New era for mucosal mast cells: their roles in inflammation,allergic immune responses and adjuvant development

To achieve immune homeostasis in such a harsh environment as the intestinal mucosa, both active and quiescent immunity operate simultaneously. Disruption of gut immune homeostasis leads to the development of intestinal immune diseases such as colitis and food allergies. Among various intestinal innate immune cells, mast cells (MCs) play critical roles in protective immunity against pathogenic microorganisms, especially at mucosal sites. This suggests the potential for a novel MC-targeting type of vaccine adjuvant. Dysregulated activation of MCs also results in inflammatory responses in mucosal compartments. The regulation of this yin and yang function of MCs remains to be elucidated. In this review, we focus on the roles of mucosal MCs in the regulation of intestinal allergic reaction, inflammation and their potential as a new target for the development of mucosal adjuvants.     

A tetrahedral framework nucleic acid based multifunctional nanocapsule for tumor prophylactic mRNA vaccination

Synthetic antigen-encoding mRNA plays an increasingly significant role in tumor vaccine technology owing to its antigen-specific immune-activation. However, its immune efficacy is challenged by inferior delivery efficiency and demand for suitable adjuvants. Here, we develop a novel mRNA nanovaccine based on a multifunctional nanocapsule, which is a dual-adjuvant formulation composed of cytosine-phosphate-guanine motifs loaded tetrahedral framework nucleic acid (CpG-tFNA) and an immunopeptide murine β-defensin 2 (mDF2β). This mRNA nanovaccine successfully achieves intracellular delivery, antigen expression and presentation of dendritic cells, and proliferation of antigen-specific T cells. In a tumor prophylactic vaccination model, it exerts an excellent inhibitory effect on lymphoma occurrence through cellular immunity. This mRNA nanovaccine has promising prophylactic applications in tumors and many other diseases.     

Injectable Biodegradable Chitosan‐Alginate 3D Porous Gel Scaffold for mRNA Vaccine Delivery

mRNA vaccines have proven to be more stable, effective, and specific than protein/peptide‐based vaccines in stimulating both humoral and cellular immune response. However, mRNA's fast degradation rate and low‐transfection efficiency in vivo impede its potential in vaccination. Recent research in gene delivery has focused on nonviral vaccine carriers and either implantable or injectable delivery systems to improve transgene expression in vivo. Here, an injectable chitosan‐alginate gel scaffold for the local delivery of mRNA vaccines is reported. Gel scaffold biodegradation rates and biocompatibility are quantified. Scaffold‐mediated mRNA in vivo transgene expression as well as ovalbumin antigen specific cellular and humoral immune responses are evaluated in vivo. Luciferase reporter protein expression resulting from mRNA lipoplex‐loaded gel scaffolds is five times higher than systemic injection. Compared to systemic injections of naked mRNA or mRNA:lipoplexes, elevated levels of T cell proliferation and IFN‐γ secretion are seen with in vivo scaffold‐mediated mRNA lipoplex delivery. Furthermore, a humoral response (ovalbumin antigen specific IgG levels) is observed as early as week 1 for scaffold‐mediated mRNA lipoplex delivery, while protein‐based immunization did not elicit IgG production until 2 weeks post‐injection. Results suggest that injectable scaffold mRNA vaccine delivery maybe a viable alternative to traditional nucleic acid immunization methods.     

壳聚糖及其衍生物作为疫苗佐剂的研究进展

疫苗佐剂能够增强机体对抗原的免疫应答反应或改变免疫应答反应类型,延长疫苗在体内作用时间,提高疫苗效力。壳聚糖能有效地将疫苗递送到靶抗原递呈细胞或组织,激活抗原提呈细胞,诱导产生免疫应答,促进Th1/Th2应答反应的平衡,因此,壳聚糖作为疫苗佐剂具有一定的潜力。为了解决壳聚糖在中性和碱性溶液中溶解性差,以及进一步提高其黏膜黏附性和靶向性等问题,通过对壳聚糖进行化学改性,生成一系列壳聚糖衍生物,提高其佐剂性能。本论文就近年来有关壳聚糖及其衍生物作为疫苗佐剂和递送系统在疫苗中的应用进行了综述,总结并提出了壳聚糖及其衍生物在疫苗佐剂应用领域所面临的问题以及其未来的发展方向,使读者对其有全面的了解。     

Surface display on gram positive bacteria

Heterologous surface display on Gram-positive bacteria was first described almost a decade ago and has since then developed into an active research area. Gram-positive bacterial surface display has today found a range of applications, in immunology, microbiology and biotechnology. Live bacterial vaccine delivery vehicles are being developed through the surface display of selected foreign antigens on the bacterial surfaces. In this field, "second generation" vaccine delivery vehicles are at present being generated by the addition of mucosal targeting signals through co-display of adhesins, in order to achieve targeting of the live bacteria to immunoreactive sites to thereby increase immune responses. Engineered Gram-positive bacteria are further being evaluated as novel microbial biocatalysts with heterologous enzymes immobilized as surface exposed on the bacterial cell surface. A discussion has started whether bacteria can find use as new types of whole-cell diagnostic devices since single-chain antibodies and other variants of tailor-made binding proteins can be displayed on bacteria. Bacteria with increased binding capacity for certain metal ions can be created and potential environmental or biosensor applications for such recombinant bacteria as biosorbents are being discussed. This article explains the basis of Gram-positive bacterial surface display, and discusses current uses and possible future trends of this emerging technology.     

高分子材料调控肿瘤免疫微环境的研究进展

随着肿瘤免疫疗法在临床应用取得巨大突破,通过抗肿瘤免疫反应提高抗肿瘤疗效的治疗方式受到了广泛的关注.然而,肿瘤组织存在复杂的免疫抑制性微环境,严重限制了部分免疫疗法的效果.长期以来,高分子材料作为重要的药物递送载体受到广泛关注,但是其在调控肿瘤免疫微环境的功能及应用方面尚未引起足够的重视.在本文中,我们一方面介绍了肿瘤组织形成免疫抑制性微环境的成因,如肿瘤组织存在多种免疫抑制性细胞,如调节性T细胞(Tregs)、髓系来源抑制性细胞(MDSCs)和肿瘤相关巨噬细胞(TAMs)等,以及免疫细胞、肿瘤细胞等分泌的大量细胞因子、趋化因子、代谢产物等.另一方面,重点介绍了近年来高分子材料作为载体递送免疫调节分子或发挥自身免疫调节功能,调控或逆转免疫抑制性微环境的策略和典型代表,证明了高分子材料在调控肿瘤免疫微环境,改善肿瘤治疗效果方面的巨大潜力.     

乙型肝炎疫苗预防效果探讨

目的分析研究乙型肝炎疫苗预防效果。方法选取2013年4月—2014年4月中山市三乡医院收治的220例儿童作为研究对象,220例对象全部以经过乙型肝炎疫苗接种,随机分为观察组、对照组,对照组儿童没有给予增强乙型肝炎疫苗接种,观察组儿童则在乙肝疫苗接种的基础之上给予加强乙型肝炎疫苗接种,对两组儿童乙型肝炎疫苗的预防效果进行对比。结果观察组儿童的抗-HBs阳性率和抗-HBc阳性率显著高于对照组(P0.05);观察组儿童的HBs Ag阳性率明显低于对照组(P0.05)。结论乙肝疫苗预防接种免疫人群效果良好,增强乙肝疫苗接种可以有效降低大规模疫苗接种当中不可预知因素对其预防效果的影响,进而使乙肝疫苗预防性接种效果明显提高。     

Rectal administration of nanosystems: from drug delivery to diagnostics

The rectal administration of drugs has been an enduring medical practice for either the management of local or systemic conditions. Although mostly regarded as an alternative to other delivery routes, the colorectal mucosa offers an effective pathway for enhanced systemic bioavailability of many active molecules. The fairly stable physicochemical and enzymatic environment of the mucosa and the possibility of partially avoiding the hepatic first-pass effect are some of the potential advantages of rectal drug delivery. At the same time, higher drug levels of drugs can be achieved at colorectal fluids and tissues, which can aid management of local conditions. However, problems with patient acceptability as well as poor and erratic drug absorption may impair efficient use of the rectal drug delivery route. The valuable features of nanotechnology-based systems for mucosal use are well recognized, and their potential as carriers for drug delivery has already been proven for different medical applications/delivery routes. Although still limited, the development of rectal nanomedicines with therapeutic, diagnostic, and prophylactic purposes is steadily emerging and may circumvent some of the problems associated with the more standard delivery approaches. This review discusses the rationale behind the use of nanotechnology-based strategies for rectal drug delivery and provides a critical overview on the various types of nanosystems proposed so far.     

Modulation of protective immunity against herpes simplex virus via mucosal genetic co-transfer of DNA vaccine with ��2-adrenergic agonist

Cholera toxin, which has been frequently used as mucosal adjuvant, leads to an irreversible activation of adenylyl cyclase, thereby accumulating cAMP in target cells. Here, it was assumed that β₂-adrenergic agonist salbutamol may have modulatory functions of immunity induced by DNA vaccine, since β₂-adrenergic agonists induce a temporary cAMP accumulation. To test this assumption, the present study evaluated the modulatory functions of salbutamol co-administered with DNA vaccine expressing gB of herpes simplex virus (HSV) via intranasal (i.n.) route. We found that the i.n. co-administration of salbutamol enhanced gB-specific IgG and IgA responses in both systemic and mucosal tissues, but optimal dosages of co-administered salbutamol were required to induce maximal immune responses. Moreover, the mucosal co-delivery of salbutamol with HSV DNA vaccine induced T_h2-biased immunity against HSV antigen, as evidenced by IgG isotypes and T_h1/T_h2-type cytokine production. The enhanced immune responses caused by co-administration of salbutamol provided effective and rapid responses to HSV mucosal challenge, thereby conferring prolonged survival and reduced inflammation against viral infection. Therefore, these results suggest that salbutamol may be an attractive adjuvant for mucosal genetic transfer of DNA vaccine.     

A single HBsAg DNA vaccination in combination with electroporation elicits long-term antibody responses in sheep

Vaccines continue to be the most cost effective method to reduce the burden of disease in both human and animal health. However, there is a need to improve the duration of immunity following vaccination, since maintenance of protective levels of antibody in serum or the ability to rapidly respond upon re-exposure (memory) is critical if vaccines are to provide long-term protective immunity. The purpose of this experiment was to test the duration of antibody responses and the ability to generate anamnestic responses following a single immunization with a DNA vaccine encoding hepatitis B surface antigen (HBsAg) delivered by a variety of routes. Sheep immunized with the conventional HBsAg subunit vaccine (Engerix-B) as well as sheep immunized with a HBsAg DNA vaccine, combined with electroporation, generated significant antibody responses that were sustained for 25 weeks after primary immunization. At 25 weeks, all experimental groups received a secondary immunization with the HBsAg subunit vaccine. Sheep that received a primary DNA immunization, in combination with electroporation, mounted an anamnestic response similar to the cohort immunized with the HBsAg subunit vaccine. In contrast, animals immunized with DNA vaccines administered without electroporation elicited no detectable memory response. The presence of immune memory was significantly correlated with the induction of a prolonged primary immune response. Thus, a single DNA vaccination, in combination with electroporation, approached the efficacy of the commercial subunit vaccine in the maintenance of long-term protective serum antibody titres and immune memory.     

DNA vaccines for cervical cancer: from bench to bedside

More than 99% of cervical cancers have been associated with human papillomaviruses (HPVs), particularly HPV type 16. The clear association between HPV infection and cervical cancer indicates that HPV serves as an ideal target for development of preventive and therapeutic vaccines. Although the recently licensed preventive HPV vaccine, Gardasil, has been shown to be safe and capable of generating significant protection against specific HPV types, it does not have therapeutic effect against established HPV infections and HPV-associated lesions. Two HPV oncogenic proteins, E6 and E7, are consistently co-expressed in HPV-expressing cervical cancers and are important in the induction and maintenance of cellular transformation. Therefore, immunotherapy targeting E6 and/or E7 proteins may provide an opportunity to prevent and treat HPV-associated cervical malignancies. It has been established that T cell-mediated immunity is one of the most crucial components to defend against HPV infections and HPV-associated lesions. Therefore, effective therapeutic HPV vaccines should generate strong E6/E7-specific T cell-mediated immune responses. DNA vaccines have emerged as an attractive approach for antigen-specific T cell-mediated immunotherapy to combat cancers. Intradermal administration of DNA vaccines via a gene gun represents an efficient way to deliver DNA vaccines into professional antigen-presenting cells in vivo. Professional antigen-presenting cells, such as dendritic cells, are the most effective cells for priming antigen-specific T cells. Using the gene gun delivery system, we tested several DNA vaccines that employ intracellular targeting strategies for enhancing MHC class I and class II presentation of encoded model antigen HPV-16 E7. Furthermore, we have developed a strategy to prolong the life of DCs to enhance DNA vaccine potency. More recently, we have developed a strategy to generate antigen-specific CD4(+) T cell immune responses to further enhance DNA vaccine potency. The impressive pre- clinical data generated from our studies have led to several HPV DNA vaccine clinical trials.     

A Rapidly Dissolvable Microneedle Patch with Tip-Accumulated Antigens for Efficient Transdermal Vaccination

Dissolvable microneedles (DMNs) are an attractive alternative for vaccine delivery due to their user-friendly, skin-targeted, and minimally invasive features. However, vaccine waste and inaccurate dosage remain significant issues faced by DMNs, as the skin's elasticity makes it difficult to insert MNs completely. Here, a simple and reliable fabrication method are introduced based on two-casting micromolding with centrifugal drying to create a rapidly DMN patch made of hyaluronic acid. Ovalbumin (OVA), as the model antigens, is concentrated in the tip parts of the DMNs (60% of the needle height) to prevent antigen waste caused by skin elasticity. The time and temperature of the initial centrifugal drying significantly affect antigen distribution within the needle tips, with lower temperature facilitating antigen accumulation. The resulting DMN patch is able to penetrate the skin with enough mechanical strength and quickly release antigens into the skin tissue within 3 min. The in vivo study demonstrates that immunization of OVA with DMNs outperforms conventional vaccination routes, including subcutaneous and intramuscular injections, in eliciting both humoral and cellular immunity. This biocompatible DMN patch offers a promising and effective strategy for efficient and safe vaccination.     

Functional polymer/inorganic hybrid nanoparticles for macrophage targeting delivery of oligodeoxynucleotides in cancer immunotherapy

To efficiently deliver CpG oligodeoxynucleotides (ODN) in cancer immunotherapy, a multifunctional macrophage targeting delivery system was designed and prepared. Mannosylated carboxymethyl chitosan/protamine sulfate/CaCO₃/ODN (MCMC/PS/CaCO₃/ODN) nanoparticles were prepared using a facile self-assembly method. The functional components, including MCMC to endow the nanoparticles with macrophage targeting ability, PS to improve the ODN loading capacity and enhance the cell uptake, and CaCO₃ to encapsulate ODN and induce the favorable pH sensitivity, were introduced to the delivery systems by self-assembly. Due to the mannose mediated endocytosis and the favorable effects of PS in overcoming delivery barriers, MCMC/PS/CaCO₃/ODN nanoparticles exhibit a much higher ODN delivery efficiency and a significantly enhanced immune stimulation capacity as compared with Lipofectamine 2000/ODN complexes. The regulation of NF-κB activity by our ODN delivery system results in dramatically increased production of proinflammatory cytokines including IL-12, IL-6, and TNF-α in RAW264.7 cells. The significantly increased CD80 expression after stimulation by the ODN delivery systems indicates the successful modulation of the macrophage polarity to the anti-tumor M1 phenotype. The multifunctional macrophage targeting delivery system developed has promising applications in delivery of CpG ODN in cancer immunotherapy.     

Mucosal dendritic cells shape mucosal immunity

Dendritic cells (DCs) are key modulators that shape the immune system. In mucosal tissues, DCs act as surveillance systems to sense infection and also function as professional antigen-presenting cells that stimulate the differentiation of naive T and B cells. On the basis of their molecular expression, DCs can be divided into several subsets with unique functions. In this review, we focus on intestinal DC subsets and their function in bridging the innate signaling and adaptive immune systems to maintain the homeostasis of the intestinal immune environment. We also review the current strategies for manipulating mucosal DCs for the development of efficient mucosal vaccines to protect against infectious diseases.     

Nanovaccines for cancer immunotherapy: Current knowledge and future perspectives

Cancer immunotherapy harnesses the immune system to attack tumors and has received extensive attention in recent years. Cancer vaccines as an important branch of immunotherapy are designed for delivering tumor antigens to antigen-presenting cells (APCs) to stimulate a strong immune response to against tumors, representing a potentially therapeutic and prophylactic effect with the long-term anti-cancer benefits. Nevertheless, the disappointing outcomes of their clinical use might be attributed to dilemma in antigen selection, immunogenicity, lymph nodes (LNs) targeting ability, lysosomal escape ability, immune evasion, etc. Nanotechnology, aiming to overcome these barriers, has been utilized in cancer vaccine development for decades. Numerous preclinical and clinical studies demonstrate positive results in nanomaterials-based cancer vaccines with considerable improvement in the vaccine efficacy. In this review, we systematically introduced the characteristics of nanovaccines and highlighted the different types of nanomaterials used for cancer vaccine design. In addition, the opportunities and challenges of the emerging nanotechnology-based cancer vaccines were discussed.     

Catalytic immunotherapy-photothermal therapy combination for melanoma by ferroptosis-activating vaccine based on artificial nanoenzyme

Tumor cell vaccine is a promising approach for cancer therapy to activate tumor immune, which can be achieved by tumor cells immunogenic cell death (ICD), converting in situ tumors into endogenous vaccination strategy, and ferroptosis has been proved to induce ICD occurrence. Ferroptosis is triggered by artificial nanoenzyme copper telluride mimicking peroxidase and glutathione oxidase, based on which the ferroptosis-activating vaccine (termed as CM CTNPs@OVA) was designed and established for catalytic immunotherapy. Owing to photothermal effect of copper telluride, photothermal therapy (PTT) was combined for an intensive cancer therapeutic effect. CM CTNPs@OVA was composed of solid mesoporous copper telluride nanoparticles, ovalbumin (OVA) loaded in mesoporous, and melanoma cell membrane coating surface. In in vitro and in vivo investigations, CM CTNPs@OVA, with particle size of 113.7 ± 1.7 nm, was certified to release copper ions for ferroptosis initiation, and OVA directly maturated dendritic cell (DC) as exogenous antigens extracellularly. ICD was then induced by ferroptosis pathway and PTT to release damage-associated molecular patterns for DC maturation and subsequent T cells recruitment. CM CTNPs@OVA-treated melanoma with exited inhibition rate, proving that the strategy of catalytic immunotherapy-PTT combination by ferroptosis-activating vaccine possessed massive potential for melanoma therapy based on nanoenzyme copper telluride.     

Development and Optimization of Chitosan Nanoparticle-Based Intranasal Vaccine Carrier

Chitosan is a natural polysaccharide, mainly derived from the shell of marine organisms. At present, chitosan has been widely used in the field of biomedicine due to its special characteristics of low toxicity, biocompatibility, biodegradation and low immunogenicity. Chitosan nanoparticles can be easily prepared. Chitosan nanoparticles with positive charge can enhance the adhesion of antigens in nasal mucosa and promote its absorption, which is expected to be used for intranasal vaccine delivery. In this study, we prepared chitosan nanoparticles by a gelation method, and modified the chitosan nanoparticles with mannose by hybridization. Bovine serum albumin (BSA) was used as the model antigen for development of an intranasal vaccine. The preparation technology of the chitosan nanoparticle-based intranasal vaccine delivery system was optimized by design of experiment (DoE). The DoE results showed that mannose-modified chitosan nanoparticles (Man-BSA-CS-NPs) had high modification tolerance and the mean particle size and the surface charge with optimized Man-BSA-CS-NPs were 156 nm and +33.5 mV. FTIR and DSC results confirmed the presence of Man in Man-BSA-CS-NPs. The BSA released from Man-BSA-CS-NPs had no irreversible aggregation or degradation. In addition, the analysis of fluorescence spectroscopy of BSA confirmed an appropriate binding constant between CS and BSA in this study, which could improve the stability of BSA. The cell study in vitro demonstrated the low toxicity and biocompatibility of Man-BSA-CS-NPs. Confocal results showed that the Man-modified BSA-FITC-CS-NPs promote the endocytosis and internalization of BSA-FITC in DC2.4 cells. In vivo studies of mice, Man-BSA-CS-NPs intranasally immunized showed a significantly improvement of BSA-specific serum IgG response and the highest level of BSA-specific IgA expression in nasal lavage fluid. Overall, our study provides a promising method to modify BSA-loaded CS-NPs with mannose, which is worthy of further study.     

Increased in vivo immunological potency of HB-110, a novel therapeutic HBV DNA vaccine, by electroporation

Pulse-induced permeabilization of cellular membranes, generally referred to as electroporation (EP), has been used for years as a tool to increase macromolecule uptake in tissues, including nucleic acids, for gene therapeutic applications, and this technique has been shown to result in improved immunogenicity. In this study, we assessed the utility of EP as a tool to improve the efficacy of HB-110, a novel therapeutic DNA vaccine against chronic hepatitis B, now in phase 1 of clinical study in South Korea. The potency of HB-110 in mice was shown to be improved by EP. The rapid onset of antigen expression and higher magnitude of humoral and cellular responses in electric pulse-treated mice revealed that EP may enable a substantial reduction in the dosage of DNA vaccine required to elicit a response similar in magnitude to that achievable via conventional administration. This study also showed that EP-based vaccination at 4-week-intervals elicited a cellular immune response which was about two-fold higher than the response elicited by conventional vaccination at 2-week intervals. These results may provide a rationale to reduce the clinical dose and increase the interval between the doses in the multidose vaccination schedule. Electric pulsing also elicited a more balanced immune response against four antigens expressed by HB-110: S, preS, Core, and Pol.     

Synthesis of DNP-modified GM3-based anticancer vaccine and evaluation of its immunological activities for cancer immunotherapy

Tumor-associated carbohydrate antigens (TACAs) are attractive targets for vaccine development. In this context, we described a strategy combining artificial TACA and glycoengineering for cancer vaccine development. A 2,4-ditrophenyl (DNP)-modified GM3 intermediate was synthesized chemoenzymatically and conjugated to keyhole limpet hemocyanin (KLH), and the resulting bioconjugate was tested for its potential as a vaccine candidate. Mice immunological studies revealed that the DNP-modified GM3 (GM3-NHDNP) analog elicited strong and rapid immune responses by recruiting anti-DNP antibodies to facilitate the targeted delivery of the vaccine construct to antigen processing cells (APCs). Moreover, the endogenously produced anti-DNP antibodies, together with the elicited antibodies against GM3-NHDNP, may synergistically promote tumor binding and cancer cell death when the cancer cell surfaces are glycoengineered to express the GM3-NHDNP antigen.     

Designing polymeric particles for antigen delivery

By targeting dendritic cells, polymeric carriers in the nano to lower micron range constitute very interesting tools for antigen delivery. In this critical review, we review how new immunological insights can be exploited to design new carriers allowing one to tune immune responses and to further increase vaccine potency (137 references).