动物实验,疫苗免疫功能 z-bo 2020-12-17 306

　　The immune system has diverse functions and can respond delicately to hundreds of different types of infectious diseases. The initiation of the immune response after infection requires the cooperation between innate immune cells (which are pathogens that recognize general functions) and lymphocytes of the adaptive immune system (its highly variable antigen receptors can bind to specific pathogens). The activation of T lymphocytes depends on the direct activation of the innate immune system of antigen presenting cells (APCs) by pathogens, which are phagocytosed and ruminated, and then presented to activated T cells. T cells proliferate and differentiate to protect the body by activating other immune cells or killing infected cells. Among the immune cells activated by T lymphocytes, the most important thing is that B lymphocytes produce antibodies. T lymphocytes determine the type of antibodies secreted by B lymphocytes and the activity of other immune cells to guide the immune response and provide the best protection against different types of infections.

　　At the end of the immune response, most of the activated B cells and T cells will undergo apoptosis, but a small number are reserved for memory cells, which can be used for immune defense when the host is exposed to the same infection. The vaccine must work in a similar way, activating antigen-specific T cells and B cells, some of which are converted into memory cells, which will control secondary infections through the invader of the vaccine. In addition, similar to the infection itself, the vaccine must generate the best type of immune response to fight against specific pathogens.

　　A brief summary of the different methods of the immune system against antigens. T lymphocytes are divided into two categories, cytotoxic (CD8) cells and helper (CD4) cells, and their main actions. For example, viral infections can be eliminated by cytotoxic T cells (CTL) or antibodies, and the bacteria and parasites in the cells are the most effective control of cytokines produced by T cells, specifically to activate specific innate immune cells. The immune system is believed to be able to recognize different types of pathogens through the recognition of innate immune cells to microbial pathogens (PAMPs) related molecular mechanisms. The reactive cells can directly obtain adaptive responses through these pathways to better resist microbial invasion. PAMPs include, for example, the compositional characteristics of bacterial cell walls, double-stranded RNA, which is found in certain viruses, and CpG-rich DNA, which can exist in bacteria and viruses. These microbial components are detected by the inherent signal pattern recognition receptors (PRRS), the most famous and most commonly used are Toll-like receptors (TLRs). Different members of the membrane-bound receptors of the TLR family specialize in detecting different types of pathogens. In addition, many cytoplasmic proteins have recently been recognized as important substances during innate immunity, such as PRRS. The recognition of PAMPS by reactive cells promotes the recruitment of natural immune cells and antigen-presenting APCs and activates antigen-presenting APCs. Improve the absorption of antigens and induce cell surface molecules and soluble mediators, which are necessary for T cell activation. These effects affect the size of T cell and B cell responses and the number of memory cells produced. Not only can they be used to warn and activate adaptive immune response cells, but importantly, they can also control the following types of immune responses. Currently, three main types of vaccines are used in humans: live attenuated vaccines, composed of bacteria and viruses, similar to real pathogens but less pathogenic than theirs; inactivated vaccines, heat-inactivated or chemically inactivated pathogen particles ; Subunit vaccine, made from pathogen components. Vaccines do not only contain antigens (antigens are the target of an acquired immune response). However, whether molecules or other substances can amplify or affect the adaptive immune response, these are called adjuvants. In live attenuated vaccines, antigens are recognized by cells of the adaptive immune system and PAMPs can activate specialized antigen-presenting cells. Just like the pathogen itself, it also provides a natural adjuvant. By including the purified antigen on the Bia unit vaccine, lymphocytes specifically recognize it. Although they are safer than whole body vaccines, as far as we know, they are not the best vaccines to activate the immune system because they lack the recognition of intrinsic PAMPs. This vaccine requires enhanced adjuvants to enhance immunogenicity and affect the magnitude and response of natural immunity.

　　adjuvant may promote immune response by recruiting professional antigen presenting cells to the vaccination site. By increasing antigen presentation to APCs, or by activating cells to produce cytokines and provide T cell activation signals. An adjuvant that has been used for a long time in the history of human vaccines is aluminum salt (sometimes called alum). Proteins from pathogenic bacteria are adsorbed to aluminum salts, creating a suspension, which is immunized by intramuscular injection. Despite its long-term and widespread use in human vaccines, it is still unclear how it assists. Although it is generally believed that aluminum adjuvants maintain the immune response by slowly releasing antigens. It is now clear that their influence on the innate immune system is manifold. In addition, the removal of nodules caused by the use of aluminum adjuvant has no effect on the immune response, and the slow-release function of aluminum adjuvant is questioned.

　　After intentionally introducing an adjuvant into a vaccine for the first time, the results show that aluminum salts and other particles can enhance the body's immune response. Currently, few licensed vaccine adjuvants are used clinically. In the United States, aluminum salts have been used for many years, and it can be said that aluminum salts are the only adjuvant used in human vaccines. Recently, lipid monophosphate A (MPL), a derivative of lipopolysaccharide (LPS), a cell wall component of highly immunogenic bacteria, has been approved for use in GlaxoSmithKline vaccine, human papillomavirus HPV and aluminum hydroxide. Vaccine use. In Europe, some other adjuvants are used, including water and oil adjuvants MF59 and AsO3, developed by Novartis and GlaxoSmithKline, respectively. In addition to these, a large number of new vaccine adjuvants have been studied in the laboratory, and some of them have also been used in human clinical trials. Whether these adjuvants will pass the two standard general requirements (effectiveness and safety) remains to be determined.

　　Adjuvant and antibody production: Most of the current vaccines induce long-lived plasma cells-terminally differentiated B cells, which can continuously secrete antibodies for a long time. Before damage to the body occurs, antibodies move quickly to stop pathogens, or their products. Therefore they are ideal for controlling many diseases, including viral infections and toxins produced by a certain amount of bacterial products, including tetanus and diphtheria toxin. Many virus vaccines use attenuated viruses as agents and can produce good, long-lasting antibodies. However, this is not true for subunit vaccines. For example, the tetanus vaccine consists of a toxoid (an inactivated toxin that retains its antigenicity) adsorbed to an aluminum adjuvant, resulting in the production of cells that produce antibodies against tetanus toxin. However, this vaccine is routinely given to individuals every 10 to 15 years to produce specific plasma cells, which continue to produce antibodies until they die. Plasma cells infected with live measles virus are predicted to have a half-life of 3014 years. How the infection is controlled and how an adjuvant can achieve this remains unclear. The type of antibody produced is also affected by the adjuvant. There are different types of antibodies. Ideally, vaccines should be designed to induce such antibodies. This will be the most effective way to deal with pathogens. Immunoglobulin A (IgA) passes through the mucosal surface and is a very effective anti-infective. This factor may be the reason why the (live attenuated vaccine) is more effective than the Shaq (inactivated) polio vaccine. This is because the secretion of intestinal and respiratory tract IgA induced by oral live vaccines, while inactivated intramuscular injection of Shaq vaccine cannot induce its secretion. Adjuvants can be selected to increase the production of secreted IgA, possibly through their influence on the differentiation of APC and T cells.

　　Antibodies are sometimes not enough: Influenza vaccines induce antibodies against the main surface proteins of the virus and hemagglutinin and neuraminidase. In doing so, they can effectively protect the body from infection by influenza virus strains, and there are substances expressing these proteins in the vaccine. However, the changes in the two proteins, hemagglutinin and neuraminidase, act as a mutation, and the vaccine must be reconstituted annually to contain the hemagglutinin and neuraminidase of the expected strain. In addition, no adjuvant vaccine is currently used in the United States. This means that large doses of immunity must be given and it is difficult to induce immunity to proteins found in emerging strains, such as those caused by the H5N1 virus. This may be because there are memory cells that can recognize viruses but cannot recognize emerging virus strains. Memory cells respond in the absence of high levels of co-stimulation. Therefore, it can be activated without an adjuvant. However, a major response is required to prevent the emergence of new strains due to their annual influenza strain antigenic differences. In the absence of adjuvant-induced inflammation, this primary response cannot be activated. Adding adjuvants (MF59, AsO3 or aluminum salt; Table 1) to influenza vaccine can increase antibody titer and durability. However, these methods do not provide cross-reactivity to viruses of different subtypes. Attenuation of the influenza virus subunit vaccine FluMist is real, and it is modified every year, although the vaccine can activate cross-reactive CD8 + T cells, at least in children. CD8 + T cells recognize the immutable parts of the virus-for example, the core protein. And can provide more cross-reactions, which can be induced by new vaccines. In addition to influenza, there are many other infectious diseases, AIDS and malaria. For example, their antibodies are not all, or they are not enough to provide adequate protection. In these cases, humoral immunity, antibody-mediated, and cell-mediated immunity, depending on cytokine activation of immune cells, cytotoxic T cells or T cells, may have effective protection.

　　Contribution of T cell priming adjuvant: Dendritic cells (DC) are the main antigen presenting cells at the beginning of the T cell response, so it may be the main target of adjuvants to exert their effectiveness. In the absence of infection, dendritic cells are distributed throughout the tissue as phagocytes. These cells signal the existence of infection, recognize microbial components through pattern receptors (PRRs), and indirectly, recognize microbial components through inflammatory cytokines released by natural immune cells. The dendritic DCs activated by these signals undergo a process called maturation and migrate to secondary lymphoid organs, where they activate the initial T cells. DC maturation involves increasing the processed microbial protein. Some of these are presented to the major histocompatibility complex (MHC) molecules on T cells (discussed below). This is the first activation signal required. In addition, the activation of DCs by PRRS leads to the expression of so-called costimulatory molecules on the cell surface and secretion of cytokines by DCs. The costimulatory signal is the second signal, which requires DC to activate naive T cells and cytokines, provide the third signal, and directly differentiate along different pathways. One way, such as the aluminum salt adjuvant MF59 acts by promoting inflammation and DC infiltration to the vaccination site and increasing the uptake of related antigens by DCS.

　　The adjuvant effect is better understood as a signal that induces helper T cell 1 (Th1) response. It is characterized by that helper T cells can produce high levels of IFNγ, and other cytokines, which can activate the antibacterial effect at the effector site. These Th1 driving signals are known to induce the secretion of interleukin (IL) 12 through TLR, which can induce Th1 cell differentiation. Adjuvant QS21 or other saponins drive Th1 response, which is thought to work through the induction of IL-12 in DCS. Aluminum salt, but does not directly cause signals through Toll-like receptors and does not stimulate IL-12 production through DCS. On the contrary, the mechanism of aluminum adjuvant driving Th2 reaction is not well understood.

　　is the requirement for CD8 + T cells (cytotoxic cells) to present antigen, which is different from those of CD4 helper T cells. CD8+ T cells are designed to monitor antigens, such as viruses. They invade into the cytoplasm. The way from the cytoplasm to the surface of the antigen is different, and the way to distinguish the internal antigen. In short, all antigens are presented to the cell surface through the coding of MHC molecules. The internalized antigen is delivered to the cell surface through a class of MHC molecules, and MHC class II can promote the activation of CD4 cells. On the contrary, endogenous antigens reach the cell surface through different molecules, MHC class I molecules can activate CD8 cells. To activate cytotoxic T cells, DCs internalized antigens must pass through the MHC class I pathway before they appear on the cell surface. In a process called cross-presentation, the epitopes of DCs are specific. The adjuvant system may have an important influence on the process of antigen cross-presentation. For example, certain TLR ligands, such as lipopolysaccharide, can facilitate the transportation of an important part of the MHC class I antigen processing pathway to the internalized vesicle pathway, presumably to increase cross-presentation. Other adjuvants, such as immune complexes, microparticle adjuvants are composed of lipids, cholesterol and saponin adjuvant Quil A, which can promote cross-presentation and CD8 cell activation by bypassing the antigen processing pathway.

　　The inflammatory response caused by adjuvant can enhance the initiation of T cell response is an open question. For example, a key complex of the inflammatory pathway, the inflammator, is recommended for research. Instead of recruiting APCs and enhancing T and B cell responses accompanied by immune responses with aluminum adjuvants. Although a similar method is used on genetically identical mice. It is also unclear whether the PRR-mediated pathway must be activated by DCS itself, or the inflammatory response in local tissue cells can explain some adjuvant effects. For example, inflammatory cytokines can recruit monocytes to the injection site and differentiate into dendritic cells. These may be activated and migrate to the lymph nodes, presenting antigens to T cells. Such recruitment is an act of MF59, oil-in-water emulsion and aluminum salt adjuvant.

　　The establishment of cellular memory: Although after years of research, immunologists still do not know that these signals are necessary to generate memory T cells. This may be a random process in which a percentage of cells are randomly selected to survive, or there is a selective process in which cells are designated to survive and generate memory cell banks in the early stages of the reaction. The signature markers that memory cells can recognize may be useful in measuring the effectiveness of different antigen adjuvant combinations. In some cases, the production of memory cells expressing lymph node markers is related to long-term survival and thus plays a protective role. In contrast, other researchers believe that memory cells may migrate to non-lymphatic organs, where re-infection may occur, providing the most effective protection. Therefore, the measurement of protective capacity (for example, the challenge after reducing virus titer or bacterial load) is a more useful indicator of a successful vaccine than the phenotype of memory cells.

　　Many variables can affect the number and phenotype of memory cells. For example, a large dose of antigen can activate a large number of cells, but vaccines are more inclined to low doses. If it activates high-affinity receptor cells, it may be more effective for some infections. This seems to be true in the mouse model of Mycobacterium tuberculosis infection. The low dose triggers the induction of highly sensitive T cells to produce a broad cytokine response, which is related to protection. Similarly, the degree of inflammation can be changed by adding adjuvants to the vaccine. Influencing the phenotype and number of memory cells is partly due to the need for inflammatory signals for effective expansion and survival of T cells. However, by reducing the inflammatory response during the start-up process, the rate of memory cell production can be increased, resulting in faster production of memory cells. It is critical, therefore, antigen and The adjustment of the amount of adjuvant depends on how many and what type of memory cells are needed to provide protection. This brings us to the question of how important the two main types of T cells, CD4 and CD8, are in providing protection.

　　 CD4 T cell-mediated protection: It is clear that CD4 + T cells are an important factor in cellular and humoral immunity. It has been verified for many years that CD4 + T cells can help B cells. But CD4 + T cells are also crucial for producing effector CD8 memory T cells. Any vaccine, regardless of its intended purpose, must activate helper CD4 + T cells. Perhaps the most important consideration is to determine which adjuvant is used in a vaccine, and it is necessary to consider what type of CD4 + T cell response is the ideal immune response that directly follows. It is now recognized that there are at least five subgroups of CD4 + T: Th1 and Th2 cells, which activate macrophages in different ways, and induce B cells to produce different types of antibodies. Th17 cells cause inflammation; T follicular cells, which specifically activate B cells. Regulatory T cells, which are thought to prevent autoimmune diseases. These subpopulations have been extensively reviewed elsewhere. Here, we will mainly discuss Th1 cells related to post-vaccination protection. Although it is obvious that CD4 + T cells must be activated after vaccination, the importance of generating CD4 memory cells is not yet obvious. We have discussed some details recently, so we won't discuss the details here. A careful analysis of the available evidence shows that relatively little protective immune response depends on CD4 + T cell memory. However, the protection of Mycobacterium tuberculosis is a good example of how CD4 memory cells behave. The cytokine interferon (IFNγ) produced by CD4 + T cells activates macrophages to provide important protection for the lungs infected by Mycobacterium tuberculosis. The current tuberculosis vaccine, Bacillus Calmette-Guerin (BCG), provides protection for children against serious diseases. But its use by adults is limited. Therefore, many researches on tuberculosis vaccines have focused on a main stimulus method, a series of vaccines, with BCG as the main vaccine and the second experimental vaccine, aimed at reactivating and increasing the protective memory response. Tried with some substance stimulation. For example, a modified vaccinia virus (MVA) protein expressed from Mycobacterium tuberculosis, 85A, has been tested on animals and humans. With poxvirus vectors, a wide range of immune responses are generated, including IL-12 produced by DCS and IFNγ secreted by CD4 cells. In mouse research models, MVA85A reduces bacterial levels in animal models. The vaccine can also successfully promote human antigen-specific cells and subsequent memory cells to produce a series of cytokines, including interferon IFNγ and tumor necrosis factor TNFα. This multifunctional cell can produce higher levels of cells Factors have been shown to provide protection in mouse models against infections, including tuberculosis.

　　 CD8 T cell-mediated protection: Although it is difficult to prove that CD4 memory T cells have a direct protective effect, the differentiation of CD8 + T cells into CTL has always been a measure of its protective effect. After activation of the lymphatic organs and clonal expansion, CTL lymphocytes migrate to the site of inflammation, where they kill the infected cells by inducing apoptosis, thereby limiting and eventually clearing the infection. CTLS has been shown to provide protection in various mouse infection models. In vitro experiments show that human CD8 + T cells can verify CTL activity. CTL is also related to the protection of humans after influenza infection. The focus of the development of influenza virus subunit vaccines has shifted to generating memory CD8 + T cells, which can provide more cross-reactive protection. As mentioned above, this is because CD8+ T cells recognize a small amount of the virus. Several methods have been developed, the most interesting of which is the lungs where the vaccine is targeted, generating memory cells in the correct location to provide the fastest protection. For example, CD8 + T cell recognition peptide can bind to the lipid moiety, Pam-2-Cys, and activate TLR receptors on DCs to successfully protect CD8 + T cells. When delivered intranasally, the CD8 + T cells produced by this vaccine are transferred to the lungs to provide immediate protection. The use of peptide fragments instead of the entire antigen, because different fragments are recognized by T cells of different individuals, it is necessary to recognize and include a very large number of different fragments. As an alternative, the entire detergent-inactivated influenza virus can be combined with immunostimulatory complexes. It can present antigen to DCS and activate a series of natural immune cells to generate Th1 cells and CTL response. The immunostimulatory complex containing inactivated influenza virus has been used to make an intranasal vaccine, including all viral proteins and protection that can induce cross-reactivity. The CTL and antibodies required for this protection indicate that the ISCOM vaccine can induce effective cellular immunity and humoral response.

　　 Killing infected cells by CTLs and Th1 cells is an effective way to eliminate pathogens from intracellular infection. However, in some cases, such as liver infection with hepatitis B virus, interferon IFNγ produces CD8+ T cells to provide more effective protection, because there is no large number of dead virus from the host cell that can be eliminated. In the same way, the production of CD8+ T cells by interferon gamma is related to the protection of RTS and malaria vaccinators. This vaccine contains a protein from the parasite and the surface protein of the hepatitis B virus. Although the mechanism by which immune individuals resist infection is not very clear. It is believed that during malaria infection, humoral and cellular immunity against multiple antigens expressed at different stages of the parasite's life cycle is necessary. The most successful adjuvant system for malaria vaccine is AS02, which is prepared with saponin components and TLR agonist MPL in a microparticle system. It is worth noting that saponins and MPL need to induce moderate levels in the process of individual protection immunity. In contrast, using the same antigen with aluminum hydroxide adjuvant and MPL adjuvant (AS04) or oil-in-water emulsion (AS03) can induce high levels of antibodies, but cannot prevent infection. A deeper understanding of the protective body's response may help to effectively identify more effective combinations of antigen adjuvants. For example, the successful adjuvant, AS02, promotes immune response, Th1 cell differentiation and extensive antibody response. This shows that antibodies and cell-mediated immunity play an important role in defense against complex pathogens.

　　Ideal adjuvant: The immune system has a variety of mechanisms to deal with infectious organisms. A successful vaccine should activate the immune response and create a redundant protective response that can deal with pathogen mutations and escape strategies. Although live attenuated virus and bacterial vaccines can activate all immune systems, adjuvants have not reached this goal so far. By combining adjuvants, such as aluminum salts with MPL, or using DNA and bacterial or viral vector stimulation strategies, as discussed above, it has been reported that humoral immunity and cell-mediated responses can be activated. Yellow fever and smallpox (vaccinia) are very effective live virus vaccines to promote T and B memory cells and promote life-long protection.