surface antigen (anti-HBs) was detected in 89% of children by radioimmunoassay (RIA) and in 83% by enzyme immunoassay (EIA). Seroconversion rates and anti-HBs titres were significantly greater in 1-4-year-olds than in older children (p less than 0.01). Eighteen children with no anti-HBs or other markers of HBV at this time were given 10 micrograms of vaccine and tested one month later. Seventeen developed anti-HBs, 12 at levels consistent with an anamnestic response. Forty-nine HBV-marker-negative children seroconverted for antibody to hepatitis B core antigen (anti-HBc) in the 8-month period before or the 12-month period following vaccination. Forty-six of these children were positive for anti-HBs, and one has been confirmed as a chronic carrier of hepatitis B surface antigen (HBsAg). Three cases of clinical hepatitis B in children have been seen in the community since the vaccination programme began. Two of these were amongst the estimated 5% of children who were not vaccinated. The third was in a vaccinee and occurred 4 1/2 months after the last dose of vaccine.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

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