Mottram Patricia L, Leong David, Crimeen-Irwin Blessing, Gloster Simone, Xiang Sue D, Meanger Jayesh, Ghildyal Reena, Vardaxis Nicholas, Plebanski Magdalena
The Burnet Institute at Austin (Austin Research Institute), Studley Road, Heidelberg 3084, VIC, Australia.
Mol Pharm. 2007 Jan-Feb;4(1):73-84. doi: 10.1021/mp060096p.
Previous studies compared uptake by dendritic cells (DC) of 20, 40, 100, 200, 500, 1000, and 2000 nm beads in vivo. When beads were used as antigen carriers, bead size influenced antibody responses and induction of IFN-gamma-producing CD4 and CD8 T cells. Beads of 40-50 nm were taken up preferentially by DC and induced particularly strong immunity. Herein, we examine immunity induced by minute differences in nanobead size, specifically within a narrow viral-sized range (20, 40, 49, 67, 93, 101, and 123 nm), to see if bead carrier size influenced the induction of type 1 or type 2 cells as demonstrated by the production of IFN-gamma or IL-4. In vivo uptake by DC was assessed for selected sizes in this range. Responses to whole ovalbumin (OVA) or the OVA-derived CD8 T cell peptide epitope (SIINFEKL) were tested. After one immunization with beads-OVA, IFN-gamma responses to both OVA and SIINFEKL were significantly better with 40 and 49 nm beads than other sizes, while, in contrast, IL-4 responses to OVA were higher after immunization with OVA conjugated to larger beads (93, 101, and 123 nm). Thus IFN-gamma induction from CD8 T cells was limited to 40-49 nm beads, while CD4 T cell activation and IL-4 were induced by 93-123 nm beads-OVA. After two immunizations, there were comparable high levels of IFN-gamma produced with 40 and 49 beads and IL-4 reactivity was still higher for larger beads (93, 101, 123 nm). Production of IgG1 was seen across the full range of bead sizes, increasing after two immunizations. Since protection against respiratory syncytial virus (RSV) depends on strong IFN responses, while IL-4 responses are reported to cause asthma-like symptoms, immunization with RSV antigens on the 49 nm carrier beads could provide the basis for a suitable vaccine. When the 49 nm beads were conjugated to RSV proteins G88 (surface) or M2.1 (internal capsid), one immunization with G88 induced high levels of IFN-gamma and low levels of IL-4. IL-4 increased with two immunizations. Beads-M2.1 induced only moderate levels of IFN-gamma and low titer antibody after two immunizations. Mice vaccinated once with G88-conjugated 49 nm beads and challenged intranasally with RSV strain A2 subtype showed reduced viral titers and recovered from weight loss more rapidly than mice immunized with M2.1-conjugated 49 nm beads or naive control mice. These results show that precise selection of nanobead size for vaccination can influence the type 1/type 2 cytokine balance after one immunization, and this will be useful in the development of effective vaccines against common human pathogens such as RSV.
先前的研究比较了树突状细胞（DC）在体内对20、40、100、200、500、1000和2000纳米珠子的摄取情况。当珠子用作抗原载体时，珠子大小会影响抗体反应以及产生干扰素-γ的CD4和CD8 T细胞的诱导。40 - 50纳米的珠子优先被DC摄取，并诱导出特别强的免疫反应。在此，我们研究了纳米珠子大小的微小差异，特别是在狭窄的病毒大小范围内（20、40、49、67、93、101和123纳米）所诱导的免疫反应，以观察珠子载体大小是否如通过干扰素-γ或白细胞介素-4的产生所证明的那样影响1型或2型细胞的诱导。评估了该范围内选定大小的珠子在体内被DC摄取的情况。测试了对完整卵清蛋白（OVA）或OVA衍生的CD8 T细胞肽表位（SIINFEKL）的反应。用珠子-OVA进行一次免疫后，40和49纳米的珠子对OVA和SIINFEKL的干扰素-γ反应明显优于其他大小的珠子，而相比之下，用与较大珠子（93、101和123纳米）偶联的OVA免疫后，对OVA的白细胞介素-4反应更高。因此，CD8 T细胞诱导的干扰素-γ仅限于40 - 49纳米的珠子，而93 - 123纳米的珠子-OVA诱导CD4 T细胞活化和白细胞介素-4。两次免疫后，40和49纳米的珠子产生的干扰素-γ水平相当高，而较大珠子（93、101、123纳米）的白细胞介素-4反应性仍然更高。在整个珠子大小范围内都观察到了IgG1的产生，两次免疫后增加。由于针对呼吸道合胞病毒（RSV）的保护依赖于强烈的干扰素反应，而据报道白细胞介素-4反应会导致类似哮喘的症状，在49纳米载体珠子上用RSV抗原进行免疫接种可为合适的疫苗提供基础。当49纳米的珠子与RSV蛋白G88（表面）或M2.1（内部衣壳）偶联时，用G88进行一次免疫诱导出高水平的干扰素-γ和低水平的白细胞介素-4。两次免疫后白细胞介素-4增加。珠子-M2.1在两次免疫后仅诱导出中等水平的干扰素-γ和低滴度抗体。用G88偶联的49纳米珠子接种一次的小鼠，经鼻内用RSV A2亚型毒株攻击后，病毒滴度降低，体重减轻恢复得比用M2.1偶联的49纳米珠子接种的小鼠或未免疫的对照小鼠更快。这些结果表明，精确选择用于疫苗接种的纳米珠子大小可在一次免疫后影响1型/2型细胞因子平衡，这将有助于开发针对RSV等常见人类病原体的有效疫苗。
