| CD40L对单纯疱疹病毒2型DNA疫苗的免疫增强作用研究 |
| 投稿时间：2015-07-07 修订日期：2015-07-21 点此下载全文 |
| 引用本文：胡如西,陶薇,傅婷,何卓晶,贾岚,洪艳,陈勇.CD40L对单纯疱疹病毒2型DNA疫苗的免疫增强作用研究[J].医学研究杂志,2015,44(10):29-33,37 |
| DOI：
10.11969/j.issn.1673-548X.2015.10.009 |
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| 基金项目:浙江省自然科学基金资助项目(LY12H19009) |
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| 中文摘要:目的 研究重组质粒pcDNA3-Kan/CD40L辅佐HSV-2 DNA疫苗增强小鼠体液免疫和细胞免疫的作用效果,探讨其作为HSV-2 DNA疫苗佐剂的潜力。 方法 (1)构建鼠CD40L基因的重组真核表达质粒pcDNA3-Kan/CD40L。(2)体外细胞实验:检测重组质粒pCD40L刺激小鼠外周血淋巴细胞增殖情况和脾细胞分泌IFN-γ的能力。(3)体内动物实验:48只雌性BALB/c小鼠随机分为4个免疫组pK组、pgD组、pcCD40L+pgD组和pK+pgD组。小鼠后腿肌内注射,共免疫2次,间隔3周。末次免疫3周后,每组随机取4只小鼠进行致死剂量攻毒实验验证疫苗的保护作用。 ELISA检测小鼠血清抗HSV-2 IgG抗体水平和趋化因子RANTES;流式细胞术检测全血中CD4+和CD8+T细胞百分率以及分泌IFN-γ和IL-4的T细胞的百分率;MTS法检测小鼠脾脏T细胞的增殖能力。 结果 (1)体外实验结果:重组质粒pcDNA3-Kan/CD40L对小鼠外周血淋巴细胞的增殖能力和刺激脾细胞分泌IFN-γ的能力均显著大于空质粒pcDNA3-Kan(P<0.05)。(2)体内实验结果:小鼠血清抗HSV-2 IgG水平、趋化因子RANTES、脾淋巴细胞刺激指数和外周血CD4+T细胞数和分泌IFN-γ的Th1细胞数均高于其他免疫组(P<0.05)。pcDNA3-Kan/CD40L+pgD组预防小鼠感染HSV-2效果好于其他免疫组。 结论 (1)重组质粒pcDNA3-Kan/CD40L能够诱导外周血淋巴细胞增殖并刺激脾细胞分泌IFN-γ具有作为疫苗佐剂的潜力。(2)pcDNA3-Kan/CD40L可以辅助HSV-2 DNA疫苗诱导BALB/c小鼠产生特异性抗HSV-2的体液免疫和细胞免疫,具备作为HSV-2 DNA疫苗免疫佐剂的能力。 |
| 中文关键词:CD40配体(CD40L) 单纯疱疹病毒2型(HSV-2) 疫苗佐剂 |
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| Plasmid CD40L Enhances Immune Response of HSV-2 DNA Vaccine |
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| Abstract:Objective To detect HSV-2-specific humoral immunological response and cellular immunological response in BALB/c mice which were induced by plasmid CD40L-adjuvanted HSV-2 DNA vaccine. Methods 1The murine CD40L gene transcript was inserted into the pcDNA3 vector to obtain the recombinant plasmid pcDNA3-Kan/CD40L. 2In vitro study: MTS colorimetric method was employed in the detection of the rat peripheral blood lymphocytes proliferation and SYBRgreen qPCR assay was used to test the IFN-γ secretion ability of spleen cells. 3In vivo study: Forty eight female BALB/c mice were randomly divided into four groups: pKan, pgD, pCD40L+pgD and pKan+pgD, and inoculated through intramuscular immunization at the weeks 0 and 3. After 6 weeks the protection given to the mice was assayed by a fatal dose of HSV-2. The humoral immunological response and the cellular immunological response were detected by enzyme linked immunosorbent assay (ELISA), MTS colorimetric assay and flow cytometry (FCM). Results 1The ability of stimulation lymphocytes proliferation of rat PBMC and IFN-γ level in spleen cells of cDNA3-Kan/CD40L group were significantly better than that of pK group (P<0.05).2The level of anti-HSV-2 IgG, RANTES, stimulation index (SI), CD4+ and IFN-γ in pCD40L+pgD group were significantly higher than anther groups. Furthermore,mice of pCD40L+pgD group were prophylactically protected from challenge with a high dose of HSV-2. Conclusion ①The potential of pcDNA3-Kan/CD40L could be used as an adjuvant in vaccines.②The potential of pCD40L-adjuvantd HSV-2 DNA vaccine could induce systemic humoral immune responses and cellular immune responses intramuscular vaccinated mice. |
| keywords:CD40 ligand (CD40L) Herpes simplex virus type 2 (HSV-2) Vaccine adjuvant |
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