靶向多肽功能化的智能响应型疫苗佐剂递送龋齿抗原用于龋齿的防治

Dental caries endangering oral health is one of the most common oral diseases worldwide, particularly in developing countries. It is of great significance to find effective ways to prevent dental caries and improve the oral health of human beings. In recent years, due to the significant role in the prevention and treatment of dental caries, dental caries vaccine has been widely concerned and studied. However, due to the weak immunogenicity of the antigen in dental caries vaccine, the function of the vaccine is greatly reduced. Therefore, it is the key point to increase the immunogenicity of the antigen and enhance the immune response. Vaccine adjuvant is a kind of substance that can effectively improve the immune response. To design the proper vaccine adjuvant in dental caries vaccine can effectively increase the immunogenicity of the antigen and enhance specific immune response. In the previous study, our research group designed and synthesized different types of nanocarriers used as vaccine adjuvants, which could enhance the immune response. Based on the previous research, This project intends to design the pH-responsive mesoporous hydroxyapatite as the core loading the antigen PAc from Streptococcus mutans causing dental caries, which were modified with redox-responsive poly(amido amine), and were functionalized with CPE30 targeted with M cell, and then to prepare a vaccine delivery system which is more effective, can target immune system cell, has high cell penetration ability and property of intelligent responsive release of antigen to achieve the effective prevention and treatment of dental caries.

龋病是人类最常见的细菌感染性疾病之一，严重危害口腔健康。寻找有效的龋病防治方法、改善人类口腔健康具有重要的意义。近年来，龋齿疫苗因在龋齿防治中的显著作用，而被广泛关注和研究。但是因为龋齿疫苗中抗原弱的免疫原性，而大大降低了疫苗的效价，所以增加抗原免疫原性、提高免疫应答成为研究的关键。疫苗佐剂是一类可以有效提高免疫应答的物质，在龋齿疫苗制备过程中设计恰当的疫苗佐剂可以有效的增加龋齿疫苗的效价，从而解决这一关键问题。我们课题组前期研究中制备不同类型的纳米载体用于疫苗佐剂，在一定程度上提高了机体的免疫应答水平。本项目拟在前期研究的基础上，以pH响应型介孔羟基磷灰石为内核负载引起龋齿的变异链球菌的抗原蛋白PAc，用还原响应型聚酰胺胺修饰，并用具有黏膜上皮M细胞靶向功能的多肽CPE30功能化，制备具有免疫系统细胞靶向功能的、高细胞渗透能力和胞内环境响应智能释放抗原的疫苗递送系统，实现龋齿的有效防治。

龋病是人类最常见的细菌感染性疾病之一，严重危害人类口腔健康。龋病疫苗具有显著防治龋病作用，被广泛研究。然而，龋病疫苗中抗原免疫原性低，显著降低了疫苗效价。因此，需要疫苗佐剂增加抗原免疫原性，提高免疫应答。所以项目在国家自然科学基金委员会的资助下，从体外和体内深入研究了CPE30靶向多肽功能化的智能响应型纳米疫苗佐剂的制备表征、促进抗原递呈、提高免疫应答用于龋病预防。首先，基因工程技术构建PAc抗原蛋白体外表达系统，成功表达纯化PAc抗原蛋白和PAc-GFP荧光标记抗原；合成靶向多肽CPE30功能化聚酰胺胺聚合物，用其修饰介孔羟基磷灰石制备疫苗佐剂，负载抗原成功制备龋病疫苗。然后，在体内外研究疫苗的生物安全性、黏膜渗透、细胞内吞和溶酶体逃逸能力，显示其具有良好的生物安全性、有效渗透进入鼻黏膜下层、促进抗原内吞和pH响应释放。最后，采用滴鼻免疫接种方式，间隔两周免疫小鼠三次，采集小鼠血清、唾液、脾细胞等检测血清和唾液抗体滴度、脾细胞增殖、细胞因子分泌和T细胞分型，研究表明疫苗可以诱导小鼠产生高水平血清IgG抗体和唾液SIgA抗体、分泌高水平细胞因子（IL-4、IL-6、IL-10、IFN-γ）、强的脾细胞增殖能力和显著的中心及效应记忆T细胞，阐明疫苗佐剂增强抗原免疫原性，提高免疫应答的作用机制；并构建龋病大鼠模型，疫苗免疫后显著增强PAc特异免疫应答，抑制口腔变形链球菌生长达到龋齿预防的目标，阐明龋病疫苗预防龋病的作用机制。