铝佐剂和免疫程序对Sabin IPV免疫原性影响的研究

目的 研究铝佐剂和免疫程序对不同配比的Sabin IPV免疫原性的影响.方法 用Ⅰ、Ⅱ、Ⅲ型脊髓灰质炎病毒原液制备四批Sabin IPV,分别为中剂量无佐剂组、中剂量铝佐剂组、低剂量无佐剂组、低剂量铝佐剂组,采用0、1、2月和0、2、4月两种免疫程序对大鼠进行3针基础免疫,每针基础免疫一个月后采血,并检测免疫后血清中3个型别的中和抗体水平.结果 经过3针基础免疫后,各组Ⅰ、Ⅱ、Ⅲ中和抗体几何平均滴度均显著提高,阳转率均达到100％.对不同组别间两种免疫程序比较无显著差异,但相同条件下,0、2、4月免疫程序可诱导产生更高的抗体水平.对相同免疫程序下不同组别进行比对,添加铝佐剂组与无佐剂组相比可以诱导更高的抗体水平.结论 铝佐剂和免疫程序可提高Sabin IPV免疫原性.     

新型CpG ODN增强乙肝疫苗诱导IgG2a类抗体产生的实验研究

目的：寻找能增强乙肝疫苗刺激IgG2a类抗体产生，使机体处于Th1样免疫环境的新型乙肝疫苗佐剂。方法：选用自行设计的A、B、C型CpGODN，并以发表的A、B型CpGODN作为阳性对照，与重组乙肝疫苗混合后于第0．4周免疫BALB／C小鼠，用ELISA法检测免疫小鼠血清中HBsAb水平及种类。结果：各型CpGODN都能增强乙肝疫苗刺激HBsAb的产生水平，CpGODN＋乙肝疫苗免疫的小鼠血清中HBsAb类型为IgG2a〉〉IgG1，而单独应用商品化重组乙型肝炎疫苗的小鼠血清中HBsAb类型为IgG1〉〉IgG2a。结论：各型CpGODN对重组乙型肝炎疫苗[含AI（OH）3佐剂]均具有增效作用，而且可以诱导机体产生倾向于Th1途径的免疫应答反应。     

铝佐剂类型及吸附率对吸附无细胞百白破联合疫苗免疫原性的影响

目的研究铝佐剂类型及吸附能力对吸附无细胞百白破联合疫苗免疫原性的影响,筛选最佳佐剂。方法用磷酸铝(AP)、不同浓度磷酸盐处理的氢氧化铝(PTAH)以及氢氧化铝(AH)佐剂吸附百白破抗原,检测不同佐剂对各抗原的吸附率。制备不同佐剂类型和吸附率的制剂,腹腔免疫NIH小鼠,ELISA法测定免疫后各组小鼠血清抗体滴度。结果百日咳丝状血凝素(FHA)在所有佐剂中吸附率≥95%,百日咳类毒素(PT)、百日咳溶血素(PRN)、精制白喉类毒素(DT)和破伤风类毒素(TT)则随着铝佐剂中磷酸根增多吸附率降低。动物实验产生PT抗体水平大小关系:AP/AHPTAH/无佐剂组;产生FHA抗体水平大小关系:AP/PTAH-3/AHPTAH-2/PTAH-3无佐剂组;产生PRN抗体水平大小关系:PTAH/AP/AH无佐剂组;产生DT抗体水平大小关系:APPTAHAH无佐剂组;产生TT抗体水平大小关系:PTAHAH/AP/无佐剂组。结论铝佐剂吸附力不影响抗原(DT除外)免疫原性,磷酸铝更适合作吸附无细胞百白破联合疫苗的佐剂。     

IL-23、IL-27对RSV重组蛋白疫苗免疫原性的影响

目的:以编码IL-23和IL-27的真核表达质粒pcDNA3.1-IL-23(以下简称IL-23)和pcDNA3.1-IL-27(以下简称IL-27)为佐剂,与呼吸道合胞病毒(RSV)重组蛋白疫苗G1F/M2共免疫小鼠,观察IL-23和IL-27对疫苗的免疫原性的影响。方法:以IL-23、IL-27质粒和Al(OH)3为免疫佐剂,与G1F/M2共免疫BALB/c小鼠,末次免疫后10天杀死小鼠,用ELISA检测特异性IgG、IgG1和IgG2a水平;流式细胞术检测小鼠脾细胞CD4+、CD8+细胞的变化;用乳酸脱氢酶(LDH)释放法检测特异性小鼠脾细胞杀伤活性。结果:G1F/M2+Al(OH)3+IL-23和G1F/M2+Al(OH)3+IL-27可诱导高效价的IgG、IgG1、IgG2a抗体,且显著高于这三种佐剂单独使用诱导的抗体效价;流式细胞检测结果显示G1F/M2+IL-23和G1F/M2+Al(OH)3+IL-23刺激的CD4+T细胞和CD8+T细胞水平显著高于其他组;单独的IL-23或IL-27对G1F/M2诱导的脾细胞杀伤活性没有增强作用,而Al(OH)3+IL-23和Al(OH)3+IL-27佐剂组的杀伤率显著高于单独的IL-23、IL-27或Al(OH)3组。结论:这些结果表明:IL-23或IL-27质粒与传统铝盐佐剂Al(OH)3联合使用能显著增强RSV重组疫苗G1F/M2的免疫原性。     

灭活SARS病毒的免疫原性的实验研究

研究灭活SARS病毒的免疫原性。SARS病毒F6 9株经甲醛灭活后 ,加入氢氧化铝佐剂 ,以 3种剂量接种BALB/c小鼠 ,定时采血 ,测定特异性抗体的滴度及其中和活性 ,同时用化学发光酶联免疫法测定抗血清与SARS病毒结构蛋白的反应特异性及其相对强度。结果发现 ,小鼠接种疫苗 4d后 ,血清中可检测到IgM抗体 ,直至 2 6d后逐渐下降 ;IgG抗体在初次免疫 8d后出现 ,4 7d时达最高峰 ,6 3d后进入稳定期 ;不同剂量组的抗体滴度具有明显剂效关系 ,低剂量组和中剂量组滴度峰值为 1∶192 0 0 ,高剂量组滴度峰值为 1∶384 0 0。中和实验结果表明 ,小鼠所产生的抗体具有中和病毒活性 ,在 6 3d时 ,低剂量组和中剂量组血清的中和效价为 1∶12 80 ,高剂量组血清的中和效价为 1∶5 12 0。抗体分类结果表明 ,小鼠抗血清中含有针对多种SARS病毒结构蛋白的特异性抗体 ,其中 ,针对N蛋白、S4蛋白和S2蛋白的抗体水平相对较高 ,而抗M抗体、抗 3CL抗体的水平相对较低。上述结果说明 ,SARS病毒F6 9株经甲醛灭活后 ,各主要结构蛋白仍保持较强免疫原性 ;免疫小鼠后 ,可以诱导产生高滴度的特异性混合抗体 ,在体外可以保护敏感细胞不受SARS病毒攻击     

空肠弯曲菌外膜蛋白亚单位脂质体佐剂疫苗的免疫原性研究

目的 初步探讨自制的空肠弯曲菌（campylobacter jejuni）亚单位（M,28 000-31 000外膜蛋白）脂质体佐剂疫苗的免疫原性,为该疫苗的临床前研究打下基础。方法 用双向免疫琼脂扩散试验、试管凝集试验、ELISA（间接法）分别检测经GJ疫苗免疫后的新西兰兔血清IgG抗体效价。结果 GJ阴离子组与GJ阳离子组比较,血清IgG A值显著增高（P〈0．01）,与GJ弗氏组无统计学差别（P〉0．1）;CJ阴、阳离子组脂质体疫苗接种剂量在0．50mg／kg时,血清IgGA值最大;CJ阳离子皮下注射组与肌肉注射组IgGA值无统计学差别（P〉0．1）;特异性IgG抗体经6个月动态观察,5个月后抗体效价下降。结论 实验组全程免疫后均有特异性抗体产生;同一剂量的阴离子脂质体疫苗比阳离子脂质体疫苗血清抗体水平显著增高;脂质体疫苗最佳接种剂量为0．50mg／kg,肌肉注射与皮下注射免疫效果无显著差异;特异性抗体高水平维持5、6个月后应加强免疫。     

三种不同载体的C群脑膜炎球菌荚膜多糖-蛋白质结合疫苗比较研究

目的 通过3种不同载体即破伤风类毒素（TT）、白喉类毒素（DT）和重组铜绿假单胞菌外毒素A（rEPA）与C群脑膜炎球菌荚膜多糖（GCMP）结合，经免疫小鼠后比较3种结合疫苗的免疫原性。方法将GCMP以ADH作为间隔剂分别与TT、DT和rEPA等3种蛋白质载体结合形成GCMP-蛋白质结合疫苗，以GCMP和3种结合疫苗免疫NIH小鼠以研究在其体内的免疫原性。结果GCMP和结合疫苗免疫NIH小鼠后，其血清ELISA结果显示多糖疫苗和结合疫苗均可诱生免疫应答，但使用GCMP免疫小鼠后仅能产生较低水平抗GCMP的IgG抗体，而用3种GCMP-蛋白质结合疫苗免疫小鼠后其血清中产生了较GCMP免疫显著增高的抗GCMP的IgG抗体，并且3种GCMP-蛋白质结合疫苗第2次、第3次免疫与初次免疫相比，IgG抗体水平均有显著升高（P〈0．001），表明GCMP-蛋白质结合疫苗具有免疫记忆和加强应答效应，且3种GCMP-蛋白质结合疫苗在小鼠体内产生的抗GCMP水平差异有统计学意义（P〈0．01）。补体介导的血清抗体体外杀菌试验结果证明，3种GCMP-蛋白质结合疫苗免疫小鼠诱导的IgG抗体均比GCMP具有增强的体外杀菌活性。结论 经3种不同载体与GCMP结合后免疫小鼠均产生了比GCMP显著增强的免疫应答。比较研究表明，rEPA是实验中GCMP-蛋白质结合疫苗较好的备选载体。     

恶性疟原虫裂殖子表面蛋白3功能区片段的制备及其免疫原性分析

目的构建恶性疟原虫裂殖子表面蛋白3功能片段MSP3（69）与谷胱甘肽（GST）的融合蛋白，并在大肠杆菌中进行表达，分析其免疫原性。方法采用大肠杆菌系统表达融合蛋白GST—MSP3（69），以疟疾病人血清进行Western-blot，并以ISA720、佛氏、氢氧化铝与CpG联合3种佐剂与GST—MSP3（69）融合蛋白乳化，免疫Balb／ca小鼠．分析其免疫原性。结果在大肠杆菌系统中成功表达GST-MSP3（69）融合蛋白，疟疾病人血清能与融合蛋白反应。3种佐剂免疫Balb／ca小鼠均能诱发Balb／ca小鼠产生较高水平的特异抗体，3次免疫后的抗体滴度分别为4．5×10^5，4．1×10^5和3．5×10^5。3次免疫后血清抗体亚型分析显示该蛋白能够诱导Balb／ca小鼠产生较高滴度的亲细胞亚型IgG1和IgG2b．ISA720、佛氏、氢氧化铝与CpG联合佐剂组IgG1分别为8×104、7．8×10^4、6．8×10^4．IgG2b分别为1．2×10^4，1．25×10^4、1．5×10^4。统计学分析显示不同佐剂组的抗体滴度及抗体亚型滴度之间差异均无统计学意义（P〉0．05）。结论制备了在大肠杆菌表达的GST-MSP3（69）融合蛋白，该蛋白具有高免疫原性，并产生亲细胞亚型抗体，这些为研究MSP3蛋白功能区的免疫保护作用打下了基础。     

弗氏佐剂与氢氧化铝佐剂对诱导小鼠获得性免疫应答作用的比较

为研究、比较不同佐剂对诱导小鼠产生获得性免疫应答的不同作用,以卵清白蛋白(OVA)为抗原,分别混合完全弗氏佐剂(CFA)或Al(OH)3佐剂,对C57BL/6小鼠进行常规免疫,采用流式细胞技术对细胞内细胞因子IFN-γ和IL-4进行检测;ELISA方法对特异性抗OVA抗体滴度及抗体亚型进行了检测。结果显示在免疫后CFA组产生以IFN-γ为主的细胞因子而Al(OH)3组产生以IL-4为主的细胞因子;两组中均产生特异性抗OVA IgG抗体,但CFA组以IgG2a亚型为主,而Al(OH)3组则以IgG1亚型为主,不产生IgG2a亚型抗体。实验表明,经CFA加抗原免疫后机体产生的免疫应答以Th1型细胞免疫为主,抗体类型为IgG2a;而Al(OH)3佐剂则诱导机体产生Th2型细胞免疫应答,抗体类型为IgG1。     

鼠疫F1-V重组蛋白疫苗滴鼻免疫应答效果的研究

目的 以重组霍乱毒素B亚单位（rCT-B）为鼠疫F1-V重组蛋白的佐剂制备黏膜疫苗,观察小鼠诱导的黏膜免疫和系统免疫应答效果。方法以制备的鼠疫黏膜疫苗滴鼻免疫小鼠4次免疫后,采用间接ELISA检测血清特异性抗F1-V的IgG和IgA抗体及抗体亚型分类,检测鼻咽喉、肺、小肠及阴道灌洗液中特异性抗F1-V的黏膜分泌型IgA;采用流式细胞术检测鼻相关淋巴组织淋巴细胞、脾淋巴细胞、肠系膜淋巴结及小肠PP结T淋巴细胞表型的变化。结果以rCT-B为佐剂的鼠疫F1-V重组蛋白黏膜疫苗滴鼻免疫后,能够诱导血清中IgG、IgA抗体比正常对照组显著升高（P〈0．01）,同时诱导鼻咽、肺、小肠和阴道内特异性黏膜抗体升高,尤其是肺和生殖道冲冼液内抗体升高极为显著（P〈0．01）。与单纯的F1-V组相比,不同剂量比例疫苗组都能诱导较高、较快的血清IgG、IgA和黏膜sIgA,其中1：2疫苗组能诱导更强的系统免疫和黏膜免疫,但是相比之下,5：1疫苗组是最合适的免疫剂量。结论rCT-B佐剂不仅能提高鼠疫F1-V黏膜疫苗的系统全身免疫应答,还能促进诱导呼吸道、消化道和生殖道等局部黏膜sIgA抗体,增强局部免疫应答,提示rCT-B佐剂能显著提高鼠疫感染的免疫应答作用,这为下一步疫苗的免疫保护评价奠定了基础。     

Optimizing oral vaccines: induction of systemic and mucosal B-cell and antibody responses to tetanus toxoid by use of cholera toxin as an adjuvant.

Cholera toxin (CT) is an effective mucosal antigen and acts as an adjuvant when given orally with various antigens; however, few studies have compared the levels of antibody responses to CT and coadministered protein in systemic and mucosal tissues. In this study, we used tetanus toxoid (TT) for assessment of immune responses. Time course and dose-response studies established that 250 micrograms of TT given orally with 10 micrograms of CT three times at weekly intervals induced high serum and gastrointestinal tract anti-TT and anti-CT antibody responses. Oral immunization with TT alone induced no detectable mucosal immunoglobulin A (IgA) antibodies in fecal extracts and only weak serum IgG anti-TT responses. The coadministration of CT and TT induced peak serum IgG anti-TT responses following two oral doses that remained constant after the third oral immunization, while optimal mucosal IgA responses were seen after the third oral immunization. The serum anti-TT response obtained with CT and TT proved protective against TT challenge (100 minimum lethal doses), whereas mice orally given CT or TT alone died. Antigen-specific B-cell responses were assessed with an isotype-specific Elispot assay of isolated lymphoid cells from the spleen, Peyer's patches, and the small intestinal lamina propria. Interestingly, approximately fourfold-higher numbers of IgA anti-CT than of anti-TT antibody-producing (spot-forming) cells occurred in lymphocytes from the lamina propria of mice orally immunized with both TT and CT. The adjuvant CT did not induce polyclonal B-cell responses in mice given CT by the oral route, since no significant differences in total numbers of B cells producing IgA, IgG, or IgM were found compared with the numbers in mice given TT alone. The results clearly indicate that serum and mucosal antibody responses develop with different kinetics and that protective TT-specific antibody responses are generated in the systemic compartment when TT is administered with CT via the oral route.     

The immune response to anti-idiotype antibodies bearing an internal image epitope of tetanus toxin/toxoid. I. Induction of the humoral immune response

Primary immune responses to tetanus toxoid (TT) and primary and secondary immune responses to a rabbit TT internal image bearing anti-idiotype antibody (Ab2 beta 1) inoculated in Freund's complete adjuvant (FCA), saline (SAL) or syntex adjuvant formulation vehicle (SAF) via the intraperitoneal or subcutaneous route, were examined in mice. High anti-TT antibody (Ab3) titres are reported although the titre and persistence of the antibody response varied according to the adjuvant used in the priming and challenge inocula of Ab2 beta 1. Mouse Ab3 antibodies were elicited in mice inoculated with rabbit Ab2 beta 1 antibodies which in turn were elicited by an inoculum of mouse monoclonal anti-TT Ab1 antibody. Ab3 was shown to be identical to Ab1 by immunoblot analysis. Primary and secondary immune responses elicited by rabbit Ab2 beta 1 antibody protected mice against a lethal dose of tetanus toxin.     

Combined meningococcal serogroup A and W135 outer-membrane vesicles activate cell-mediated immunity and long-term memory responses against non-covalent capsular polysaccharide A

Outer-membrane vesicles (OMVs) have inherent adjuvant properties, and many vaccines use OMV as vaccine components. Utilizing the adjuvant properties of OMV could lead to the formulation of vaccines that are less expensive and potentially more immunogenic than covalently conjugated polysaccharide vaccines. We evaluated the adjuvant effect in Balb/c mice of combinations of OMV from Neisseria meningitidis serogroup A and W₁₃₅ as compared to that of the non-covalently conjugated capsular polysaccharide A. Both antigens were adsorbed onto aluminum hydroxide. The mice were given a booster dose of plain polysaccharide A to stimulate an immunologic memory response. Subclasses determination and cytokine assays demonstrated the capacity of OMV to induce a IgG2a/IgG2b isotype profile and IFN-γ production, suggesting the induction of a Th1 pattern immune response. Lymphoproliferative responses to OMVs were high, with affinity maturation of antibodies observed. Bactericidal titers after the booster dose were also observed. Memory B cells and long-term memory T cells were also detected. The results of this study indicate that combined meningococcal serogroup A and W₁₃₅ OMV can activate cell-mediated immunity and induce a long-term memory response.     

Parenteral adjuvant activities of Escherichia coli heat-labile toxin and its B subunit for immunization of mice against gastric Helicobacter pylori infection

The heat-labile toxin (LT) of Escherichia coli is a potent mucosal adjuvant that has been used to induce protective immunity against Helicobacter felis and Helicobacter pylori infection in mice. We studied whether recombinant LT or its B subunit (LTB) has adjuvant activity in mice when delivered with H. pylori urease antigen via the parenteral route. Mice were immunized subcutaneously or intradermally with urease plus LT, recombinant LTB, or a combination of LT and LTB prior to intragastric challenge with H. pylori. Control mice were immunized orally with urease plus LT, a regimen shown previously to protect against H. pylori gastric infection. Parenteral immunization using either LT or LTB as adjuvant protected mice against H. pylori challenge as effectively as oral immunization and enhanced urease-specific immunoglobulin G (IgG) responses in serum as effectively as aluminum hydroxide adjuvant. LT and LTB had adjuvant activity at subtoxic doses and induced more consistent antibody responses than those observed with oral immunization. A mixture of a low dose of LT and a high dose of LTB stimulated the highest levels of protection and specific IgG in serum. Urease-specific IgG1 and IgG2a antibody subclass responses were stimulated by all immunization regimens tested, but relative levels were dependent on the adjuvant used. Compared to parenteral immunization with urease alone, LT preferentially enhanced IgG1, while LTB or the LT-LTB mixture preferentially enhanced IgG2a. Parenteral immunization using LT or LTB as adjuvant also induced IgA to urease in the saliva of some mice. These results show that LT and LTB stimulate qualitatively different humoral immune responses to urease but are both effective parenteral adjuvants for immunization of mice against H. pylori infection.     

Adjuvant effects of aluminum silicate on IgE and IgG1 antibody production in mice

The adjuvant effects of aluminum silicate on IgE and IgG1 antibody production were investigated. BALB/c mice were immunized intraperitoneally with 10 micrograms ovalbumin (OA) adsorbed on 0.2, 2, or 20 mg aluminum silicate. The enhancement of anti-OA IgE antibody production was observed in the mice injected with aluminum silicate and antigen compared with the mice injected with antigen alone. Anti-OA IgE antibody production with 2 and 20 mg aluminum silicate was greater than that with aluminum hydroxide (alum) as an adjuvant. Similar adjuvant effects with 2 mg aluminum silicate or alum were observed in AKR and C57BL/6 mice using 10 micrograms OA, and in BALB/c mice using 2 micrograms DNP-KLH (dinitrophenyl keyhole limpet hemocyanin): IgE antibody production induced by aluminum silicate adjuvant persisted for weeks in these experiments. The enhancement of IgG1 antibody production to OA mixed with aluminum silicate was also demonstrated. However, no difference between aluminum silicate and alum was observed on the IgG1 antibody production.     

Synthesis and immunological properties of conjugates composed of group B streptococcus type III capsular polysaccharide covalently bound to tetanus toxoid

A synthetic scheme for covalently binding group B streptococcus type III to tetanus toxoid (TT), using adipic acid dihydrazide as a spacer, is described. Type III alone or as a conjugate with TT was injected subcutaneously into laboratory mice, and the type-specific and TT antibody responses elicited by these immunogens were assayed. Type III-TT elicited significantly higher levels of type-specific antibodies after each immunization than did the type III alone. These levels were related to the dosage of the conjugate, enhanced by Freund adjuvant, and exhibited booster responses. Type III alone elicited only immunoglobulin M (IgM) antibodies in Swiss albino mice and mostly IgM and low levels of IgG antibodies of the IgG3 subclass in BALB/c mice. Type III-TT conjugates, in contrast, elicited mostly IgG antibodies in both strains of mice. IgA type III antibodies were not detected. The first two immunizations with the conjugates elicited type III antibodies in the IgG1 and in the IgG3 subclasses. Low levels of IgG2a type III antibodies were detected after a third injection of type III-TT. Conjugate-induced antibodies facilitated opsonization of group B streptococcus type III organisms and did not react with the structurally related pneumococcus type 14. TT alone or as a component of type III-TT induced mostly antibodies of the IgG class: IgG1 levels were the highest of the four subclasses. No IgA TT antibodies were detected. The conjugation procedure, therefore, enhanced the immunogenicity of and conferred T-cell dependent properties to the type III while preserving the immunogenicity of the TT component. The T-cell dependent properties of the conjugates were responsible for stimulating IgG type III antibodies which could be boosted. Evaluation of type III-TT conjugates in antibody-negative women of child-bearing age is planned.     

Immune responses of young mice to pneumococcal type 9V polysaccharide-tetanus toxoid conjugate.

Pneumococcal type 9V polysaccharide (PS), contained in the current pneumococcal vaccine, induces only a weak antibody response in young children and therefore is not an effective vaccine for young children. To increase its immunogenicity, a conjugate of PS to a protein carrier, tetanus toxoid (TT), was prepared. To quantify the immune response, mouse anti-9V PS immunoglobulin G (IgG) and IgM reference standards were established. Young mice immunized at 2 weeks of age produced IgM antibody in response to 9V PS alone or 9V PS conjugated to TT. However, only the 9V PS-TT conjugate induced an IgG antibody response and an anamnestic effect. Thus, a covalent linkage between TT and 9V PS was required for isotype switching from IgM to IgG. 9V PS-TT adsorbed with aluminum hydroxide adjuvant resulted in a fivefold or greater increase in the IgG antibody level. We also studied the effect of maternal immunization on the immune response of young mice to 9V PS-TT. Maternal immunization before mating or before mating and during gestation primed 2-week-old progeny given two injections of 9V PS-TT to produce more IgM antibody than progeny from unimmunized mothers. The IgG antibody level of neonates at birth was similar to that observed in the mothers and was probably passive antibody. These results indicate that maternal immunization with an optimum dose of a PS-protein conjugate before and/or during pregnancy, followed by immunization of the offspring with the conjugate, could provide young children with an enhanced IgM antibody response to pneumococcal PSs.     

Effect of vitamin A supplementation on immunoglobulin G subclass responses to tetanus toxoid in children.

Previously, we demonstrated that administering vitamin A supplements to children resulted in a significant increase in the immunoglobulin G (IgG) response generated against a vaccine dose of tetanus toxoid (TT) (R. D. Semba et al., J. Nutr. 122:101-107, 1991). However, from these analyses we could not determine whether there was an increase in levels of IgG of the subclass presumed to be important for protection against challenge by the toxin or whether there was simply a general increase in the levels of all the IgG subclasses expressing anti-TT activity. The goal of this study was to determine the profile of the anti-TT IgG subclasses in children receiving vitamin A supplementation or a placebo in order to assess the potential utility of the enhanced anti-TT response. In a randomized, double-masked, placebo-controlled clinical trial, the levels of the different anti-TT IgG subclasses were measured in 139 Indonesian preschool children (3 to 6 years of age) 2 weeks before and 3 weeks after immunization. Baseline anti-TT levels and immunization histories were used to separate those children who were responding to TT for the first time from those who responded in a secondary fashion because of previous exposure to TT. Children who were given vitamin A prior to immunization had significant increases in IgG1 levels regardless of whether they were undergoing primary or memory reactions. In the group of individuals who underwent a secondary response to TT, vitamin A supplementation was also associated with a modest but significant change in the levels of anti-TT IgG3. There were only minor changes in the levels of anti-TT IgG2 and IgG4. Since IgG1 is the subclass associated with a protective response to TT immunization, these results suggest that vitamin A supplementation may be a safe and effective intervention to enhance the relevant humoral response to TT and other vaccine antigens.     

Reduction of human anti-tetanus toxoid antibody in hu-PBL-SCID mice by immunodominant peptides of tetanus toxoid

Immunotherapy of murine autoimmune and allergic diseases by administration of peptides corresponding to the dominant T cell epitope is a reality. However, problems remain in applying this therapy to reduce antibody responses in humans. To overcome these difficulties, a preclinical system was developed to test the effect of immunodominant peptides from a common antigen, tetanus toxoid (TT), on the long-term human anti-TT response. Individuals whose T cells proliferated against dominant TT peptides were identified. Peripheral blood leucocytes (PBL) from these donors were injected intraperitoneally (i.p.) into mice with severe combined immunodeficiency (SCID) that had been depleted of murine natural killer (NK) cells (hu-PBL-SCID mice). Peptides or PBS were injected i.p. before a further injection of PBL and immunization with TT. The concentration of human IgG and anti-TT in murine plasma was followed for 10 weeks. The total IgG was similar in both groups. By contrast, there was a statistically significant reduction in IgG anti-TT from eight weeks onwards. It is considered that the hu-PBL-SCID model system may provide a means by which the efficacy of peptide immunotherapy for reduction of pathological antibodies in humans can be examined.     

Interleukin-12 profoundly up-regulates the synthesis of antigen-specific complement-fixing IgG2a,IgG2b and IgG3 antibody subclasses in vivo

The influence of the cytokine interleukin-12 (IL-12) on humoral immune responses was studied in vivo. CBA/J mice immunized with protein antigens (keyhole limpet hemocyanin, phospholipase A₂) adsorbed to aluminum hydroxide (Alum) develop a Th2-like immune response characterized by the production of large amounts of IgG1 as well as some IgE but little IgG2a, IgG2b and IgG3 antibodies. IL-12 is a cytokine that promotes the development and the activation of Th1 cells. Th1 cells are involved in the induction of cellular immunity, which is characterized by low or absent antibody production. On the other hand, some Th1-like immune responses are associated with a strong antibody production of the IgG2a, IgG2b and IgG3 subclasses. Thus, we investigated whether treatment with IL-12 would down-regulate the humoral immune response or stimulate antibody production of the IgG2a, IgG2b and IgG3 subclasses. We observed that: 1) administration of IL-12 to mice together with protein antigens adsorbed to Alum strongly enhanced the humoral immune response by increasing the synthesis of antigen-specific antibodies of the IgG2a, IgG2b and IgG3 subclasses 10- to 1000-fold. The synthesis of IgG1 was not or only slightly (2–5-fold) enhanced, whereas that of the IgE isotype was suppressed. 2) These effects of IL-12 were observed when high (10 μg, 100 μg) or low doses (0.1 μg) of antigen were used for immunization. 3) Titration of IL-12 in vitro revealed that IgG2a is strongly up-regulated over a wide dose range of IL-12 (10 to 1000 ng/day). 4) The effects of IL-12 in vivo are at least partially interferon (IFN)-γ-dependent because an anti-IFN-γ mAb in combination with IL-12 prevented most of the enhanced IgG2a production. 5) Mice receiving IL-12 showed a strong up-regulation of IFN-γ but no inhibition of IL-5 synthesis by spleen cells activated ex vivo with antigen. These results suggest that IL-12 is a potent adjuvant for enhancing humoral immunity to protein antigens adsorbed to Alum, primarily by inducing the synthesis of the complement-fixing IgG subclasses 2a, 2b and 3.