基因药物载体重组古菌组蛋白的免疫原性研究

目的 观察基因药物载体重组古菌组蛋白(HphA)在鼠模型中的免疫原性,为开发安全有效的基因药物载体重组古菌组蛋白奠定基础.方法 Balb/c小鼠随机分为重组古菌组蛋白实验组、重组古菌组蛋白佐剂实验组、牛血清白蛋白佐剂对照组,测定免疫小鼠中体液和细胞免疫应答水平.体液免疫应答水平通过采用酶联免疫吸附试验(ELISA)考察测定血清中的抗体水平;细胞免疫应答水平通过测定脾淋巴细胞的增殖指数和其分泌的IFN-γ浓度进行评价.结果 ELISA法未检测出重组古菌组蛋白免疫小鼠血清中产生特异性抗体,淋巴细胞的刺激指数及其分泌IFN-γ浓度与对照组相比无统计学意义(P＞0.05).结论 重组古菌组蛋白免疫在鼠模型中没有免疫原性,不能诱导免疫小鼠产生特异性体液应答和细胞免疫应答.     

壳聚糖包裹IL-21与PEB1核酸纳米粒子疫苗免疫应答的研究

目的研究壳聚糖包裹PEB1和IL-21核酸纳米粒子疫苗诱导小鼠免疫应答水平。方法复合物凝聚法制备壳聚糖纳米粒子,肌肉注射Balb/c小鼠,检测免疫期和感染期小鼠血清特异性Ig G水平、脾细胞悬液细胞因子水平、脾细胞增殖指数及疫苗保护率。结果扫描电镜观察纳米颗粒平均粒径在(300±23)nm左右,包封率为(91.23±3.24)%;实验组小鼠,特别是IL-21基因佐剂组,在血清特异性Ig G、脾细胞培养上清中IFN-γ和IL-4、脾细胞增殖水平和疫苗的临床保护效应上都显著高于对照组(P0.05);但壳聚糖纳米颗粒组效果不明显(P0.05)。结论 IL-21基因佐剂可以显著提高并维持小鼠稳定持久的免疫应答水平。但壳聚糖对免疫应答仅起到一定的促进作用。本研究的顺利开展,为空肠弯曲菌疫苗的临床应用提供了一定的理论支持。     

铜绿假单胞菌重组Bb-OprF疫苗诱导小鼠细胞免疫应答的研究

目的研究铜绿假单胞菌重组双歧杆菌(rBb)-OprF疫苗免疫小鼠,PAO1株攻击后产生的细胞免疫应答。方法将rBb-OprF疫苗分别采用皮下注射、肌肉注射、鼻腔黏膜和口服灌胃4种途径免疫Balb/c小鼠,免疫后8周用5×106CFU的PAO1株攻击,攻击1周后杀鼠取脾,MTT法检测特异性脾淋巴细胞增殖,流式细胞术检测脾CD4+T和CD8+T细胞比率,用ELISA法检测脾细胞培养上清IFN-γ、IL-12、TNF-α和IL-10的水平。结果脾T淋巴细胞增殖明显;脾细胞CD4+和CD8+T细胞亚群显著增加;脾细胞培养上清IFN-γ、IL-12、TNF-α和IL-10的水平显著升高,其中IFN-γ最为显著。结论铜绿假单胞菌rBb-OprF疫苗可诱导小鼠产生混合型的Th1和Th2免疫应答。     

黑大蒜提取物对小鼠细胞免疫应答影响的初步探讨

目的:探讨黑大蒜提取物对BALB/c小鼠细胞免疫功能的影响。方法:连续5天给予小鼠腹腔注射黑大蒜提取物溶液。从第6天开始处死鼠,分离培养脾细胞,生物学方法检测自然杀伤(NK)细胞杀伤活性,Griess法检测脾细胞培养上清一氧化氮(NO)的分泌水平,ELISA法检测脾细胞培养上清IL-2、IL-4、IFN-γ和TNF-α的水平,RT-PCR方法检测脾细胞FasLmRNA的表达水平。结果:黑大蒜提取物能显著增强NK细胞杀伤活性,促进NO和Th1型细胞因子IL-2、IFN-γ和TNF-α的分泌,IL-4水平略有降低,脾细胞FasLmRNA的表达水平未见明显变化。结论:黑大蒜提取物对小鼠细胞免疫应答具有一定的促进效应。     

CpG ODN佐剂促进HBsAg疫苗诱导小鼠细胞免疫应答的探讨

目的 探讨cpG ODN对重组乙型肝炎表面抗原（HBsAg）诱导小鼠细胞免疫应答的影响。方法HBsAg和HBsAg＋CpG ODN分别肌肉注射免疫Balb／c小鼠后,从淋巴组织（脾和淋巴结）和非淋巴组织（肺）分离淋巴细胞,体外经HBsAg抗原刺激后,利用ELISA检测细胞培养液中IFN-γ的水平,再利用流式细胞仪在单个细胞水平上检测IFN-γ^＋细胞的频率,分析IFN-7^＋细胞亚群。结果对照组和HBsAg免疫组IFN-γ产生的水平很低,当HBsAg加强免疫1～2次后,IFN-γ表达仍然没有明显升高;而CpG ODN＋HBsAg免疫1次或加强免疫后,CpG显著促进HBsAg诱导的IFN-γ产生。进一研究结果表明CpG促进HBsAg诱导淋巴器官和非淋巴器官内CD4^＋和CD8^＋T细胞IFN-γ的表达。结论CpG免疫佐剂不仅能诱导细胞免疫应答同时也诱导体液免疫应答,提示CpG ODN可以用于HBsAg预防和治疗性佐剂。     

IL-2 与IL-33 佐剂增强铜绿假单胞菌外膜囊泡的免疫保护效果

目的:探究IL-2、IL-33佐剂对铜绿假单胞菌(Pa)外膜囊泡(OMVs)抗感染免疫保护作用的影响。方法:体外培养Pa提取OMVs后加入IL-2、IL-33佐剂,并于0、2、4周皮下注射免疫雌性BALB/c小鼠,每组18只。末次免疫2周后,ELISA检测各组小鼠血清IgG、IgA及IgG亚类水平以及小鼠脾细胞上清中IFN-γ、IL-2、IL-4、IL-6等细胞因子水平,同时检测CD4、CD8蛋白水平;CCK-8法评估IL-2、IL-33分子佐剂对小鼠脾细胞增殖情况的影响;末次免疫2周后Pa滴鼻感染小鼠,感染后10 d统计各组小鼠死亡率,检测肺组织中菌体载量,并通过HE染色评估肺组织病理损伤情况。结果:ELISA结果显示,OMVs-IL-2、OMVs-IL-33组小鼠血清中IgG、IgA以及IgG亚类水平均显著高于各对照组(P0.05),且其中各实验组IgG2a/IgG1均大于1;IL-2、IL-33均能够显著促进OMVs诱导小鼠脾细胞分泌IFN-γ、IL-2及IL-12(P0.05),但对细胞因子IL-4、IL-6分泌水平影响无统计学意义(P0.05);此外,OMVs-IL-2、OMVs-IL-33免疫小鼠脾细胞上清中CD4、CD8蛋白水平均显著高于对照组(P0.05);CCK-8结果显示,OMVs-IL-2、OMVs-IL-33组的淋巴细胞增殖水平最为显著(P0.05);Pa感染后,OMVs-IL-2、OMVs-IL-33组小鼠的死亡率及肺组织菌体载量均显著低于各对照组(P0.05),且小鼠肺组织的炎症侵润程度均明显减轻。结论:IL-2、IL-33佐剂能够增强Pa-OMVs诱导的体液及细胞免疫应答水平,显著提高其抗感染保护作用,具有较大临床意义。     

Flt3L及CCL5对prime/boost免疫策略中抗原特异性免疫应答的增强及抗肿瘤作用

目的:研究Flt3L与CCL5作为联合佐剂在prime/boost免疫策略中对HBc抗原特异性免疫应答的增强及抗肿瘤作用.方法:将两种细胞因子质粒与携带HBc抗原的DNA疫苗经肌内注射法共免疫小鼠, 免疫3次后再用原核表达的HBc颗粒蛋白或HBc DNA疫苗加强, 观察对稳定表达HBcAg 的小鼠黑色素瘤细胞(B16-HBc)的生长抑制作用;并分别采用MTT法检测荷瘤小鼠脾淋巴细胞增殖、流式细胞术检测脾CD8 T淋巴细胞中IFN-γ表达、ELISA法检测脾淋巴细胞培养上清IL-2、IL-4含量及乳酸脱氢酶(LDH)释放法检测特异性CTL杀伤活性.结果:与对照组相比, 佐剂联合DNA疫苗免疫经蛋白加强组(DDP/Adj)显著抑制肿瘤生长;佐剂联合DNA疫苗免疫组(DDD/Adj)及DDP/Adj组均可促进特异性淋巴细胞增殖反应(P＜0.05), 且DDP/Adj 高于DDD/Adj组(P＜0.05);DDD/Adj 及DDP/Adj组小鼠脾脏CD8 T淋巴细胞中IFN-γ表达、IL-2 表达水平及CTL杀靶活性均高于对照组(P＜0.01或P＜0.05), IL-4 表达水平在各组无显著区别(P＞0.05).结论:在prime/boost免疫策略中, 采用Flt3L与CCL5两种细胞因子联合应用可显著促进荷瘤小鼠产生抗原特异性免疫应答及抗肿瘤作用.     

调节性T细胞修饰前炎症应答对小鼠脑型疟疾发生的影响

为探讨调节性T细胞（Tregs）对伯氏疟原虫感染所致鼠脑型疟发生和感染结局的影响机制,用伯氏疟原虫ANKA株分别感染对照组和抗CD25单克隆抗体注射组C57BL／6小鼠,计数红细胞感染率;感染前和感染后3、5、8天制备脾细胞悬液,流式细胞术检测脾Tregs百分含量;ELISA和Griess方法检测脾细胞培养上清IFN-γ、IL-10和NO水平。结果表明大多数C57BL／6鼠于感染后8—11天死于脑疟,抗CD25单克隆抗体注射组小鼠感染后3～4周死于贫血和过度原虫血症。对照组小鼠脾细胞培养上清IFN-γ、NO、IL—10水平于感染后开始升高,感染后5天达到峰值,感染后8天与感染后5天相比,IFN-γ、NO轻微下降,IL-10显著下降。感染后3、5天,实验组IFN-γ、NO水平显著高于对照组,IL—10水平显著低于对照组。感染后8天,实验组和对照组IFN-γ、NO、IL-10水平得到逆转。这表明Tregs通过修饰前炎症应答影响伯氏疟原虫感染鼠脑型疟发生和感染结局。     

粘膜佐剂LTB优化的抗淋病ltB-nspA融合基因疫苗鼻饲免疫效果研究

目的:用真核重组质粒pcDNA3.1(-)/ltB-nspA鼻饲免疫小鼠,探索粘膜佐剂LTB辅佐NspA所诱发的特异性体液免疫应答和细胞免疫应答水平。方法:对真核重组质粒行PCR及双酶切鉴定后大量制备,经鼻饲途径免疫雌性BALB/c小鼠,间接ELISA法检测血清中NspA特异性IgG抗体水平及小鼠生殖道灌洗液中NspA特异性sIgA抗体水平;MTT法检测脾淋巴细胞增殖水平,ELISA双抗体夹心法检测脾淋巴细胞培养上清IFN-γ含量。结果:融合基因pcDNA3.1(-)/ltB-nspA组小鼠生殖道灌洗液中sIgA(A450:0.316±0.045)明显高于pcDNA3.1(-)/nspA单基因组(P0.05)和其它对照组(P0.01);小鼠血清中IgG(A450:0.643±0.156)水平、脾淋巴细胞刺激指数(SI:1.65±0.32)和脾淋巴细胞诱生的IFN-γ(160.56±25.67pg/ml)水平均明显高于pcDNA3.1(-)/ltB、空质粒pcDNA3.1(-)和PBS对照组(P0.01)。结论:pcDNA3.1(-)/ltB-nspA融合基因疫苗经鼻饲免疫,能够诱导小鼠产生较强的特异性体液免疫应答和细胞免疫应答;粘膜佐剂LTB可辅佐NspA诱导小鼠产生更高水平的生殖道粘膜免疫。     

副黏病毒Tianjin株缺损性干扰颗粒增强小鼠免疫应答的初步研究

目的 探讨副黏病毒Tianjin株缺损性干扰(DI)颗粒作为新型生物佐剂的可行性.方法 分离提纯副黏病毒Tianjin株DI颗粒并鉴定,和脊髓灰质炎病毒(PV)灭活抗原等量混合后给小鼠皮下多点注射,并设氢氧化铝凝胶佐剂组、无佐剂组以及空白对照组.于免疫后14 d摘眼球取血检测免疫血清PV特异性IgG;无菌取脾脏制备脾细胞悬液,分别用MTT法进行T、B淋巴细胞增殖试验以及检测脾淋巴细胞接受PV灭活抗原再次刺激后IFN-γ、IL-4的产生情况.结果 副黏病毒Tianjin株DI颗粒佐剂组T、B淋巴细胞刺激指数(SI)、脾淋巴细胞分泌IFN-γ的量与其他组相比较,差异有统计学意义(P＜0.05).结论 副黏病毒Tianjin株DI颗粒具有一定的增强机体免疫应答能力的作用,且以诱导Th1型细胞免疫应答为主.     

不同刺激剂诱导肺癌患者引流淋巴结细胞分泌细胞因子的研究

目的:探讨多种刺激剂对肿瘤引流淋巴结(TDLN)细胞分泌细胞因子的影响及其抗肿瘤效应和机制。方法:采用ELISA和Griess法测定不同刺激组(分别为IL2、IL2 自身肺癌细胞抗原、IL2 GMCSF IL4 自身肺癌细胞抗原和IL2 GMCSF IL4 LPS)刺激TDLN后第7、14、21天,分泌IL12p70、IFNγ、TNFα和NO的水平。结果:IL2 GMCSF IL4 自身肺癌细胞抗原和IL2 GMCSF IL4 LPS组刺激的TDLN细胞中,CD83 细胞的比率明显增多,分泌IL12p70、IFNγ及TNFα的水平也明显高于IL2和IL2 自身肺癌细胞抗原刺激组,IL2 GMCSF IL4 LPS组刺激的TDLN细胞分泌NO的水平明显高于其他3组。各种刺激剂刺激TDLN细胞后,分泌IL12p70、IFNγ和NO的水平在第14天时达到高峰。结论:不同刺激剂诱导TDLN细胞中的CD83 细胞数量不同,故具有不同的抗肿瘤活性。     

低免疫原性肿瘤可诱导调节性T细胞增生

目的以小鼠低免疫原性瘤细胞和高免疫原性瘤细胞与同系脾细胞混合培养为肿瘤免疫体外模型,研究培养后的脾细胞中CIM（＋）CD25（＋）调节性T细胞（TR）的分布,揭示肿瘤免疫逃逸的机制。方法3种不同高低免疫原性瘤细胞与同系脾细胞混合培养,建立模拟肿瘤免疫的体外模型,以放射性核素掺入法检测混合培养后的脾细胞的增殖,流式细胞术分析TR、CIM（＋）干扰素（IFN）γ（＋）以及CIM（＋）白细胞介素（IL）-10（＋）T细胞分布状况,ELISA法检测混合脾细胞和肿瘤细胞培养卜清液中IFN-1和IL-10水平。结果高免疫原性瘤细胞FBL3或H22刺激的同系脾细胞增殖指数较低免疫原性瘤细胞D5刺激的旧系脾细胞要高,分别高3倍（D5：FBL3=4．94：12．20）和10倍（D5：H22=4．94：44．60）,而3种瘤细胞刺激的同种脾细胞其增殖指数均较相应的肿瘤免疫组要高（D5 1．9倍,FBL3 2．1倍,H22 1．1倍）。在与低免疫原性瘤细胞D,混合培养的同系脾细胞中,与高免疫原性瘤细胞FBL3或H22相比,含更多的TR（D5：H22P〈0．05）和CD4^＋IL-10^＋细胞（D5：FBL3P〈0．01,D5：H22P〈0．01）,上清液中含更高水平的IL-10（P〈0．01,P〈0．01）,而IFN-γ水平很低（P〈0．01,P〈0．01）。结论低免疫原性肿瘤可诱导TR细胞的增生,TR细胞在肿瘤免疫逃逸中有重要作用。     

The comparative selectivity of adjuvants for humoral and cell-mediated immunity. I. Effect on the antibody response to bovine serum albumin and sheep red blood cells of Freund''s incomplete and complete adjuvants, alhydrogel, Corynebacterium parvum, Bordetella pertussis, muramyl dipeptide and saponin

A comparison was made of seven recognized adjuvants, Freund's incomplete and complete, alhydrogel, Corynebacterium parvum, Bordetella pertussis, muramyl dipeptide and saponin, administered with BSA or SRBC by the S.C. route of immunization. Strong selectivity as well as differences in potency were revealed in relation to these two antigens. Only FIA, FCA, alhydrogel and MDP promoted the primary response to 50 microgram of BSA, and FIA was significantly superior to FCA. Immunological memory to a low dose (0.5 microgram) of BSA, which did not evoke a primary response with any adjuvant, was potentiated by alhydrogel and by MDP and, relatively poorly, by FIA. Radioimmunoelectrophoresis showed that potentiation of the response with MDP was confined to IgG1, whereas alhydrogel, FIA and FCA stimulated both IgG1 and IgG2. Saponin was outstandingly the best adjuvant for both primary and secondary haemagglutinin responses to SRBC. Of the others, alhydrogel for the primary, and alhydrogel and B. pertussis for the secondary were active to a lesser degree. The results show that the relative potency of adjuvants differs markedly according to the antigen used, and suggest that saponin may be a particularly effective adjuvant for antigens in cell membranes.     

不同刺激诱导肺癌患者引流淋巴结细胞抗自体肿瘤的体外研究

目的：探索适合于临床大规模应用的有效刺激、活化肿瘤引流淋巴结（Tumor-draining lymph node，TDLN）细胞的方法。方法：对3种刺激剂（IL-2、IL-2＋自身肺癌细胞抗原和IL-2＋GM-CSF＋IL-4＋自身肺癌细胞抗原）诱导TDLN细胞体外抗自体肿瘤作用，通过乳酸脱氢酶法测定了CTL杀伤活性，观察了细胞形态学变化，通过流式细胞技术测定了细胞CD83的阳性表达率。结果：经MTT法检测，IL-2＋GM-CSF＋IL-4＋自身肺癌细胞抗原诱导的TDLN细胞的增殖程度明显高于IL-2＋自身肺癌细胞抗原刺激组和IL-2刺激组（P〈0．01）。IL-2＋GM-CSF＋IL-4＋自身肺癌细胞抗原诱导的肿瘤特异性CTL活性明显高于IL-2＋自身肺癌细胞抗原刺激组和IL-2刺激组，CD83阳性表达细胞率也明显高于IL-2＋自身肺癌细胞抗原刺激组和IL-2刺激组。结论：用IL-2＋GM-CSF＋IL-4＋自身肺癌细胞抗原刺激和活化TDLN，优于单独用IL-2刺激或IL-2＋自身肺癌细胞抗原刺激，该作用可能与其诱导TDLN细胞产生DC数量的增加有关。     

Adjuvants Influence the Immunoglobin Subclass Distribution of Immune Responses in Vivo

Mice were immunized against fluorescein isothiocyanate (FITC)-labelled human gamma globulin (HGG) in the absence or presence of different adjuvants. The immune response was assayed every other day with regard to both total Ig-secreting cells and FITC-specific plaque-forming cells (PFC). The adjuvants influenced the type of immune response induced to the same antigenic determinant. Thus, addition of Freund's complete (FCA) or incomplete (FIA) adjuvant preferentially led to the secretion of IgG1 PFC of an average high affinity. Most newly appearing IgG-secreting cells were also detected as FITC-specific PFC. The use of lipopolysaccharide (LPS) as an adjuvant resulted in the induction of both IgM and IgG, particularly of the IgG3 and IgG2b subclasses. However, these antibodies had relatively low affinity, and a large number of total IgG-secreting cells induced by LPS had no detectable FITC specificity. The FCA/FIA- and LPS-induced responses to FITC-HGG were additive when injected together, indicating that they act on distinct subpopulations of B lymphoid cells. The adjuvant response to LPS, but not the response to FCA/FIA, was totally absent in mice of the C3H/Hej strain, which are non-responders to the polyclonal activating properties of LPS. Finally, the response induced by FCA or FIA was T-cell-dependent and the LPS response T-cell-independent as assayed in nude mice.     

Changes of cytokines in mice immunized with recombinant Bb-EmII/3-Em14-3-3 vaccine of Echinococcus multilocularis

AIM: To study the effects of recombinant Bb-EmII/3-Em14-3-3 vaccine on cytokine secretion in mice challenged with protoscoleces of Echinococcus multilocularis. METHODS: BALB/c mice were immunized with rBb-EmII/3-Em14-3-3 vaccine by subcutaneous injection, intramuscular injection, nasal mucosa inoculation or oral administration. After 12 weeks of immunization,all the mice were challenged with 50 protoscoleces of Echinococcus multilocularis by intraperitoneal injection. 18 weeks later, the mice were sacrificed and the splenocytes were stimulated with EmAg and ConA or LPS in vitro, then IL-12, IL-10, IFN-gamma and TNF-alpha in the culture supernatant were measured by ELISA. RESULTS: The level of IFN-gamma, IL-12 and TNF-alpha were greatly higher than that of PBS control group (P<0.05 or P<0.01), and the level of IL-10 was lower. The level of IFN-gamma, IL-12, TNF-alpha and IL-10 in each group with EmAg and ConA or LPS stimulation was greatly higher than that in the group without stimulation(P<0.05 or P<0.01). CONCLUSION: Th1 response is induced in mice challenged with Echinococcus multilocularis by rBb-EmII/3-Em14-3-3 vaccine, and the vaccine may enhance protective response against the challenge with protoscoleces of Echinococcus multilocularis.     

Adjuvant modulation of the immune responses and the outcome of infection with Chlamydia pneumoniae

Immunization with different adjuvants resulted in antithetic outcomes of infection with Chlamydia pneumoniae. Immunization with the outer major protein-2 from C. pneumoniae (OMP-2) emulsified in Freund's complete adjuvant (FCA) thus increased the susceptibility of mice to infection with the bacteria. The detrimental effect was not observed upon inoculation of irrelevant antigens or major outer membrane protein (MOMP) in FCA, but was also observed after immunization with FCA-chlamydial heat shock protein-60 (HSP-60). The harmful effect of FCA-OMP-2 depended on the presence of both CD4+ and CD8+ cells and was mediated by IL-10, as shown using gene-ablated mice. The increased susceptibility to infection caused by FCA-OMP-2 immunization was long-lasting and observed in mice infected 4 months after the last dose of immunogen. In contrast, partial protection against C. pneumoniae was observed when FCA was replaced with oligodeoxynucleotides containing immunostimulatory CpG motifs mixed with Freund's incomplete adjuvant (FIA-IS-CpG). These polar outcomes of infection related to the cytokine pattern: antigen-stimulated spleen cells from FCA-OMP-2-immunized mice showed higher IL-10/IFN-gamma ratios than FIA-IS-CpG-OMP-2-immunized animals. In agreement, sera from FCA-OMP-2 showed higher anti-OMP-2 IgG1/IgG2a ratios than FIA-IS-CpG-OMP-2-immunized animals. Finally, OMP-2 also generated a protective response when delivered by a eukaryotic expression vector in tandem with CTLA4, a procedure that targeted OMP-2 to antigen-presenting cells.     

Regulation of cytokine gene expression by adjuvants in vivo

Antibody isotype affects biological activity of the antibodies and therefore should be considered in prevention of disease by vaccination. In previous reports, we demonstrated that adjuvants affect the antibody isotype switching process and favour the production of certain isotypes. The present study extends these findings and shows fundamental differences in the cytokine induction pattern according to the adjuvant used. Cytokine mRNA levels were determined by in situ RNA–RNA hybridization performed on splenocytes isolated from mice injected with different adjuvants. The results revealed that Freund's complete adjuvant (FCA), Freund's incomplete adjuvant (FIA), Al(OH)₃ and QuilA administration results in a type-2 (humoral) response, increasing IL-4, IL-5 and IL-13 gene expression, while poly I:C exhibits a type-1 (cell-mediated) response, increasing the production of interferon-gamma (IFN-γ), IL-2 and IL-6 mRNA. Finally, BeSO₄ and poly A:U augment IL-5 and IL-6 mRNA production, while lipopolysaccharide (LPS) and LiCl augment IL-6 and tumour necrosis factor-alpha (TNF-α) mRNA production. Also, the adjuvants appear capable of overcoming the inherent IL-2/IFN-γ and IL-4 dichotomy of C57Bl/6 and BALB/c mice, respectively, in response to cellular antigens such as Leishmania and herpessimplex virus (HSV). The overall data suggest that adjuvants direct the isotype switching process via induction of certain cytokines, a finding that can be useful in selection of the most efficient isotype of protective antibodies for disease prevention by vaccination.     

Contact sensitization to oxazolone: involvement of both interferon-gamma and interleukin-4 in oxazolone-specific Ig and T-cell responses.

The synthesis and role of several lymphokines were examined during contact sensitization to oxazolone (OX). Application of OX to the skin of mice increased the delayed-type hypersensitivity (DTH) response to challenge, serum titres of OX-specific IgG1 and IgG2a, and draining lymph node cell (LNC) numbers. At day 3, LN contained detectable interleukin-4 (IL-4), interferon-gamma (IFN-gamma) and granulocyte-macrophage colony-stimulating factor (GM-CSF) but not IL-2 or IL-3 mRNAs; IL-3 and higher levels of IL-4, IFN-gamma and GM-CSF mRNAs were measured after 24 hr culture with anti-CD3 antibody in OX-primed but not unprimed LNC. As a result of sensitization, LNC secreted IL-3 constitutively and produced elevated levels of IL-2, IL-3, IL-4 and IFN-gamma in response to anti-CD3 antibody; a similar but weaker lymphokine response was recalled by OX-protein conjugate. CD4+ cells were the major source of the anti-CD3-induced lymphokines except IFN-gamma, which was derived mainly from CD8+ cells. Since both IL-4 and IFN-gamma were synthesized by OX-primed LNC in vivo and in vitro, their role was investigated by administering anti-lymphokine antibodies at the time of sensitization. Anti-IL-4 treatment reduced OX-specific serum IgG1 titres without affecting IgG2a titres, whereas anti-IFN-gamma treatment reduced IgG2a but not IgG1 titres. Although neither antibody altered DTH responsiveness, anti-IFN-gamma treatment markedly increased IL-4 production by CD4+ LNC and reduced IFN-gamma production in vitro, particularly by CD4+ cells. We conclude that endogenous IL-4 and IFN-gamma reciprocally influence the isotype of the Ig response to OX and that IFN-gamma also affects the relative levels of IL-4 and IFN-gamma synthesis by CD4+ LNC.     

人乳头瘤病毒18型L1-E6、E7嵌合基因DNA疫苗的体内免疫效应

目的：检测HPV18 L1-E6、E7嵌合基因DNA疫苗在小鼠体内的体液和细胞免疫效应。方法：将实验动物BALB/c小鼠54只随机分为9组，按不同免疫方式(肌肉接种或鼻内滴注)分别给予不同的重组质粒(pVAX1-L1-E6M3或pVAX1-L1-E7M3)和免疫佐剂(pLXHDmB7-2或LTB)。用免疫原免疫3次，末次免疫后取眼球后血进行ELISA抗体检测。末次免疫后进行小鼠足垫迟发型超敏反应(DTH)试验。断足进行小鼠足垫HE染色。取小鼠脾脏制成单细胞悬液，进行脾细胞增殖试验，并进行CD4^ /CD8^ T细胞中IFN-γ^ 或IL-4^ 细胞的FACS分析。结果：与对照组相比，各实验组均获得明显的免疫效果。实验组免疫后血清抗体4值均高于相应组别免疫前；肌肉注射组每次免疫后抗体水平较前次明显升高。实验组小鼠注射VLP抗原的左后足垫局部有红肿硬结，镜下观察可见大量单核细胞侵润。肌肉接种组的DTH反应强度、脾细胞增碹刺激指数(SI)和CD8^ IFN-γ^ 细胞数均高于鼻内滴注组；而鼻内滴注组CD4^ IL-4^ 细胞数高于单纯质粒肌肉接种组；加入pLXHDmB7-2的联合免疫组各项指标均高于单纯质粒组。结论-证实了重组pVAX1-HPV18L1/E6、E7嵌和基因DNA疫苗能诱导小鼠的体液免疫和细胞免疫效应：同时，证实B7-2分子能显著提高该质粒在小鼠体内的免疫反应效果.