Role of immunoglobulin CDR-H3 in heterosubtypic immunity to influenza virus

描述(由申请人提供)：我们寻求更好地了解抗体库在提供对流感病毒的异亚型免疫方面的整体作用。我们已经发现，我们可以将抗原结合部位分组，按特定的物理化学性质定义。我们已经证明，在生命的关键阶段，某些类别是首选的，而其他类别似乎是积极选择的，特别是在年轻人中，他们通常比老年人更能抵抗异型感染。初步研究表明，在老年人中可以发现使用不受欢迎的抗原结合部位类别的免疫球蛋白。由于抗体库控制的一般原理对人类和小鼠似乎是共同的，为了研究抗体库控制在异型免疫中的作用，我们创造了小鼠，其抗体库仅限于这些高度不受欢迎的类别之一。我们以前使用这些小鼠来表明，当携带属于正常首选类别的抗原结合位点的抗体完全被使用不受欢迎的类别的抗体取代时，对H1N1毒株的异亚型攻击的保护严重受损。然而，这些研究没有回答是否由于缺乏正常的首选谱系或存在属于不受欢迎的类别的序列而损害了异亚型免疫的问题。在目前的提案中，我们测试了这样的假设，即不受欢迎的免疫球蛋白的存在可以对预防异亚型感染产生显性负面影响。这是一个关键问题，因为如上所述，剧目的组成可以随着年龄或疾病的变化而变化。杂合子的改变的免疫球蛋白等位基因包含突变的DH基因座产生正常的谱系从正常的等位基因，并不受欢迎的抗体从基因的目标等位基因。这些小鼠将被用来测试这些不受欢迎的抗体的存在是否足以增加小鼠的发病率和死亡率，在小鼠接种H3N1或H1N1毒株，然后用同源或异种病毒攻击后。发病率和死亡率的增加将证实，控制抗体库的全球组成对于适当保护对流感病毒的免疫反应至关重要。随后将对可能导致感染易感性增加的两种潜在机制进行测试。小鼠将被注射来自NAOVE或免疫小鼠的血清或骨髓单个核细胞，然后用同源或异种病毒攻击。使用血清后发病率的增加将表明可溶性不受欢迎的抗体的直接影响。只有在细胞转移后发病率才会增加，这表明免疫球蛋白作为B细胞受体的直接作用，可能是通过改变T细胞的抗原呈递。然后我们将滴定效果以确定超过该阈值的异种亚型免疫受损。再加上评估曲目多样性的新方法，这些信息将被用来告知流感病毒感染的易感性，并指导疫苗接种和治疗战略。 与公共卫生相关：我们试图了解人体产生的全部抗体的组成如何影响对流感病毒的保护。我们计划使用这些信息来识别有更高发病率和死亡率的风险个人，并指导疫苗接种和治疗策略。

DESCRIPTION (provided by applicant): We seek to gain a better understanding of the role of the antibody repertoire as a whole in providing heterosubtypic immunity to influenza virus. We have found that we can group of antigen binding sites into categories defined by specific physical-chemical properties. We have shown that some categories are preferred at key stages of life, whereas others appear to be actively selected against, especially in young adults who typically are more resistant to heterosubtypic infection than aged individuals. Preliminary studies indicate that immunoglobulins using 'disfavored' categories of antigen binding sites can be found in aged individuals. Because the general principles of antibody repertoire control appear common to human and mouse, in order to study the role of repertoire control in heterosubtypic immunity we created mice whose repertoire is limited to one of these highly disfavored categories. We previously used these mice to show that when antibodies bearing antigen binding sites belonging to normally preferred categories were completely replaced by antibodies using a disfavored category, protection against heterosubtypic challenge with an H1N1 strain was severely compromised. However, these studies did not answer the question of whether it was the absence of the normally preferred repertoire or the presence of sequences belonging to a disfavored category that had impaired heterosubtypic immunity. In the present proposal, we test the hypothesis that the presence of disfavored immunoglobulin can have a dominant negative effect on protection against heterosubtypic infection. This is a key issue because, as noted above, the composition of the repertoire can change with age or disease. Mice heterozygous for altered IgH alleles containing mutations in the DH locus produce a normal repertoire from the normal allele, and disfavored antibodies from the gene targeted allele. These mice will be used to test whether the presence of these disfavored antibodies is sufficient to increase morbidity and mortality after mice are immunized with either an H3N2 or an H1N1 strain, and then challenged with homologous or heterologous viruses. The increase in morbidity and mortality will confirm that control of the global composition of the antibody repertoire is essential for properly protective immune responses to influenza virus. Two potential mechanisms that could underlie this increases susceptibility to infection will then be tested. Mice will be injected with sera or infused with bone marrow mononuclear cells from naove or immunized mice, and then challenged with homologous or heterologous viruses. An increase in morbidity after administration with the sera will point to a direct effect of soluble disfavored antibody. An increase in morbidity only after transfer of cells will suggest a direct role of immunoglobulin as a B cell receptor, perhaps by altering antigen presentation to T cells. We will then titer the effect to determine the threshold beyond which heterosubtypic immunity is impaired. Coupled with new means to evaluate repertoire diversity, this information will then be used to inform susceptibility to influenza virus infections and to guide vaccination and treatment strategies. PUBLIC HEALTH RELEVANCE: We seek to understand how the composition of the totality of antibodies produced by the body may affect protection against influenza virus. We plan to use this information both to identify individuals at risk for greater morbidity and mortality, and to guide vaccination and treatment strategies.