微生物学报

Oral delivery vehicles	Drug loading mode	Characteristic
Bacillus surface display system	Recombination surface display	No need to produce antigens; different antigens can be displayed
	Non-recombinant surface display	Highly efficient; exhibits antigen in its natural form; requires cleavage to exhibit antigen
Yeast microcapsule system	Electrostatic adsorption	Protects drugs from destruction; accurately targets drugs; efficiently loads all types of drugs; safe and non-toxic
	Non-covalent force modification
	Passive diffusion/hydrophobicity
	Layer-by-layer synthesis (LBL)
	Post-derivatization combination
Recombinant probiotic expression system	Surface display	Stable binding; no need to penetrate cells; high efficiency
	Secrete	High delivery efficiency; avoids drug toxicity to cells
	Intracellular	Avoids environmental destruction of drugs; toxic to cells
Bacterium-like particles, BLPs	Surface display	Safe and stable; high drug loading; insignificant therapeutic immunization effect
Bacterial ghosts, BGs	Anchors and fills in internal spaces	Safe and stable; efficient targeting; its structure decreases drug concentration

Oral delivery vehicles	Drug loading mode	Characteristic
Bacillus surface display system	Recombination surface display	No need to produce antigens; different antigens can be displayed
	Non-recombinant surface display	Highly efficient; exhibits antigen in its natural form; requires cleavage to exhibit antigen
Yeast microcapsule system	Electrostatic adsorption	Protects drugs from destruction; accurately targets drugs; efficiently loads all types of drugs; safe and non-toxic
	Non-covalent force modification
	Passive diffusion/hydrophobicity
	Layer-by-layer synthesis (LBL)
	Post-derivatization combination
Recombinant probiotic expression system	Surface display	Stable binding; no need to penetrate cells; high efficiency
	Secrete	High delivery efficiency; avoids drug toxicity to cells
	Intracellular	Avoids environmental destruction of drugs; toxic to cells
Bacterium-like particles, BLPs	Surface display	Safe and stable; high drug loading; insignificant therapeutic immunization effect
Bacterial ghosts, BGs	Anchors and fills in internal spaces	Safe and stable; efficient targeting; its structure decreases drug concentration

Type	Exogenous proteins/drugs	Anchoring proteins/attachment	Applications	References
Recombination	CsPmy	CotC	Oral vaccine for protection of grass carp against Ascaris lumbricoides	[42]
	Glucagon-like peptide-1 (GLP-1)	CotC	GLP-1 (28−36) oral delivery system for the treatment of type 2 diabetes mellitus	[35]
Non-recombinant	Grass carp reovirus (GCRV) Vp7 antigen	CotB & CotC	Enhancement of protective immunity to GCRV in grass carp	[43]
	OmpK or green fluorescence protein (GFP)	CotY	Enhancement of zebrafish immunogenicity to Vibrio pathogen infections	[44]
	Helicobacter pylori urease B	CotC	Oral administration of urease-carrying B spores prevents Helicobacter pylori infection	[45]
	Protective antigen (PA)	Adsorption	Development of an oral vaccine for the surface presentation of PA for the treatment of anthrax	[46]
	PfCSP	Coupling	Oral vaccines for malaria prevention	[47]
	TTFC	Adsorption	Effective in inducing an immune response and can be used as an adjuvant for vaccine immunization	[48]

Type	Exogenous proteins/drugs	Anchoring proteins/attachment	Applications	References
Recombination	CsPmy	CotC	Oral vaccine for protection of grass carp against Ascaris lumbricoides	[42]
	Glucagon-like peptide-1 (GLP-1)	CotC	GLP-1 (28−36) oral delivery system for the treatment of type 2 diabetes mellitus	[35]
Non-recombinant	Grass carp reovirus (GCRV) Vp7 antigen	CotB & CotC	Enhancement of protective immunity to GCRV in grass carp	[43]
	OmpK or green fluorescence protein (GFP)	CotY	Enhancement of zebrafish immunogenicity to Vibrio pathogen infections	[44]
	Helicobacter pylori urease B	CotC	Oral administration of urease-carrying B spores prevents Helicobacter pylori infection	[45]
	Protective antigen (PA)	Adsorption	Development of an oral vaccine for the surface presentation of PA for the treatment of anthrax	[46]
	PfCSP	Coupling	Oral vaccines for malaria prevention	[47]
	TTFC	Adsorption	Effective in inducing an immune response and can be used as an adjuvant for vaccine immunization	[48]

Protein expression mode	Bacterial strain type	Applications	References
Surface display	S. cerevisiae	Expression of avian influenza HA protein for use as an oral vaccine against avian influenza	[91]
	L. plantarum	Expression of Clostridium nucleatum outer membrane FomA protein inhibits IBD	[92]
	L. plantarum	Expression of Staphylococcus aureus non-toxic mutant alpha hemolysin, protective against Staphylococcus aureus lung and skin infections	[93]
	E. faecalis	Separate expression of four Eimeria aphidis proteins of Tennessee and protection against homologous infection in chickens by combined immunization	[94]
	Bifidobacterium	Expression of WT1 protein for use as an oral vaccine against oral cancer	[95]
	L. plantarum	Expression of the influenza virus antigen HA1-DCpep, which can play an immunizing role against influenza virus	[96]
	L. casei	Expression of AHA1-CK6 and VP2 proteins and oral immunization induces prevention of infectious pancreatic necrosis in rainbow trout	[97]
	L. casei	Expression of DCpep, Col, and CoE antigenic fusion proteins effectively induces an immune response against porcine epidemic diarrhea virus	[98]
	L. casei	Expression of an OmpK fusion CTB protein for use as an adjuvant against Vibrio anisopliae infection in crucian carp	[99]
Secrete	P. pentosaceus	Secretion of therapeutic protein P8 for tumor suppression and regulation of intestinal microflora	[100]
	L. paragasseri	Express and efficiently secrete epitope-4	[101]
	B. subtilis	Expression of the multi-epitope protein OmpC-FliC-SopF-SseB-IL-18 induces a strong immune response against Salmonella enteritidis	[102]
	L. lactis	Expression of extracellular TGFβR2 with significant anti-hepatic fibrosis effects	[103]
	L. lactis	Expression of soluble CD80 enhances anti-tumor immunity to inhibit tumor growth	[104]
Intracellular	L. casei	Expression of an antigen directed against SARS-CoV-2 Omicron variant B.1.1.529	[105]
	L. lactis	Expression of heat shock protein 65 and inhibition of atherosclerosis	[106]
	L. lactis	Expression of Helicobacter lipoprotein Lpp20 produces significant immune effects	[107]

Protein expression mode	Bacterial strain type	Applications	References
Surface display	S. cerevisiae	Expression of avian influenza HA protein for use as an oral vaccine against avian influenza	[91]
	L. plantarum	Expression of Clostridium nucleatum outer membrane FomA protein inhibits IBD	[92]
	L. plantarum	Expression of Staphylococcus aureus non-toxic mutant alpha hemolysin, protective against Staphylococcus aureus lung and skin infections	[93]
	E. faecalis	Separate expression of four Eimeria aphidis proteins of Tennessee and protection against homologous infection in chickens by combined immunization	[94]
	Bifidobacterium	Expression of WT1 protein for use as an oral vaccine against oral cancer	[95]
	L. plantarum	Expression of the influenza virus antigen HA1-DCpep, which can play an immunizing role against influenza virus	[96]
	L. casei	Expression of AHA1-CK6 and VP2 proteins and oral immunization induces prevention of infectious pancreatic necrosis in rainbow trout	[97]
	L. casei	Expression of DCpep, Col, and CoE antigenic fusion proteins effectively induces an immune response against porcine epidemic diarrhea virus	[98]
	L. casei	Expression of an OmpK fusion CTB protein for use as an adjuvant against Vibrio anisopliae infection in crucian carp	[99]
Secrete	P. pentosaceus	Secretion of therapeutic protein P8 for tumor suppression and regulation of intestinal microflora	[100]
	L. paragasseri	Express and efficiently secrete epitope-4	[101]
	B. subtilis	Expression of the multi-epitope protein OmpC-FliC-SopF-SseB-IL-18 induces a strong immune response against Salmonella enteritidis	[102]
	L. lactis	Expression of extracellular TGFβR2 with significant anti-hepatic fibrosis effects	[103]
	L. lactis	Expression of soluble CD80 enhances anti-tumor immunity to inhibit tumor growth	[104]
Intracellular	L. casei	Expression of an antigen directed against SARS-CoV-2 Omicron variant B.1.1.529	[105]
	L. lactis	Expression of heat shock protein 65 and inhibition of atherosclerosis	[106]
	L. lactis	Expression of Helicobacter lipoprotein Lpp20 produces significant immune effects	[107]

基于益生菌口服药物递送系统的研究进展

PDF下载

李文 , 刘夫锋 ^*

微生物学报 | 综述 2024,64(11): 4086-4105

收起

微生物学报 | 综述 2024, 64(11): 4086-4105

基于益生菌口服药物递送系统的研究进展

全屏

李文, 刘夫锋^*

作者信息

天津科技大学生物工程学院, 天津 300457

Research progress in oral delivery systems with probiotics as carriers

Wen LI, Fufeng LIU^*

Affiliations

College of Biotechnology, Tianjin University of Science and Technology, Tianjin 300457, China

出版时间: 2024-09-30 doi: 10.13343/j.cnki.wsxb.20240307

文章导航

摘要

收起

随着当今人们对健康的不断重视，益生菌因其安全、有益于肠道健康的特点逐渐进入人们的视野，并成为食品、医药等领域的热点。然而在口服给药方面，选择使用一种安全、方便、稳定的载体是一个难题。以益生菌为载体的口服给药系统具有优异的安全性和稳定性，同时又可以保护被递送药物通过复杂的体内环境而不受破坏。本文综述了5种常见的基于益生菌的口服药物递送方式：芽孢表面展示、酵母微胶囊、重组益生菌表达、细菌样颗粒和细菌影，并详细介绍了它们各自的结构和优缺点，为口服递送载体的开发奠定了坚实的理论基础。

关键词

口服给药 / 益生菌 / 芽孢 / 递送载体

Abstract

收起

With the increasing emphasis on health, probiotics have garnered increasing attention due to their safe and beneficial features for intestinal health and have become a hot spot in the fields of food and medicine. In terms of oral drug delivery, it is difficult to select a safe, convenient, and stable carrier. Probiotics can be used as carriers to deliver drugs through the complex in vivo environment without damage and have excellent safety and stability. This review describes the structures, advantages, and disadvantages of five common probiotic-based oral drug delivery approaches: display on spore surface, yeast microcapsules, recombinant probiotic expression, bacterium-like particles, and bacterial ghost, aiming to provide options for the application of oral delivery carriers.

Key words

oral drug delivery / probiotics / spore / delivery carrier

引用本文

李文, 刘夫锋. 基于益生菌口服药物递送系统的研究进展. 微生物学报, 2024 , 64 (11) : 4086 -4105 . DOI: 10.13343/j.cnki.wsxb.20240307

Wen LI, Fufeng LIU. Research progress in oral delivery systems with probiotics as carriers[J]. Acta Microbiologica Sinica, 2024 , 64 (11) : 4086 -4105 . DOI: 10.13343/j.cnki.wsxb.20240307

正文

收起

2001年，世界卫生组织(World Health Organization, WHO)首次将益生菌定义为“摄入足够的量时可以给予宿主健康益处的一类活的微生物”^[1]。益生菌包括许多不同种属的微生物，例如乳酸杆菌(Lactobacillus)、链球菌(Streptococcus)、乳球菌(Lactococcus)、双歧杆菌(Bifidobacterium)、丙酸杆菌(Propionibacterium)、益生芽孢杆菌(Bacillus)和大肠杆菌(Escherichia coli) Nissle1917等。此外，一些酵母，例如酿酒酵母(Saccharomyces cerevisiae)、卡氏酵母(Saccharomyces carlsbergensis)、布拉氏酵母(Saccharomyces boulardii)和黑曲霉(Aspergillus niger)、米曲霉(Aspergillus oryzae)等真菌也被认为是益生菌^[2]。益生菌既可以是天然存在的微生物，也可以是由于某些特殊效应而导致基因改变的微生物^[3]。过去十多年间，益生菌因其健康效益受到越来越多的关注。许多研究表明，益生菌具有调节人体肠道菌群、治疗癌症、缓解炎症性肠病、治疗胃肠道疾病等健康效益^[4-7]。时至今日，益生菌及其制剂已经广泛应用于食品、医药、农业、渔业和日化等领域中^[8-12]。

口服给药是传统给药方式之一，因其方便性和优秀的患者依从性，口服给药已经成为许多疾病治疗方式的主要给药途径^[13-14]。尽管如此，许多药物存在着稳定性差、溶解度低和靶向作用弱等问题，直接口服的生物利用度较低、治疗效果较差^[15]。为应对此困境，口服给药通常会使用相关载体，不仅可以提高口服药物的稳定性，增强口服药物的生物利用度和治疗效果，还可以维持药物浓度在合适的范围内，降低其副作用^[16]。因此，如何开发和利用新兴的口服递送载体已经成为了一大研究热门。伴随着纳米医药技术的进步，现在的口服递送载体的种类纷繁复杂，例如脂质体、纳米颗粒、纳米复合材料、微胶囊、树状体和细胞载体等已被广泛应用于口服药物递送^[17-20]。

益生菌因其可以定殖在肠道黏膜表面、不引起强烈的免疫反应，以及可以发挥一些积极的免疫调节功能的特点^[21]，在用作口服递送载体的方面具有很好的潜力。本课题组Wang等^[22]的研究证明了一种植物乳杆菌MA2具有很好的在肠道内定殖的能力，其可以调节肠道微生物群和阿尔茨海默病(Alzheimer’s disease, AD)代谢紊乱，能够改善AD大鼠的认知缺陷和焦虑样行为。然而，许多的益生菌，例如乳酸菌和益生芽孢杆菌、大肠杆菌Nissle1917 (EcN)等，已经被广泛开发用作各类药物递送途径的载体^[23-26]。本综述聚焦于益生菌作为口服递送载体的几种不同形式应用，如微胶囊、重组载体、芽孢表面展示、细胞样颗粒和细菌影，并介绍各装载药物模式与其特点(表1)。

1 芽孢杆菌表面展示系统

收起

芽孢杆菌是一类具有较大尺寸、需氧或兼性厌氧的革兰氏阳性菌，其最大的特点是能够产生无代谢活性的芽孢^[27]。芽孢，又称为内生孢子，其可以在休眠状态下长期存活，并且可以抵抗高温、缺水、缺乏营养、高酸碱度和有毒化学物质等极端恶劣条件^[28-30]。

芽孢的结构在芽孢的抗性中起着关键的作用。如图1所示，芽孢的结构内向外看依次为核芯、内膜、皮层、外膜、芽胞内壳、芽孢外壳和芽孢外壁(但并非所有种类的芽孢都有芽孢外壁)。芽孢的结构与其抗性的具体关系在Setlow的研究中已经进行了详细的综述^[31]。因其特殊结构与优秀的抗性，使得芽孢可以在口服后穿过胃部并在肠道中存活和发芽^[32]。芽孢杆菌具有较大的疏水性比表面积，因此可以通过吸附或共价结合的方式装载大量的疏水性药物^[33]。芽孢杆菌的芽孢不仅易于进行基因改造，而且不具有致病性^[34-35]。基于上述的特性，使得芽孢杆菌的芽孢成为一种良好的口服药物递送平台。

芽孢杆菌表面展示系统可以分为重组芽孢杆菌表面展示型和非重组芽孢杆菌表面展示型。重组芽孢杆菌表面展示型是通过将目标基因和编码芽孢衣壳蛋白(称为锚蛋白)的基因导入其自身的启动子中，然后宿主菌株在苛刻的培养环境中诱导芽孢形成，最后在宿主菌株芽孢表面表达外源蛋白或多肽^[36]。Yang等^[37]将衣壳蛋白CotB)和曲霉毒素的甘油醛-3-磷酸脱氢酶(glyceraldehyde-3-phosphate dehydrogenase of H. contortus, HcGAPDH)的基因融合进质粒pDG364中，使得HcGAPDH在枯草芽孢杆菌芽孢表面成功表达。Chen等^[38]通过同源双交叉重组将由锚蛋白CotB和冠状病毒多肽HR2编码基因组成的融合基因通过质粒pDG364插入到枯草芽孢杆菌中，成功设计出一种芽孢表面展示HR2肽的猫冠状病毒(feline coronavirus, FCoV)疫苗。Sun等^[34]将华支睾吸虫副肌球蛋白CsPmy的编码序列克隆到PEB03-CotC质粒里，并将其转化到枯草芽孢杆菌WB600菌株中，成功构建出一个CsPmy芽孢表面展示平台B.s-CotC-CsPmy。

非重组芽孢杆菌表面展示型是通过芽孢表面与外源蛋白的静电与疏水作用力或通过交联剂将外源蛋白吸附于芽孢表面^[39]。宋庆龄^[40]在芽孢表面修饰脱氧胆酸(deoxycholic acid, DA)并装载盐酸阿霉素(doxorubicin hydrochloride, DOX)和索拉菲尼(sorafenib, SOR)，构建出一种口服自主纳米药物“生成器”。Yin等^[33]将姜黄素分别与芽孢杆菌的外壳和叶酸共价连接，开发出了一种新型口服结肠靶向载体(SPORE-CUR-FA)用于治疗结肠癌。值得注意的是，灭活的芽孢也可以稳定表面吸附蛋白质，其免疫保护效果仍与未灭活芽孢相当。Song等^[41]研究发现，灭活的芽孢仍然能够与甲型流感(H5N1)病毒粒子结合，并且可以有效增强口服后产生的免疫反应。表2列出了更多重组和非重组的芽孢杆菌表面展示系统的应用。

芽孢杆菌表面展示系统因为芽孢的结构与特性，在作为口服递送载体时具有显著的优势：(1) 芽孢耐酸、耐高温等对于极端条件的抗性，使得芽孢作为口服递送载体具有很好的稳定性，能够在胃肠道环境中稳定存在而不会被破坏并将药物递送至指定部位^[49]；(2) 现在广泛应用于益生菌制剂、膳食补充剂的几种芽孢杆菌具有极高的安全性，这对口服递送载体是必需的^[50]；(3) 芽孢的芽胞壳成分与融合的抗原不需要经过细胞壁的转位步骤就可以将重组细胞暴露在表面，克服了基于细胞的展示系统的尺寸限制问题^[51]；(4) 基于芽孢外壳的紧密性和外壁的不必要性，一些大型蛋白可以展示在芽孢表面而不会影响芽孢的结构、抗性或萌发^[36]；(5) 外源蛋白或多肽可以稳定地展示在芽孢表面，使酶能够更大程度地作用于底物，并且由于芽孢的抗性提高了融合蛋白在复杂环境中的活性与稳定性，增强免疫作用^[52]。

重组和非重组芽孢杆菌表面展示系统也各自有其优缺点。重组型的一大明显优势是无需生产和纯化抗原，因为重组芽孢杆菌中包含与芽孢锚蛋白融合的抗原编码基因，所以芽孢杆菌细胞能够直接生产出需要展示的抗原，从而可以降低成本和简化生产过程^[53]。

重组型的另一个优点是可以在同一芽孢表面上展示不同的抗原或使用表达单一抗原的芽孢混合物进行免疫。Dong等^[54]将幽门螺杆菌中性粒细胞激活蛋白(HP neutrophil-activiting protein, HP-NAP)和霍乱毒素B (cholera toxin B, CTB)在枯草芽孢杆菌芽孢表面融合共表达，在研究中展现出了比单一蛋白表达载体更好的炎症抑制效果。非重组型的优点在于其较高的效率，Vetráková等^[55]使用枯草芽孢杆菌芽孢分别重组表面展示和吸附SARS-CoV-2穗状糖蛋白的受体结合域(receptor-binding domain, RBD)，而免疫荧光显微镜结果显示表面吸附RBD芽孢的荧光强度显著高于重组展示RBD芽孢，证实了非重组型的高效率。

此外，非重组型的另一个优势是其益于抗原以天然构象进行展示。大肠杆菌不耐热毒素(heat-labile enterotoxin B, LTB)是一种五聚体，其只有天然形式才具有活性功能，当LTB在芽孢表面以融合蛋白形式表达时只能以单体的形式展示，而通过吸附在芽孢表面表达时则可以以天然五聚体的形式展示^[56]。重组型展示的成功取决于锚蛋白和目标蛋白的融合，所以重组型最大的局限性在于为特定的基因选择合适的锚蛋白。Hinc等^[57]报道了使用CotB和CotC作为锚蛋白展示金线莲UreA失败，而使用衣壳蛋白CotG为锚蛋白则展示成功。

非重组型的缺点在于芽孢展示的目标蛋白不在芽孢表面，只有芽孢在体内发芽或裂解后才能展示给免疫细胞；芽孢吸附的分子机制尚未完全明确，目标蛋白会无序地在芽孢表层积聚，从而影响载体精确结构的构建^[53]。

2 酵母微胶囊系统

收起

酵母是一类单细胞真菌。酵母及其生物制剂已经广泛应用在食品工业、生物医药等领域^[58-59]。酵母微囊从酵母中提取，其成分主要是β-葡聚糖，形状呈椭圆形，具有约2−5 μm的中空多孔结构^[60] (图2)。β-葡聚糖是酵母细胞壁的主要结构成分，在结构上主要由β-1, 3-葡聚糖的主链和β-1, 6-葡聚糖的侧链组成，其单糖组成为葡萄糖^[61]。其中β-1, 3-葡聚糖既构成了葡萄糖刚性网络结构，也为酵母微囊提供了免疫活性^[62]。β-1, 3-葡聚糖是真菌的主要病原体相关分子模式，它能够被肠道免疫细胞通过多种模式识别受体(包括补体受体3和Dectin-1受体)识别出来，从而引起相关免疫反应^[63]。

酵母微囊由β-葡聚糖构成了一个内部中空、表面多孔且带负电荷的结构，因此大多数的药物都可以在酵母微囊的内部被封装或者吸附在酵母微囊表面，有利于药物的递送^[64]。随着研究的不断发展，酵母微囊的药物装载策略逐渐增多，根据刘瑛琪等^[64]和Wu等^[61]对酵母口服载体的综述，主要的药物装载策略有5种。

(1) 通过静电吸附作用将带正电的药物装载到酵母微囊内部。例如，Zhou等^[65]制备了带有正电荷的顺铂(cisplatin, CDDP)前体纳米粒(PreCDDP)，并将其封装到酵母微囊中，开发了一种安全有效的口服顺铂药物递送平台。Sabu等^[66]通过在酵母微囊表面负载带有相反电荷的胰岛素，并用海藻酸盐进行包被，成功实现了胰岛素的口服递送。

(2) 对于带负电荷的药物，可以通过非共价作用力修饰带正电荷的载体材料，再通过静电吸附作用装载到酵母微囊中。Feng等^[60]首先将小檗碱(berberine, BBR)、表没食子儿茶素没食子酸酯(epigallocatechin gallate, EGCG)和Mn²⁺通过非共价分子间作用力自组装成纳米颗粒(BBR/MPN NPs)，而后进一步装载到酵母微囊中。王大壮^[67]将DOX装载在3-巯基丙酸(3-mercaptopropionic acid, MPA)修饰的纳米氧化锌(ZnO NPs)上，构建了带负电荷的载药纳米粒(MPA-ZnO-DOX)，再用聚乙烯亚胺(polyethyleneimine, PEI)将酵母微囊内部修饰成正电荷，使MPA-ZnO-DOX能够装载在酵母微囊中。

(3) 可以利用被动扩散或疏水作用将不带电的药物修饰进酵母微囊中。Shi等^[68]通过被动扩散将牛血清蛋白(bovine serum albumin, BSA)包封在被壳聚糖包被的酵母微囊里。除此之外，咖啡因、维生素B₁₂等都可以通过相互孵育利用被动扩散装载到酵母微囊中^[69]。

(4) 部分药物还可以通过逐层合成(LBL)进行装载。Tan等^[70]通过静电相互作用将带有相反电荷的多糖依次沉积在预先装载花青素的酵母微囊上，制备出用于花青素包封和稳定的口服递送载体。He等^[71]将替莫唑胺(temozolomide, TMZ)装载在聚乳酸-羟基乙酸共聚物(poly (lactic-co-glycolic acid), PLGA)中，并通过逐层合成依次包裹O⁶-苄基鸟嘌呤(O⁶-benzylguanine, O⁶-BG)接枝的壳聚糖(BG-chitosan, BG-CS)层和酵母微囊，最终获得了一个稳定的TMZ口服递送平台。

(5) 某些药物还可以将其与衍生化的酵母微囊进行结合。Soto等^[72]使用原位合成酵母微囊包覆镍纳米颗粒，将带有His-tag的Cda2蛋白通过非共价作用力装载进酵母微囊中。Pan等^[73]使用NaIO₄对酵母微囊进行氧化，使其具有醛基，进而与卵清蛋白(ovalbumin, OVA)的伯胺基反应，将OVA通过化学连接到酵母微囊表面，从而在体内产生有效的特异性免疫反应。

酵母微囊作为口服递送载体具有诸多优势。首先，因为人体胃肠道中缺乏可以降解β-葡聚糖的酶，而且β-葡聚糖具有耐酸的特性，所以以β-葡聚糖为主要成分的酵母微囊可以避免在恶劣的肠道环境中被消化降解，保护药物可以顺利通过复杂的肠道环境而不被破坏^{[60-61, 74]}。其次，β-葡聚糖是一种来源于酵母细胞的载体，因此可以模仿酵母的行为，被M细胞上的Dectin-1识别，从而通过复杂的巨噬细胞介导运动，将各种药物准确、安全地靶向地送到病变部位^[75-76]。

酵母微囊对极性和非极性的小分子具有良好的渗透性，而且可以通过调节表面紧密连接程度促进对生物大分子的吸收，所以可以有效地装载各类大分子和小分子的药物，例如蛋白质、DNA、抗原、姜黄素等^{[63, 68, 77]}。最后，酵母微囊具有无细胞毒性以及安全的特点，这是作为口服递送载体的一大根本要求^[78-79]。

3 重组益生菌表达系统

收起

益生菌具有安全、可以进行基因工程改造的特点，伴随着分子生物学和基因工程技术的不断发展，现在工程化益生菌已经广泛应用于各类工业生产中^[80-81]。在此前的研究中，重组益生菌是通过基因编辑等技术将特定的基因重组进益生菌中，使得益生菌可以表达所需要的特定产物^[82]。随着对重组益生菌研究的不断深入，许多种属的益生菌已经被研究改造成各类外源基因的重组表达载体，除了前文提到的芽孢杆菌、酵母等，岳梦云^[83]将乳酸乳球菌改造成能够表达胰高糖素样肽-1 (glucagon-like peptide-1, GLP-1)蛋白的载体，用以治疗帕金森病；赵瑞^[84]工程化大肠杆菌Nissle1917菌株，使其能够异源表达尿酸代谢基因和提高氧气利用率基因，以实现高效地降解尿酸；Mathipa-Mdakane等^[85]还综述了鼠李糖乳杆菌作为工程益生菌的应用。

用作重组载体的益生菌的种类繁多，不同种类的重组益生菌表达外源蛋白的方式也不尽相同。乳酸菌是一种具有非致病性和非定殖性的益生菌，已被详细研究且越来越多地被用作外源蛋白的生产宿主^[86]。重组乳酸菌所携带的外源蛋白可以以3种形式表达，包括在细胞质中表达、锚定在细胞壁上表面展示和分泌到细胞外部^[87] (图3)。Muñoz等^[88]以乳酸乳球菌为载体，构建了能在大西洋鲑体内生产和释放Ⅰ型干扰素(interferon-la, IFN-la)的工程益生菌，并测定发现IFN-la在细胞质中表达。Huang等^[89]通过将非洲猪瘟病毒(African swine fever virus, ASFV)编码的p14.5基因插入到载体质粒pLP-S中，使p14.5蛋白成功锚定在植物乳杆菌的S层蛋白中，从而将p14.5蛋白表面展示在植物乳杆菌上。Zhao等^[90]克隆了来自嗜水气单胞菌的Aha1基因，并使其在干酪乳杆菌中分泌表达，而口服重组干酪乳杆菌的鲤鱼体内抗体水平显著提高。更多关于不同种益生菌表达外源蛋白的附加信息总结在表3中。

不同的外源蛋白表达方式也各有其优势。在细胞质中表达的优点在于可以避免外源蛋白在复杂的胃肠道环境中被水解破坏，因为只有当宿主菌株裂解之后外源蛋白才会被释放出来^[108]。Yang等^[109]构建了能够在细胞质内共表达SO7和DCpep基因的植物乳杆菌重组载体，与对照组相比展现出显著的抗柔嫩艾梅尔球虫感染的效应。锚定在细胞壁上表面展示的优点是使得外源蛋白与细胞紧密地结合，充分地与黏膜接触，显著增强机体免疫反应，研究表明锚定在细胞壁上表面展示的外源蛋白通常可以激发出更强的宿主免疫反应^[91]。Ai等^[110]构建出在细胞不同位置表达Der p2的重组乳酸乳球菌表达载体，其中在细胞壁锚定的E7抗原免疫效果优于表达在细胞质内E7抗原的免疫效果。

如果将合成的外源蛋白分泌到细胞外部，则可以避免外源蛋白在细胞质内水解和外源蛋白积累对宿主细胞的毒性^[111]。Ren等^[112]通过在乳酸乳球菌的不同位置表达出花生过敏原Ara h2，发现将抗原分泌到细胞外部的疫苗的免疫调节能力显著高于其他两种方式。

重组益生菌表达外源蛋白的方式对外源蛋白的活性及其数量有明显的影响，但是不同方式各有其优劣势。在实际生产应用中应结合具体条件，应综合考虑外源蛋白类型与结构、菌株种类等因素，从而选择合适的益生菌。

4 新兴的口服递送载体：细菌样颗粒和细菌影

收起

除了上述提到了几种常见的口服递送载体之外，随着研究的不断发展，现在也出现了一些新兴的口服递送载体，下面简述两种新兴的口服递送载体：细菌样颗粒(bacterium-like particles, BLPs)表面展示系统和细菌影(bacterial ghosts, BGs)。

4.1 细菌样颗粒(BLPs)

BLPs是一种将乳酸菌经过热酸处理，使得除乳酸菌的刚性细胞壁肽聚糖(peptidoglycan，PGN)之外的成分，包括DNA、蛋白质、细胞壁等全部破坏，从而产生的无活性、与活菌形状相似的空心颗粒，也被称为革兰氏阳性菌增强基质(Gram-positive enhancer matrix, GEM)^[113]。BLPs表面展示系统包括3个部分：BLPs作为支架单元、锚定蛋白(protein anchor, PA)作为锚定单元和目标蛋白作为活性单元。目标蛋白和锚定蛋白PA在体外融合表达后，通过锚定蛋白PA紧密结合在热酸处理后的乳酸菌肽聚糖表面，实现目标蛋白的表面展示^[114] (图4)。

BLPs表面展示系统具有许多的优势：(1) 无安全顾虑且性质稳定，可在室温下贮存；(2) 目标蛋白与BLPs的结合紧密，稳定性高；(3) 较乳酸菌活菌可以更高密度地装载目标蛋白；(4) 可以诱导全身和黏膜免疫，用作免疫佐剂；(5) 可以高效地同时展示多种抗原；(6) 展示和传递目标蛋白的数量已知且恒定^[115-117]。Liu等^[118]使用GEM颗粒和重组抗原CTB-UE构成了口服疫苗CUE-GEM，并证实其可以增强抗幽门螺旋杆菌感染的天然免疫反应。Mao等^[119]将单链胰岛素SCI-59与乳酸菌BLPs颗粒结合，口服后可诱导非肥胖糖尿病(non obese diabetes, NOD)小鼠的口服耐受性并预防自身免疫性糖尿病，其效果与稳定性还优于直接使用SCI-59。

然而BLPs表面展示系统也存在着一些问题：有报道发现以BLPs作为载体的疫苗一次免疫的效果并不明显，通常需要2到3次免疫才能有一定的效果^[120]。Sudo等研究发现BLPs递送的效果与递送途径有关，而滴鼻途径的效果高于口服途径^[121]。目前，关于BLPs表面递送的研究主要集中在滴鼻递送方面，而关于口服递送方面的研究还不充足。

4.2 细菌影(BGs)

BGs是通过基因或化学方法产生的非生物空包膜，常见的是革兰氏阴性细菌，如大肠杆菌Nissle1917受到裂解基因E控制表达，在活菌包膜内形成裂解隧道结构^[122-123] (图5)。BGs不含有任何细胞质内容物，包括染色体和质粒DNA，但保留了其细胞表面形态结构：外膜、内膜和肽聚糖。因为BGs保留了完整的细胞形态和天然表面抗原结构，如病原体相关分子模式，其中包括天然细菌的脂多糖、鞭毛、黏附素等与免疫相关的刺激因子，所以BGs易于被免疫细胞，如巨噬细胞、B和T细胞等识别和捕获，能够有效地激发强大的免疫反应，无需添加额外的佐剂，表明BGs具有良好的作为疫苗的潜力^[124-125]。

BGs的空洞包膜的结构，使其非常适合用作各种药物的载体。药物可以锚定在BGs的细胞内膜和外膜中，也可以锚定和填充在BGs的周质空间和内腔中，其载药机理是药物与内部空间之间的简单静电相互作用^[126-127]。Zhu等^[128]成功将埃博毒素B通过非共价作用力装载到由EcN制备的BGs的内部空间中，并可以显著提高埃博毒素B在HeLa细胞中的抗癌效果，并减少所需的剂量和相关的副作用。因为BGs是一种非活体细菌，内部不含任何有害化学物质，也不会产生细胞毒性和恢复到致病形式以造成危害，而且BGs是无DNA的，也不存在基因水平传播的风险，所以BGs具有很高的安全性^[129]。

此外，BGs在室温下具有优异的稳定性，也表明其是一种很好的候选口服递送载体^[130]。Eslaminejad等^[131]用干酪乳杆菌制备了一种干酪乳杆菌BGs用以装载克霉唑，并展现出优异的载药能力和显著的抗念珠菌作用。谢松志^[132]将抗菌药物CIP载入到EcN制备的BGs内腔中，具有靶向和刺激响应性释放药物的能力，成功构建出一种新型口服载药平台。然而，有些报道也指出了BGs存在的问题：BGs的空洞结构会使得靶点药物浓度和免疫原性下降；BGs似乎不适合免疫力低下的病人^[133]。尽管如此，BGs是一种很有前景的口服递送载体，但是对于其的研究还有很多工作要做。

5 总结与展望

收起

益生菌是理想的口服递送载体，可以保护药物顺利通过复杂胃肠道环境而不被破坏，能够将药物靶向递送到特定病变部位，并且具有很好的安全性和与药物结合的稳定性。因此许多益生菌已经被开发为递送各种药物的疫苗，如DNA疫苗、蛋白质疫苗、化学药物疫苗和多肽疫苗等^[134-137]，并且应用在治疗过敏、炎症、肿瘤等疾病中^[138-140]。

本文综述了3种常见的益生菌口服递送载体和2种新兴的口服递送载体，并介绍了它们的结构与特点。益生菌的特殊结构，如芽孢杆菌的芽孢、酵母的β-葡聚糖等可以很好地装载药物并保护它们通过复杂的肠道环境，以实现药物的高效递送，并且益生菌口服递送载体具有表面展示、分泌、在细胞质内等多种药物表达模式，可以根据药物的种类和特点进行匹配。此外，可以用作口服递送载体的益生菌种类繁多，包括乳酸菌、酵母、益生芽孢杆菌和大肠杆菌Nissle1917等，具有广泛的开发潜力。益生菌口服递送载体以其特殊的结构提高药物递送的稳定性与靶向性，多种药物表达模式为药物的选择使用提供普遍性，再佐以益生菌安全无害的特性，是一种非常有应用潜力的药物递送系统。

然而，益生菌口服递送载体目前仍然存在以下问题。许多益生菌口服递送载体是使用非活性细菌制备的，而制备过程的研究与安全控制方面的研究还不充分，使得益生菌口服疫苗的工业化生产存在阻碍。此外，使用益生菌作为递送载体的最大问题就是其在胃肠道中的稳定性，口服益生菌普遍存在生物利用度低和治疗效果不稳定的问题^[141]。针对这些问题，目前也有很多研究聚焦于在益生菌表面修饰各种涂层以达到保护益生菌的目的，包括物理修饰和化学修饰等^[142-144]。

最后，现在对于益生菌口服递送载体的研究更多是在动物模型上实现的，在临床上的报道较少。益生菌口服递送载体是一种很有应用前景的药物递送平台，相信随着研究的不断深入与丰富，其可以在药物递送领域提供一条可靠的选择。

基金

收起

国家重点研发计划(2021YFC2102700)
国家自然科学基金(32272269)

参考文献

收起

文献

收起

参考文献引证文献

排序方式：

[1]

HOTEL AC, CORDOBA A.Health and nutritional properties of probiotics in food including powder milk with live lactic acid bacteria[J].Prevention,2001,5(1):1-10.

[2]

YOHA KS, NIDA S, DUTTA S, MOSES JA, ANANDHARAMAKRISHNAN C.Targeted delivery of probiotics: perspectives on research and commercialization[J].Probiotics and Antimicrobial Proteins,2022,14(1):15-48.

https://doi.org/10.1007/s12602-021-09791-7

[3]

SANDERS ME, AKKERMANS LMA, HALLER D, HAMMERMAN C, HEIMBACH J, HÖRMANNSPERGER G, HUYS G, LEVY DD, LUTGENDORFF F, MACK D, PHOTHIRATH P, SOLANO-AGUILAR G, VAUGHAN E.Safety assessment of probiotics for human use[J].Gut Microbes,2010,1(3):164-185.

https://doi.org/10.4161/gmic.1.3.12127

[4]

MILNER E, STEVENS B, AN M, LAM V, AINSWORTH M, DIHLE P, STEARNS J, DOMBROWSKI A, REGO D, SEGARS K.Utilizing probiotics for the prevention and treatment of gastrointestinal diseases[J].Frontiers in Microbiology,2021,12: 689958.

https://doi.org/10.3389/fmicb.2021.689958

[5]

JAKUBCZYK D, LESZCZYŃSKA K, GÓRSKA S.The effectiveness of probiotics in the treatment of inflammatory bowel disease (IBD)-a critical review[J].Nutrients,2020,12(7):1973.

https://doi.org/10.3390/nu12071973

[6]

徐康, 王哲, 薛志朋, 王家朋, 徐振上, 王婷.益生菌缓解或治疗炎症性肠病的作用及机制研究进展[J]. 食品工业科技,2022,43(20):414-420.

XU K, WANG Z, XUE ZP, WANG JP, XU ZS, WANG T.Effect and mechanism of probiotics in alleviating or treating inflammatory bowel disease[J].Science and Technology of Food Industry,2022,43(20):414-420 (in Chinese).

[7]

LEGESSE BEDADA T, FETO TK, AWOKE KS, GAREDEW AD, YIFAT FT, BIRRI DJ.Probiotics for cancer alternative prevention and treatment[J].Biomedicine & Pharmacotherapy,2020,129: 110409.

[8]

TERPOU A, PAPADAKI A, LAPPA IK, KACHRIMANIDOU V, BOSNEA LA, KOPSAHELIS N.Probiotics in food systems: significance and emerging strategies towards improved viability and delivery of enhanced beneficial value[J].Nutrients,2019,11(7):1591.

https://doi.org/10.3390/nu11071591

[9]

SHOKRYAZDAN P, FASELEH JAHROMI M, LIANG JB, HO YW.Probiotics: from isolation to application[J].Journal of the American College of Nutrition,2017,36(8):666-676.

https://doi.org/10.1080/07315724.2017.1337529

[10]

PUEBLA-BARRAGAN S, REID G.Probiotics in cosmetic and personal care products: trends and challenges[J].Molecules,2021,26(5):1249.

https://doi.org/10.3390/molecules26051249

[11]

EI-SAADONY MT, ALAGAWANY M, PATRA AK, KAR I, TIWARI R, DAWOOD MAO, DHAMA K, ABDEL-LATIF HMR.The functionality of probiotics in aquaculture: an overview[J].Fish & Shellfish Immunology,2021,117: 36-52.

[12]

SUEZ J, ZMORA N, SEGAL E, ELINAV E.The pros, cons, and many unknowns of probiotics[J].Nature Medicine,2019,25(5):716-729.

https://doi.org/10.1038/s41591-019-0439-x

[13]

ZHANG L, WANG SL, ZHANG MH, SUN J.Nanocarriers for oral drug delivery[J].Journal of Drug Targeting,2013,21(6):515-527.

https://doi.org/10.3109/1061186X.2013.789033

[14]

NESLIHAN GURSOY R, BENITA S.Self-emulsifying drug delivery systems (SEDDS) for improved oral delivery of lipophilic drugs[J].Biomedicine & Pharmacotherapy,2004,58(3):173-182.

[15]

赵特, 邹鹏飞, 李佳欣, 高春生, 李志平.自组装多肽用于药物递送的研究进展[J]. 军事医学,2023,47(10):783-787.

https://doi.org/10.7644/j.issn.1674-9960.2023.10.012

ZHAO T, ZOU PF, LI JX, GAO CS, LI ZP.Progress in self-assembling peptides for drug delivery[J].Military Medical Sciences,2023,47(10):783-787 (in Chinese).

https://doi.org/10.7644/j.issn.1674-9960.2023.10.012

[16]

MEI L, ZHANG ZP, ZHAO LY, HUANG LQ, YANG XL, TANG JT, FENG SS.Pharmaceutical nanotechnology for oral delivery of anticancer drugs[J].Advanced Drug Delivery Reviews,2013,65(6):880-890.

https://doi.org/10.1016/j.addr.2012.11.005

[17]

BHUTANI U, BASU T, MAJUMDAR S.Oral drug delivery: conventional to long acting new-age designs[J].European Journal of Pharmaceutics and Biopharmaceutics,2021,162: 23-42.

https://doi.org/10.1016/j.ejpb.2021.02.008

[18]

LI WS, SU ZG, HAO MX, JU CY, ZHANG C.Cytopharmaceuticals: an emerging paradigm for drug delivery[J].Journal of Controlled Release,2020,328: 313-324.

https://doi.org/10.1016/j.jconrel.2020.08.063

[19]

LIANG J, YAN H, PULIGUNDLA P, GAO XL, ZHOU YB, WAN XC.Applications of chitosan nanoparticles to enhance absorption and bioavailability of tea polyphenols: a review[J].Food Hydrocolloids,2017,69: 286-292.

https://doi.org/10.1016/j.foodhyd.2017.01.041

[20]

LEONG KW, SUNG HW.Nanoparticle- and biomaterials-mediated oral delivery for drug, gene, and immunotherapy[J].Advanced Drug Delivery Reviews,2013,65(6):757-758.

https://doi.org/10.1016/j.addr.2013.02.002

[21]

JIANG BY, LI ZD, OU BM, DUAN QD, ZHU GQ.Targeting ideal oral vaccine vectors based on probiotics: a systematical view[J].Applied Microbiology and Biotechnology,2019,103(10):3941-3953.

https://doi.org/10.1007/s00253-019-09770-7

[22]

WANG YW, WANG DH, LV HJ, DONG QC, LI JJ, GENG WT, WANG JJ, LIU FF, JIA LG, WANG YP.Modulation of the gut microbiota and glycometabolism by a probiotic to alleviate amyloid accumulation and cognitive impairments in AD rats[J].Molecular Nutrition & Food Research,2022,66(19):e2200265.

[23]

RICCA E, BACCIGALUPI L, CANGIANO G, de FELICE M, ISTICATO R.Mucosal vaccine delivery by non-recombinant spores of Bacillus subtilis[J].Microbial Cell Factories,2014,13: 115.

[24]

WELLS JM, MERCENIER A.Mucosal delivery of therapeutic and prophylactic molecules using lactic acid bacteria[J].Nature Reviews Microbiology,2008,6(5):349-362.

https://doi.org/10.1038/nrmicro1840

[25]

MEDINA M, VINTIÑI E, VILLENA J, RAYA R, ALVAREZ S.Lactococcus lactis as an adjuvant and delivery vehicle of antigens against pneumococcal respiratory infections[J]. Bioengineered Bugs,2010,1(5):313-325.

https://doi.org/10.4161/bbug.1.5.12086

[26]

HE L, YANG HJ, TANG JL, LIU ZD, CHEN YY, LU BH, HE HC, TANG SJ, SUN YJ, LIU F, DING XZ, ZHANG YM, HU SB, XIA LQ.Intestinal probiotics E. coli Nissle 1917 as a targeted vehicle for delivery of p53 and Tum-5 to solid tumors for cancer therapy[J].Journal of Biological Engineering,2019,13: 58.

https://doi.org/10.1186/s13036-019-0189-9

[27]

SHANK EA, KOLTER R.Extracellular signaling and multicellularity in Bacillus subtilis[J].Current Opinion in Microbiology,2011,14(6):741-747.

https://doi.org/10.1016/j.mib.2011.09.016

[28]

ISTICATO R.Bacterial spore-based delivery system: 20 years of a versatile approach for innovative vaccines[J].Biomolecules,2023,13(6):947.

https://doi.org/10.3390/biom13060947

[29]

SELLA SRBR, VANDENBERGHE LPS, SOCCOL CR.Life cycle and spore resistance of spore-forming Bacillus atrophaeus[J].Microbiological Research,2014,169(12):931-939.

https://doi.org/10.1016/j.micres.2014.05.001

[30]

TAN IS, RAMAMURTHI KS.Spore formation in Bacillus subtilis[J].Environmental Microbiology Reports,2014,6(3):212-225.

https://doi.org/10.1111/1758-2229.12130

[31]

SETLOW P.Spore resistance properties[J].Microbiology Spectrum,2014,2(5):201-205.

[32]

CAO J, YU ZM, LIU WY, ZHAO JX, ZHANG H, ZHAI QX, CHEN W.Probiotic characteristics of Bacillus coagulans and associated implications for human health and diseases[J].Journal of Functional Foods,2020,64: 103643.

https://doi.org/10.1016/j.jff.2019.103643

[33]

YIN L, MENG Z, ZHANG YX, HU KK, CHEN WY, HAN KB, WU BY, YOU R, LI CH, JIN Y, GUAN YQ.Bacillus spore-based oral carriers loading curcumin for the therapy of colon cancer[J]. Journal of Controlled Release,2018,271: 31-44.

https://doi.org/10.1016/j.jconrel.2017.12.013

[34]

SUN HC, LIN ZP, ZHAO L, CHEN TJ, SHANG M, JIANG HY, TANG ZL, ZHOU XY, SHI MC, ZHOU LN, REN PL, QU HL, LIN JS, LI XR, XU J, HUANG Y, YU XB.Bacillus subtilis spore with surface display of paramyosin from Clonorchis sinensis potentializes a promising oral vaccine candidate[J]. Parasites & Vectors,2018,11(1):156.

[35]

KANG M, FENG F, GE Q, ZHU F, CHEN L, LV P, MA S, YAO Q, CHEN K.Display of quintuple glucagon-like peptide 1 (28-36) nonapeptide on Bacillus subtilis spore for oral administration in the treatment of type 2 diabetes[J].Journal of Applied Microbiology,2021,130(2):314-324.

https://doi.org/10.1111/jam.14729

[36]

ZHANG GY, AN YF, ZABED H, GUO Q, YANG MM, YUAN J, WEN L, SUN WJ, QI XH.Bacillus subtilis spore surface display technology: a review of its development and applications[J]. Journal of Microbiology and Biotechnology,2019,29(2):179-190.

https://doi.org/10.4014/jmb.1807.06066

[37]

YANG Y, ZHANG GH, WU J, CHEN XQ, TONG DN, YANG YM, SHI HZ, YAO CQ, ZHUANG LN, WANG JB, DU AF.Recombinant HcGAPDH protein expressed on probiotic Bacillus subtilis spores protects sheep from Haemonchus contortus infection by inducing both humoral and cell-mediated responses[J].mSystems,2020,5(3):e00239-20.

[38]

CHEN C, LI YL, LV FL, XU LD, HUANG YW.Surface display of peptides corresponding to the heptad repeat 2 domain of the feline enteric coronavirus spike protein on Bacillus subtilis spores elicits protective immune responses against homologous infection in a feline aminopeptidase-N-transduced mouse model[J].Frontiers in Immunology,2022,13: 925922.

https://doi.org/10.3389/fimmu.2022.925922

[39]

杨云川. 枯草芽孢杆菌表面展示AIV HA和GPV VP3及其免疫原性的初步评价[D]. 重庆: 西南大学硕士学位论文, 2023.

YANG YC. Preliminary evaluation of AIV HA and GPV VP3 displayed on the surface of Bacillus subtilis and their immunogenicity[D]. Chongqing: Master's Thesis of Southwest University, 2023 (in Chinese).

[40]

宋庆龄. 基于益生菌芽孢的口服递送系统用于结肠炎和结肠癌治疗研究[D]. 郑州: 郑州大学博士学位论文, 2023.

SONG QL. Oral delivery system based on probiotic spores for the treatment of colitis and colon cancer[D]. Zhengzhou: Doctoral Dissertation of Zhengzhou University, 2023 (in Chinese).

[41]

SONG M, HONG HA, HUANG JM, COLENUTT C, KHANG DD, van ANH NGUYEN T, PARK SM, SHIM BS, SONG HH, CHEON IS, JANG JE, CHOI JA, CHOI YK, STADLER K, CUTTING SM.Killed Bacillus subtilis spores as a mucosal adjuvant for an H5N1 vaccine[J].Vaccine,2012,30(22):3266-3277.

https://doi.org/10.1016/j.vaccine.2012.03.016

[42]

SUN HC, SHANG M, TANG ZL, JIANG HY, DONG HM, ZHOU XY, LIN ZP, SHI CB, REN PL, ZHAO L, SHI MC, ZHOU LN, PAN HJ, CHANG OQ, LI XR, HUANG Y, YU XB.Oral delivery of Bacillus subtilis spores expressing Clonorchis sinensis paramyosin protects grass carp from cercaria infection[J].Applied Microbiology and Biotechnology,2020,104(4):1633-1646.

https://doi.org/10.1007/s00253-019-10316-0

[43]

SUN R, ZHANG M, CHEN H, WEI Y, NING DG.Germination-arrest Bacillus subtilis spores as an oral delivery vehicle of grass carp reovirus (GCRV) vp7 antigen augment protective immunity in grass carp (Ctenopharyngodon idella)[J].Genes,2020,11(11):1351.

https://doi.org/10.3390/genes11111351

[44]

GONÇALVES G, SANTOS RA, COUTINHO F, PEDROSA N, CURADO M, MACHADO M, COSTAS B, BONNEVILLE L, SERRANO M, CARVALHO AP, DÍAZ-ROSALES P, OLIVA-TELES A, COUTO A, SERRA CR.Oral vaccination of fish against vibriosis using spore-display technology[J].Frontiers in Immunology,2022,13: 1012301.

https://doi.org/10.3389/fimmu.2022.1012301

[45]

ZHOU ZW, GONG ST, LI XM, YANG YY, GUAN RL, ZHOU S, YAO SW, XIE YQ, OU ZY, ZHAO JH, LIU ZG. Expression of Helicobacter pylori urease B on the surface of Bacillus subtilis spores[J]. Journal of Medical Microbiology, 2015, 64 (Pt 1):104-110.

[46]

OH Y, KIM JA, KIM CH, CHOI SK, PAN JG.Bacillus subtilis spore vaccines displaying protective antigen induce functional antibodies and protective potency[J]. BMC Veterinary Research,2020,16(1):259.

https://doi.org/10.1186/s12917-020-02468-3

[47]

de ALMEIDA MEM, ALVES KCS, de VASCONCELOS MGS, PINTO TS, GLÓRIA JC, CHAVES YO, NEVES WLL, TARRAGÔ AM, de SOUZA NETO JN, ASTOLFI-FILHO S, PONTES GS, da SILVA BALIEIRO AA, ISTICATO R, RICCA E, MARIÚBA LAM.Bacillus subtilis spores as delivery system for nasal Plasmodium falciparum circumsporozoite surface protein immunization in a murine model[J]. Scientific Reports,2022,12(1):1531.

https://doi.org/10.1038/s41598-022-05344-2

[48]

SANTOS FDS, MAZZOLI A, MAIA AR, SAGGESE A, ISTICATO R, LEITE F, IOSSA S, RICCA E, BACCIGALUPI L.A probiotic treatment increases the immune response induced by the nasal delivery of spore-adsorbed TTFC[J].Microbial Cell Factories,2020,19(1):42.

https://doi.org/10.1186/s12934-020-01308-1

[49]

TANG ZL, WU ZS, SUN HC, ZHAO L, SHANG M, SHI MC, JIANG HY, LIN ZP, ZHOU XY, LI XR, YU XB, HUANG Y.The storage stability of Bacillus subtilis spore displaying cysteine protease of Clonorchis sinensis and its effect on improving the gut microbiota of mice[J].Applied Microbiology and Biotechnology,2021,105(6):2513-2526.

https://doi.org/10.1007/s00253-021-11126-z

[50]

LEE NK, KIM WS, PAIK HD.Bacillus strains as human probiotics: characterization, safety, microbiome, and probiotic carrier[J]. Food Science and Biotechnology,2019,28(5):1297-1305.

https://doi.org/10.1007/s10068-019-00691-9

[51]

DUC LH, HONG HA, ATKINS HS, FLICK-SMITH HC, DURRANI Z, RIJPKEMA S, TITBALL RW, CUTTING SM.Immunization against anthrax using Bacillus subtilis spores expressing the anthrax protective antigen[J].Vaccine,2007,25(2):346-355.

https://doi.org/10.1016/j.vaccine.2006.07.037

[52]

CHEN HY, ZHANG TX, JIA JR, VASTERMARK A, TIAN R, NI Z, CHEN Z, CHEN KP, YANG SL.Expression and display of a novel thermostable esterase from Clostridium thermocellum on the surface of Bacillus subtilis using the CotB anchor protein[J].Journal of Industrial Microbiology & Biotechnology,2015,42(11):1439-1448.

[53]

SAGGESE A, BACCIGALUPI L, DONADIO G, RICCA E, ISTICATO R.The bacterial spore as a mucosal vaccine delivery system[J].International Journal of Molecular Sciences,2023,24(13):10880.

https://doi.org/10.3390/ijms241310880

[54]

DONG H, HUANG YM, YAO SW, LIANG BS, LONG Y, XIE YQ, MAI JL, GONG ST, ZHOU ZW.The recombinant fusion protein of cholera toxin B and neutrophil-activating protein expressed on Bacillus subtilis spore surface suppresses allergic inflammation in mice[J].Applied Microbiology and Biotechnology,2017,101(14):5819-5829.

https://doi.org/10.1007/s00253-017-8370-x

[55]

VETRÁKOVÁ A, CHOVANOVÁ RK, RECHTORÍKOVÁ R, KRAJČÍKOVÁ D, BARÁK I.Bacillus subtilis spores displaying RBD domain of SARS-CoV-2 spike protein[J]. Computational and Structural Biotechnology Journal,2023,21: 1550-1556.

https://doi.org/10.1016/j.csbj.2023.02.007

[56]

ISTICATO R, SIREC T, TREPPICCIONE L, MAURANO F, de FELICE M, ROSSI M, RICCA E.Non-recombinant display of the B subunit of the heat labile toxin of Escherichia coli on wild type and mutant spores of Bacillus subtilis[J].Microbial Cell Factories,2013,12: 98.

https://doi.org/10.1186/1475-2859-12-98

[57]

HINC K, ISTICATO R, DEMBEK M, KARCZEWSKA J, IWANICKI A, PESZYŃSKA-SULARZ G, de FELICE M, OBUCHOWSKI M, RICCA E.Expression and display of UreA of Helicobacter acinonychis on the surface of Bacillus subtilis spores[J].Microbial Cell Factories,2010,9: 2.

https://doi.org/10.1186/1475-2859-9-2

[58]

CHERNOFF YO, GRIZEL AV, RUBEL AA, ZELINSKY AA, CHANDRAMOWLISHWARAN P, CHERNOVA TA.Application of yeast to studying amyloid and prion diseases[J].Advances in Genetics,2020,105: 293-380.

[59]

KARIM A, GERLIANI N, AÏDER M.Kluyveromyces marxianus: an emerging yeast cell factory for applications in food and biotechnology[J]. International Journal of Food Microbiology,2020,333: 108818.

https://doi.org/10.1016/j.ijfoodmicro.2020.108818

[60]

FENG XX, XIE Q, XU HB, ZHANG TT, LI XN, TIAN YM, LAN HB, KONG L, ZHANG ZP.Yeast microcapsule mediated natural products delivery for treating ulcerative colitis through anti-inflammatory and regulation of macrophage polarization[J].ACS Applied Materials & Interfaces,2022,14(27):31085-31098.

[61]

WU Y, LI PY, JIANG ZZ, SUN XL, HE HQ, YAN PJ, XU Y, LIU Y.Bioinspired yeast-based β-glucan system for oral drug delivery[J].Carbohydrate Polymers,2023,319: 121163.

https://doi.org/10.1016/j.carbpol.2023.121163

[62]

BASTOS R, OLIVEIRA PG, GASPAR VM, MANO JF, COIMBRA MA, COELHO E.Brewer's yeast polysaccharides: a review of their exquisite structural features and biomedical applications[J].Carbohydrate Polymers,2022,277: 118826.

https://doi.org/10.1016/j.carbpol.2021.118826

[63]

LIU Y, YANG HS, ZHU JH, YANG ZF, ZHAO LL, ZHANG X, ZHANG HX.Novel albendazole-glucan particles for enhancing intestinal absorption and improving hepatic targeting[J].Annals of Translational Medicine,2022,10(24):1312

https://doi.org/10.21037/atm-22-5299

[64]

刘瑛琪, 李静如, 孟繁, 邢昊楠, 郑爱萍.酵母微囊作为口服药物递送载体的研究进展[J]. 中国药房,2023,34(16):2022-2027.

https://doi.org/10.6039/j.issn.1001-0408.2023.16.20

LIU YQ, LI JR, MENG F, XING HN, ZHENG AP.Research progress of yeast microcapsules as oral drug delivery carrier[J].China Pharmacy,2023,34(16):2022-2027 (in Chinese).

https://doi.org/10.6039/j.issn.1001-0408.2023.16.20

[65]

ZHOU X, LING KJ, LIU MY, ZHANG XJ, DING J, DONG Y, LIANG ZQ, LI JJ, ZHANG JX.Targeted delivery of cisplatin-derived nanoprecursors via a biomimetic yeast microcapsule for tumor therapy by the oral route[J].Theranostics,2019,9(22):6568-6586.

https://doi.org/10.7150/thno.35353

[66]

SABU C, RAGHAV D, JIJITH US, MUFEEDHA P, NASEEF PP, RATHINASAMY K, PRAMOD K.Bioinspired oral insulin delivery system using yeast microcapsules[J].Materials Science and Engineering: C,2019,103: 109753.

https://doi.org/10.1016/j.msec.2019.109753

[67]

王大壮. 酵母微囊包覆纳米氧化锌载药递送系统的构建及抗肿瘤作用研究[D]. 沈阳: 辽宁大学硕士学位论文, 2022.

WANG DZ. Construction of drug delivery system of nano-zinc oxide coated with yeast microcapsules and its anti-tumor effect[D]. Shenyang: Master's Thesis of Liaoning University, 2022 (in Chinese).

[68]

SHI GR, LIU YT, HE ZJ, ZHOU JH.Chemical treatment and chitosan coating of yeast cells to improve the encapsulation and controlled release of bovine serum albumin[J].Artificial Cells, Nanomedicine, and Biotechnology,2017,45(6):1-9.

[69]

SALOŇ I, HANUŠ J, ULBRICH P, ŠTĚPÁNEK F.Suspension stability and diffusion properties of yeast glucan microparticles[J].Food and Bioproducts Processing,2016,99: 128-135.

https://doi.org/10.1016/j.fbp.2016.04.010

[70]

TAN C, WANG J, SUN BG.Polysaccharide dual coating of yeast capsules for stabilization of anthocyanins[J].Food Chemistry,2021,357: 129652.

https://doi.org/10.1016/j.foodchem.2021.129652

[71]

HE Y, CHEN QW, YU JX, QIN SY, LIU WL, MA YH, CHEN XS, ZHANG AQ, ZHANG XZ, CHENG YJ.Yeast cell membrane-camouflaged PLGA nanoparticle platform for enhanced cancer therapy[J].Journal of Controlled Release,2023,359: 347-358.

https://doi.org/10.1016/j.jconrel.2023.05.043

[72]

SOTO ER, SPECHT CA, LEE CK, LEVITZ SM, OSTROFF GR.One step purification-vaccine delivery system[J].Pharmaceutics,2023,15(5):1390.

https://doi.org/10.3390/pharmaceutics15051390

[73]

PAN Y, LI XP, KANG TY, MENG H, CHEN ZL, YANG L, WU Y, WEI YQ, GOU ML.Efficient delivery of antigen to DCs using yeast-derived microparticles[J].Scientific Reports,2015,5: 10687.

https://doi.org/10.1038/srep10687

[74]

IVANOVA E.Yeasts in nanotechnology-enabled oral vaccine and gene delivery[J].Bioengineered,2021,12(1):8325-8335.

https://doi.org/10.1080/21655979.2021.1985816

[75]

ZHANG L, ZHANG W, PENG H, LI YK, LENG TT, XIE CX, ZHANG L.Oral gene therapy of HFD-obesity via nonpathogenic yeast microcapsules mediated shRNA delivery[J].Pharmaceutics,2021,13(10):1536.

https://doi.org/10.3390/pharmaceutics13101536

[76]

ZHOU X, ZHANG XJ, HAN SL, DOU Y, LIU MY, ZHANG L, GUO JW, SHI Q, GONG GH, WANG RB, HU J, LI XH, ZHANG JX.Yeast microcapsule-mediated targeted delivery of diverse nanoparticles for imaging and therapy via the oral route[J].Nano Letters,2017,17(2):1056-1064.

https://doi.org/10.1021/acs.nanolett.6b04523

[77]

SOTO ER, KIM HC, YAGITA H, de JESUS M, OSTROFF GR.Polydopamine coating of glucan particles increases uptake into peyer's patches[J].ACS Applied Bio Materials,2019,2(9):3748-3754.

https://doi.org/10.1021/acsabm.9b00379

[78]

REIS SF, MARTINS VJ, BASTOS R, LIMA T, CORREIA VG, PINHEIRO BA, SILVA LM, PALMA AS, FERREIRA P, VILANOVA M, COIMBRA MA, COELHO E.Feasibility of brewer's spent yeast microcapsules as targeted oral carriers[J].Foods,2023,12(2):246.

https://doi.org/10.3390/foods12020246

[79]

ZHANG L, PENG H, FENG M, ZHANG W, LI YK.Yeast microcapsule-mediated oral delivery of IL-1β shRNA for post-traumatic osteoarthritis therapy[J].Molecular Therapy-Nucleic Acids,2021,23: 336-346.

https://doi.org/10.1016/j.omtn.2020.11.006

[80]

德艳艳, 张艳花, 廖洪, 龙谭.重组益生菌的生物学潜在应用[J]. 中国预防兽医学报,2021,43(4):444-449.

DE YY, ZHANG YH, LIAO H, LONG T.Potential biological applications of recombinant probiotics[J].Chinese Journal of Preventive Veterinary Medicine,2021,43(4):444-449 (in Chinese).

[81]

KUMAR M, YADAV AK, VERMA V, SINGH B, MAL G, NAGPAL R, HEMALATHA R.Bioengineered probiotics as a new hope for health and diseases: an overview of potential and prospects[J].Future Microbiology,2016,11(4):585-600.

https://doi.org/10.2217/fmb.16.4

[82]

陈少俊, 李刚, 奈子达, 刘娣, 姜新鹏.基因编辑益生菌在动物肠道健康上的研究及应用[J]. 中国畜牧兽医,2023,50(3):1016-1024.

CHEN SJ, LI G, NAI ZD, LIU D, JIANG XP.Research and application of gene-edited probiotics in animal intestinal health[J].China Animal Husbandry & Veterinary Medicine,2023,50(3):1016-1024 (in Chinese).

[83]

岳梦云. 工程益生菌L. lactis MG1363-pMG36e-GLP-1通过抑制铁死亡改善帕金森病的机制研究[D]. 南昌: 南昌大学硕士学位论文, 2023.

YUE MY. Study on the mechanism of L. lactis MG1363-pMG36e-GLP-1, an engineering probiotic, in improving Parkinson's disease by inhibiting iron death[D]. Nanchang: Master's Thesis of Nanchang University, 2023 (in Chinese).

[84]

赵瑞. 基于Escherichia coli nissle 1917构建表达尿酸氧化酶以及氧循环系统的工程益生菌用于治疗高尿酸血症的研究[D]. 济南: 山东大学博士学位论文, 2022.

ZHAO R. Study on the treatment of hyperuricemia by constructing engineering probiotics expressing uric acid oxidase and oxygen circulation system based on Escherichia coli Nissle 1917[D]. Jinan: Doctoral Dissertation of Shandong University, 2022 (in Chinese).

[85]

MATHIPA-MDAKANE MG, THANTSHA MS.Lacticaseibacillus rhamnosus: a suitable candidate for the construction of novel bioengineered probiotic strains for targeted pathogen control[J]. Foods,2022,11(6):785.

https://doi.org/10.3390/foods11060785

[86]

WYSZYŃSKA AK, GODLEWSKA R.Lactic acid bacteria-a promising tool for controlling chicken Campylobacter infection[J].Frontiers in Microbiology,2021,12: 703441.

https://doi.org/10.3389/fmicb.2021.703441

[87]

TAGHINEZHAD-S S, MOHSENI AH, BERMÚDEZ-HUMARÁN LG, CASOLARO V, CORTES-PEREZ NG, KEYVANI H, SIMAL-GANDARA J.Probiotic-based vaccines may provide effective protection against COVID-19 acute respiratory disease[J].Vaccines,2021,9(5):466.

https://doi.org/10.3390/vaccines9050466

[88]

MUÑOZ C, GONZÁLEZ-LORCA J, PARRA M, SOTO S, VALDES N, SANDINO AM, VARGAS R, GONZÁLEZ A, TELLO M.Lactococcus lactis expressing type I interferon from Atlantic salmon enhances the innate antiviral immune response in vivo and in vitro[J]. Frontiers in Immunology,2021,12: 696781.

https://doi.org/10.3389/fimmu.2021.696781

[89]

HUANG QT, NIU TM, ZOU BS, WANG JH, XIN JH, NIU H, LI N, JIANG YX, BAO JF, ZHANG D, FENG XZ, SUN TT, WANG X, YANG KD, WANG Y, YANG GL, ZHAO DD, WANG CF.Lactobacillus plantarum surface-displayed ASFV (p14.5) can stimulate immune responses in mice[J]. Vaccines,2022,10(3):355.

https://doi.org/10.3390/vaccines10030355

[90]

ZHAO ZL, WANG H, ZHANG DX, GUAN YC, SIDDIQUI SA, XIAO FS, CONG B.Oral vaccination with recombinant Lactobacillus casei expressing Aeromonas hydrophila Aha1 against A. hydrophila infections in common carps[J].Virulence,2022,13(1):794-807.

https://doi.org/10.1080/21505594.2022.2063484

[91]

孟薇. 表面展示禽流感病毒抗原酿酒酵母菌株的构建及其生物学特性研究[D]. 天津: 天津大学硕士学位论文, 2020.

MENG W. Construction and biological characteristics of Saccharomyces cerevisiae strain with surface display of avian influenza virus antigen[D]. Tianjin: Master's Thesis of Tianjin University, 2020 (in Chinese).

[92]

张小禹. 表达具核梭杆菌外膜FomA蛋白的新型乳酸菌的构建及其抑制炎症性肠病的机制研究[D]. 长春: 吉林大学博士学位论文, 2023.

ZHANG XY. Construction of a novel Lactobacillus expressing FomA protein in the outer membrane of Fusobacterium nucleatum and its inhibitory effect on inflammatory bowel disease[D]. Changchun: Doctoral Dissertation of Jilin University, 2023 (in Chinese).

[93]

PAN N, LIU BH, BAO XM, ZHANG HC, SHENG SX, LIANG YC, PAN HT, WANG X.Oral delivery of novel recombinant Lactobacillus elicit high protection against Staphylococcus aureus pulmonary and skin infections[J].Vaccines,2021,9(9):984.

https://doi.org/10.3390/vaccines9090984

[94]

ZHI WJ, CHEN H, BAI BR, JIA ZP, PAN XH, WANG B, KONG R, LIU QJ, MA CL, MA DX.Combined oral immunization with probiotics Entercoccus faecalis delivering surface-anchored Eimeria tenella proteins provide protective efficacies against homologous infection in chickens[J].Frontiers in Immunology,2022,13: 1042143.

https://doi.org/10.3389/fimmu.2022.1042143

[95]

SHIRAKAWA T, KITAGAWA K.Antitumor effect of oral cancer vaccine with Bifidobacterium delivering WT1 protein to gut immune system is superior to WT1 peptide vaccine[J].Human Vaccines & Immunotherapeutics,2017,14(1):159-162.

[96]

NIU H, XING JH, ZOU BS, SHI CW, HUANG HB, JIANG YL, WANG JZ, CAO X, WANG N, ZENG Y, YANG WT, YANG GL, WANG CF.Immune evaluation of recombinant Lactobacillus plantarum with surface display of HA1-DCpep in mice[J].Frontiers in Immunology,2021,12: 800965.

https://doi.org/10.3389/fimmu.2021.800965

[97]

CHEN YP, HUA XJ, REN XY, DUAN KX, GAO SA, SUN JH, FENG Y, ZHOU Y, GUAN X, LI DC, WANG N, LI JH, YANG JW, XIA D, SHI W, LIU M.Oral immunization with recombinant Lactobacillus casei displayed AHA1-CK6 and VP2 induces protection against infectious pancreatic necrosis in rainbow trout (Oncorhynchus mykiss)[J].Fish & Shellfish Immunology,2020,100: 18-26.

[98]

MA ST, WANG L, HUANG XW, WANG XN, CHEN S, SHI W, QIAO XY, JIANG YP, TANG LJ, XU YG, LI YJ.Oral recombinant Lactobacillus vaccine targeting the intestinal microfold cells and dendritic cells for delivering the core neutralizing epitope of porcine epidemic diarrhea virus[J].Microbial Cell Factories,2018,17: 20.

https://doi.org/10.1186/s12934-018-0861-7

[99]

LI HJ, YANG BT, SUN YF, ZHAOT, HAO ZP, GU W, SUN MX, CONG W, KANG YH.Oral vaccination with recombinant Lactobacillus casei with surface displayed OmpK fused to CTB as an adjuvant against Vibrio mimicus infection in Carassius auratus[J].Fish & Shellfish Immunology,2023,135: 108659.

[100]

CHUNG Y, RYU Y, AN BC, YOON Y S, CHOI O, KIM TY, YOON J, AHN JY, PARK HJ, KWON SK, KIM JF, CHUNG MJ.A synthetic probiotic engineered for colorectal cancer therapy modulates gut microbiota[J].Microbiome,2021,9(1):122.

https://doi.org/10.1186/s40168-021-01071-4

[101]

ZENG Z, YU R, ZUO FL, ZHANG B, PENG DJ, MA HQ, CHEN SW.Heterologous expression and delivery of biologically active exendin-4 by Lactobacillus paracasei L14[J].PloS One,2016,11(10):e0165130.

https://doi.org/10.1371/journal.pone.0165130

[102]

LV PH, ZHANG XS, SONG MZ, HAO GJ, WANG FK, SUN SH.Oral administration of recombinant Bacillus subtilis expressing a multi-epitope protein induces strong immune responses against Salmonella enteritidis[J].Veterinary Microbiology,2023,276: 109632.

https://doi.org/10.1016/j.vetmic.2022.109632

[103]

YUAN SL, DONG M, ZHANG HL, XU HD, WANG Q, YAN CL, YE RC, JIANG XX, ZHOU HQ, CHEN L, CHENG J, XIE W, JIN WZ.Oral delivery of a Lactococcus lactis expressing extracellular TGFβR2 alleviates hepatic fibrosis[J].Applied Microbiology and Biotechnology,2021,105(14):6007-6018.

[104]

LIN ZQ, TANG YQ, CHEN ZR, LI SM, XU XY, HOU XF, CHEN ZH, WEN JJ, ZENG WS, MENG XJ, FAN HY.Soluble CD80 oral delivery by recombinant Lactococcus suppresses tumor growth by enhancing antitumor immunity[J].Bioengineering & Translational Medicine,2023,8(4):e10533.

[105]

CHAU ECT, KWONG TC, PANG CK, CHAN LT, CHAN AML, YAO XQ, TAM JSL, CHAN SW, LEUNG GPH, TAI WCS, KWAN YW.A novel probiotic-based oral vaccine against SARS-CoV-2 Omicron variant B. 1.1. 529[J].International Journal of Molecular Sciences,2023,24(18):13931.

https://doi.org/10.3390/ijms241813931

[106]

HOU J, LU Y, LIU HY, MA YJ, XING Y, ZHANG Y, LI TM, CAO RY, JIN L, WU J, ZONG L, LIU JJ.Oral administration of Lactococcus lactis delivered heat shock protein 65 attenuates atherosclerosis in low-density lipoprotein receptor-deficient mice[J].Vaccine,2011,29(24):4102-4109.

https://doi.org/10.1016/j.vaccine.2011.03.105

[107]

ZHANG RG, PENG XY, DUAN GC, SHI QF, CHEN SY, WANG C, FAN QT, XI YL.An engineered Lactococcus lactis strain exerts significant immune responses through efficient expression and delivery of Helicobacter pylori Lpp20 antigen[J].Biotechnology Letters,2016,38(12):2169-2175.

https://doi.org/10.1007/s10529-016-2209-x

[108]

GRANGETTE C, MÜLLER-ALOUF H, GOUDERCOURT D, GEOFFROY MC, TURNEER M, MERCENIER A.Mucosal immune responses and protection against tetanus toxin after intranasal immunization with recombinant Lactobacillus plantarum[J].Infection and Immunity,2001,69(3):1547-1553.

https://doi.org/10.1128/IAI.69.3.1547-1553.2001

[109]

YANG GL, YAO JY, YANG WT, JIANG YL, DU JF, HUANG HB, GU W, HU JT, YE LP, SHI CW, SHAN BL, WANG CF.Construction and immunological evaluation of recombinant Lactobacillus plantarum expressing SO7 of Eimeria tenella fusion DC-targeting peptide[J].Veterinary Parasitology,2017,236: 7-13.

https://doi.org/10.1016/j.vetpar.2017.01.023

[110]

AI CQ, ZHANG QX, REN CC, WANG G, LIU XM, TIAN FW, ZHAO JX, ZHANG H, CHEN YQ, CHEN W.Genetically engineered Lactococcus lactis protect against house dust mite allergy in a BALB/c mouse model[J].PLoS One,2014,9(10):e109461.

https://doi.org/10.1371/journal.pone.0109461

[111]

曾珠.乳酸菌作为药物分子黏膜投递载体的研究进展[J]. 生物工程学报,2021,37(7):2272-2282.

ZENG Z.Advances in the use of lactic acid bacteria as mucosal delivery vectors of therapeutic molecules[J].Chinese Journal of Biotechnology,2021,37(7):2272-2282 (in Chinese).

[112]

REN CC, ZHANG QX, WANG G, AI CQ, HU MS, LIU XM, TIAN FW, ZHAO JX, CHEN YQ, WANG M, ZHANG H, CHEN W.Modulation of peanut-induced allergic immune responses by oral lactic acid bacteria-based vaccines in mice[J].Applied Microbiology and Biotechnology,2014,98(14):6353-6364.

https://doi.org/10.1007/s00253-014-5678-7

[113]

王建忠, 赵建伟, 王春凤.细菌样颗粒: 新型乳酸菌表面展示技术及其应用[J]. 微生物学报,2019,59(3):411-419. https://actamicro.ijournals.cn/actamicrocn/article/abstract/20190301?st=search

WANG JZ, ZHAO JW, WANG CF.Bacterium-like particles: a novel surface display technology for lactic acid bacteria and its application[J].Acta Microbiologica Sinica,2019,59(3):411-419 (in Chinese). https://actamicro.ijournals.cn/actamicrocn/article/abstract/20190301?st=search

[114]

吕瑞卿, 焦翠翠, 张梦瑶, 王化磊, 张海丽.GEM-PA表面展示系统研究进展[J]. 广东畜牧兽医科技,2023,48(1):51-56.

LÜ RQ, JIAO CC, ZHANG MY, WANG HL, ZHANG HL.Research progress on GEM-PA surface display system[J].Guangdong Journal of Animal and Veterinary Science,2023,48(1):51-56 (in Chinese).

[115]

刘润杭. 基于病原菌制备的细菌样颗粒(pBLP)用作抗原递送载体和疫苗增效剂的效果分析[D]. 哈尔滨: 东北农业大学硕士学位论文, 2023.

LIU RH. Effect analysis of bacterial-like particles (pBLP) prepared from pathogenic bacteria as antigen delivery carrier and vaccine synergist[D]. Harbin: Master's Thesis of Northeast Agricultural University, 2023 (in Chinese).

[116]

李政蓉. 尼帕病毒性脑炎重组狂犬病病毒载体灭活疫苗和细菌样颗粒疫苗的构建与免疫原性研究[D]. 长春: 吉林大学博士学位论文, 2023.

LI ZR. Construction and immunogenicity of recombinant rabies virus vector inactivated vaccine and bacterial particle vaccine against Nipah virus encephalitis[D]. Changchun: Doctoral Dissertation of Jilin University, 2023 (in Chinese).

[117]

ZHOU XY, GAO MC, DE XQ, SUN T, BAI ZK, LUO JL, WANG F, GE JW.Bacterium-like particles derived from probiotics: progress, challenges and prospects[J].Frontiers in Immunology,2023,14: 1263586.

https://doi.org/10.3389/fimmu.2023.1263586

[118]

LIU W, TAN ZL, LIU H, ZENG ZQ, LUO SH, YANG HM, ZHENG LF, XI T, XING YY. Nongenetically modified Lactococcus lactis-adjuvanted vaccination enhanced innate immunity against Helicobacter pylori[J]. Helicobacter, 2017, 22 (5):DOI: 10.1111/hel.12426.

[119]

MAO RF, CHEN YY, WU Q, ZHANG T, DIAO EJ, WU DL, WANG M, LIU Y, LU L, CHANG X, ZHENG Y, WANG YF.Oral delivery of single-chain insulin (SCI-59) analog by bacterium-like particles (BLPs) induces oral tolerance and prevents autoimmune diabetes in NOD mice[J].Immunology Letters,2019,214: 37-44.

https://doi.org/10.1016/j.imlet.2019.08.008

[120]

RAMIREZ K, DITAMO Y, RODRIGUEZ L, PICKING WL, van ROOSMALEN ML, LEENHOUTS K, PASETTI MF.Neonatal mucosal immunization with a non-living, non-genetically modified Lactococcus lactis vaccine carrier induces systemic and local Th1-type immunity and protects against lethal bacterial infection[J].Mucosal Immunology,2010,3(2):159-171.

https://doi.org/10.1038/mi.2009.131

[121]

SUDO H, TOKUNOH N, TSUJII A, KAWASHIMA S, HAYAKAWA Y, FUKUSHIMA H, TAKAHASHI K, KOSHIZUKA T, INOUE N.The adjuvant effect of bacterium-like particles depends on the route of administration[J].Frontiers in Immunology,2023,14: 1082273.

https://doi.org/10.3389/fimmu.2023.1082273

[122]

CHEN HJ, JI H, KONG XJ, LEI PY, YANG QS, WU W, JIN LB, SUN D.Bacterial ghosts-based vaccine and drug delivery systems[J].Pharmaceutics,2021,13(11):1892.

https://doi.org/10.3390/pharmaceutics13111892

[123]

ALI RH, ALI ME, SAMIR R.Production and characterization of bacterial ghost vaccine against Neisseria meningitidis[J].Vaccines,2022,11(1):37.

https://doi.org/10.3390/vaccines11010037

[124]

MA Y, ZHU WJ, ZHU GS, XU Y, LI SY, CHEN R, CHEN LD, WANG JF.Efficient robust yield method for preparing bacterial ghosts by Escherichia coli phage ID52 lysis protein E[J].Bioengineering,2022,9(7):300.

https://doi.org/10.3390/bioengineering9070300

[125]

HOLAY M, GUO ZY, PIHL J, HEO J, PARK JH, FANG RH, ZHANG LF.Bacteria-inspired nanomedicine[J].ACS Applied Bio Materials,2021,4(5):3830-3848.

https://doi.org/10.1021/acsabm.0c01072

[126]

CHEN HJ, LEI PY, JI H, MA JH, FANG YM, YU HY, DU J, QU LK, YANG QS, LUO L, ZHANG K, WU W, JIN LB, SUN D.Escherichia coli Nissle 1917 ghosts alleviate inflammatory bowel disease in zebrafish[J]. Life Sciences,2023,329: 121956.

https://doi.org/10.1016/j.lfs.2023.121956

[127]

SALEM-BEKHIT MM, YOUSSOF AME, ALANAZI FK, ALEANIZY FS, ABDULAZIZ A, TAHA EI, AMARA AAAF.Bacteria from infectious particles to cell based anticancer targeted drug delivery systems[J].Pharmaceutics,2021,13(12):1984.

https://doi.org/10.3390/pharmaceutics13121984

[128]

ZHU WX, HAO LJ, LIU XL, BORRÁS-HIDALGO O, ZHANG YY.Enhanced anti-proliferative efficacy of epothilone B loaded with Escherichia coli Nissle 1917 bacterial ghosts on the HeLa cells by mitochondrial pathway of apoptosis[J].Drug Development and Industrial Pharmacy,2018,44(8):1328-1335.

https://doi.org/10.1080/03639045.2018.1449855

[129]

PANDEY M, CHOUDHURY H, VIJAYAGOMARAN PA, LIAN PNP, NING TJ, WAI NZ, XIAN-ZHUANG N, ER CL, RAHMAH NSN, KAMARUZZAMAN NDB, MAYUREN J, CANDASAMY M, GORAIN B, CHAWLA PA, AMIN MCIM.Recent update on bacteria as a delivery carrier in cancer therapy: from evil to allies[J].Pharmaceutical Research,2022,39(6):1115-1134.

https://doi.org/10.1007/s11095-022-03240-y

[130]

HAJAM IA, DAR PA, WON G, LEE JH.Bacterial ghosts as adjuvants: mechanisms and potential[J].Veterinary Research,2017,48(1):37.

https://doi.org/10.1186/s13567-017-0442-5

[131]

ESLAMINEJAD T, MOSHAFI MH, HASANPORE M, AYATOLLAHI SA, ANSARI M.Evaluation of the anticandidal activity of clotrimazole using Lactobacillus caseie ghosts as biological drug carrier[J].Daru,2022,30(1):67-73.

https://doi.org/10.1007/s40199-022-00432-0

[132]

谢松志. 益生菌作为药物靶向递送和控释载体的研究[D]. 成都: 西南交通大学博士学位论文, 2019.

XIE SZ. Study on probiotics as drug targeted delivery and controlled release carrier[D]. Chengdu: Doctoral Dissertation of Southwest Jiaotong University, 2019 (in Chinese).

[133]

GANESHPURKAR A, GANESHPURKAR A, PANDEY V, AGNIHOTRI A, BANSAL D, DUBEY N.Harnessing the potential of bacterial ghost for the effective delivery of drugs and biotherapeutics[J].International Journal of Pharmaceutical Investigation,2014,4(1):1-4.

https://doi.org/10.4103/2230-973X.127733

[134]

HAN BQ, XU K, LIU ZT, GE W, SHAO SM, LI PC, YAN NN, LI XY, ZHANG ZY.Oral yeast-based DNA vaccine confers effective protection from Aeromonas hydrophila infection on Carassius auratus[J].Fish & Shellfish Immunology,2019,84: 948-954.

[135]

STASIŁOJĆ M, HINC K, PESZYŃSKA-SULARZ G, OBUCHOWSKI M, IWANICKI A.Recombinant Bacillus subtilis spores elicit Th1/Th17-polarized immune response in a murine model of Helicobacter pylori vaccination[J].Molecular Biotechnology,2015,57(8):685-691.

https://doi.org/10.1007/s12033-015-9859-0

[136]

LIM J, KOH VHQ, CHO SSL, PERIASWAMY B, CHOI DPS, VACCA M, de SESSIONS PF, KUDELA P, LUBITZ W, PASTORIN G, ALONSO S.Harnessing the immunomodulatory properties of bacterial ghosts to boost the anti-mycobacterial protective immunity[J].Frontiers in Immunology,2019,10: 2737.

https://doi.org/10.3389/fimmu.2019.02737

[137]

WANG XN, WANG L, HUANG XW, MA ST, YU ML, SHI W, QIAO XY, TANG LJ, XU YG, LI YJ.Oral delivery of probiotics expressing dendritic cell-targeting peptide fused with porcine epidemic diarrhea virus COE antigen: a promising vaccine strategy against PEDV[J].Viruses,2017,9(11):312.

https://doi.org/10.3390/v9110312

[138]

MAI JL, LIANG BS, XIONG ZL, AI XL, GAO F, LONG Y, YAO SW, LIU YF, GONG ST, ZHOU ZW.Oral administration of recombinant Bacillus subtilis spores expressing Helicobacter pylori neutrophil-activating protein suppresses peanut allergy via up-regulation of Tregs[J].Clinical and Experimental Allergy,2019,49(12):1605-1614.

https://doi.org/10.1111/cea.13489

[139]

KITAGAWA K, GONOI R, TATSUMI M, KADOWAKI M, KATAYAMA T, HASHII Y, FUJISAWA M, SHIRAKAWA T.Preclinical development of a WT1 oral cancer vaccine using a bacterial vector to treat castration-resistant prostate cancer[J].Molecular Cancer Therapeutics,2019,18(5):980-990.

https://doi.org/10.1158/1535-7163.MCT-18-1105

[140]

ZHOU J, LI MY, CHEN QF, LI XJ, CHEN LF, DONG ZL, ZHU WJ, YANG Y, LIU Z, CHEN Q.Programmable probiotics modulate inflammation and gut microbiota for inflammatory bowel disease treatment after effective oral delivery[J].Nature Communications,2022,13(1):3432

https://doi.org/10.1038/s41467-022-31171-0

[141]

张一帆, 林思思, 吴冯, 刘尽尧.基于表面修饰的活体细菌药物及递送策略[J]. 生命科学,2023,35(3):315-328.

ZHANG YF, LIN SS, WU F, LIU JY.Fabrication and delivery of live bacterial therapeutics by surface modification[J].Chinese Bulletin of Life Sciences,2023,35(3):315-328 (in Chinese).

[142]

PATARROYO JL, FLOREZ-ROJAS JS, PRADILLA D, VALDERRAMA-RINCÓN JD, CRUZ JC, REYES LH.Formulation and characterization of gelatin-based hydrogels for the encapsulation of Kluyveromyces lactis-applications in packed-bed reactors and probiotics delivery in humans[J].Polymers,2020,12(6):1287.

https://doi.org/10.3390/polym12061287

[143]

LUAN Q, ZHANG H, WANG JH, LI Y, GAN MY, DENG QC, CAI LY, TANG H, HUANG FH.Electrostatically reinforced and sealed nanocellulose-based macrosphere by alginate/chitosan multi-layer coatings for delivery of probiotics[J].Food Hydrocolloids,2023,142: 108804.

https://doi.org/10.1016/j.foodhyd.2023.108804

[144]

CHENG QK, LIU L, XIE MZ, LI H, MA D, XUE W.A colon-targeted oral probiotics delivery system using an enzyme-triggered fuse-like microcapsule[J].Advanced Healthcare Materials,2021,10(8):e2001953.

https://doi.org/10.1002/adhm.202001953

2024年第64卷第11期

PDF下载

130

引用本文

BibTeX

文章信息

doi: 10.13343/j.cnki.wsxb.20240307

接收时间：2024-05-16
首发时间：2026-03-21
出版时间：2024-09-30

补充材料

相关文章

文章信息

作者

出版历史

收稿日期：2024-05-16
录用日期：2024-08-22

基金

National Key Research and Development Program of China(2021YFC2102700)

国家重点研发计划(2021YFC2102700)

National Natural Science Foundation of China(32272269)

国家自然科学基金(32272269)

作者信息

天津科技大学生物工程学院, 天津 300457

通讯作者:

参考文献

分享链接

https://castjournals.cast.org.cn/joweb/wswxb/CN/10.13343/j.cnki.wsxb.20240307

分享至

全文二维码

扫描看全文

引用本文

BibTeX

本文的引用情况

2种不同金属材料的力学参数

科 Family	属数 Number of genus	种数 Number of species	占总种数比例 Percentage of total species (%)	属 Genus	种数 Number of species	占总种数比例 Percentage of total species (%)
鹅膏菌科Amanitaceae	2	11	5.26	鹅膏菌属 Amanita	10	4.78
小菇科 Mycenaceae	2	12	5.74	丝盖伞属 Inocybe	5	2.39
多孔菌科 Polyporaceae	8	14	6.70	蜡蘑属 Laccaria	5	2.39
红菇科 Russulaceae	3	23	11.00	小皮伞属 Marasmius	6	2.87
				小菇属 Mycena	11	5.26
				光柄菇属 Pluteus	5	2.39
				红菇属 Russula	17	8.13
				栓菌属 Trametes	5	2.39

关闭全屏

BibTeX
EndNote
RefWorks
TxT

Oral delivery vehicles	Drug loading mode	Characteristic
Bacillus surface display system	Recombination surface display	No need to produce antigens; different antigens can be displayed
	Non-recombinant surface display	Highly efficient; exhibits antigen in its natural form; requires cleavage to exhibit antigen
Yeast microcapsule system	Electrostatic adsorption	Protects drugs from destruction; accurately targets drugs; efficiently loads all types of drugs; safe and non-toxic
	Non-covalent force modification
	Passive diffusion/hydrophobicity
	Layer-by-layer synthesis (LBL)
	Post-derivatization combination
Recombinant probiotic expression system	Surface display	Stable binding; no need to penetrate cells; high efficiency
	Secrete	High delivery efficiency; avoids drug toxicity to cells
	Intracellular	Avoids environmental destruction of drugs; toxic to cells
Bacterium-like particles, BLPs	Surface display	Safe and stable; high drug loading; insignificant therapeutic immunization effect
Bacterial ghosts, BGs	Anchors and fills in internal spaces	Safe and stable; efficient targeting; its structure decreases drug concentration

Oral delivery vehicles

Drug loading mode

Characteristic

Bacillus surface display system

Recombination surface display

No need to produce antigens; different antigens can be displayed

Non-recombinant surface display

Highly efficient; exhibits antigen in its natural form; requires cleavage to exhibit antigen

Yeast microcapsule system

Electrostatic adsorption

Protects drugs from destruction; accurately targets drugs; efficiently loads all types of drugs; safe and non-toxic

Non-covalent force modification

Passive diffusion/hydrophobicity

Layer-by-layer synthesis (LBL)

Post-derivatization combination

Recombinant probiotic expression system

Surface display

Stable binding; no need to penetrate cells; high efficiency

Secrete

High delivery efficiency; avoids drug toxicity to cells

Intracellular

Avoids environmental destruction of drugs; toxic to cells

Bacterium-like particles, BLPs

Surface display

Safe and stable; high drug loading; insignificant therapeutic immunization effect

Bacterial ghosts, BGs

Anchors and fills in internal spaces

Safe and stable; efficient targeting; its structure decreases drug concentration

Type	Exogenous proteins/drugs	Anchoring proteins/attachment	Applications	References
Recombination	CsPmy	CotC	Oral vaccine for protection of grass carp against Ascaris lumbricoides	[42]
	Glucagon-like peptide-1 (GLP-1)	CotC	GLP-1 (28−36) oral delivery system for the treatment of type 2 diabetes mellitus	[35]
Non-recombinant	Grass carp reovirus (GCRV) Vp7 antigen	CotB & CotC	Enhancement of protective immunity to GCRV in grass carp	[43]
	OmpK or green fluorescence protein (GFP)	CotY	Enhancement of zebrafish immunogenicity to Vibrio pathogen infections	[44]
	Helicobacter pylori urease B	CotC	Oral administration of urease-carrying B spores prevents Helicobacter pylori infection	[45]
	Protective antigen (PA)	Adsorption	Development of an oral vaccine for the surface presentation of PA for the treatment of anthrax	[46]
	PfCSP	Coupling	Oral vaccines for malaria prevention	[47]
	TTFC	Adsorption	Effective in inducing an immune response and can be used as an adjuvant for vaccine immunization	[48]

Type

Exogenous proteins/drugs

Anchoring proteins/attachment

Applications

References

Recombination

CsPmy

CotC

Oral vaccine for protection of grass carp against Ascaris lumbricoides

[42]

Glucagon-like peptide-1 (GLP-1)

CotC

GLP-1 (28−36) oral delivery system for the treatment of type 2 diabetes mellitus

[35]

Non-recombinant

Grass carp reovirus (GCRV) Vp7 antigen

CotB & CotC

Enhancement of protective immunity to GCRV in grass carp

[43]

OmpK or green fluorescence protein (GFP)

CotY

Enhancement of zebrafish immunogenicity to Vibrio pathogen infections

[44]

Helicobacter pylori urease B

CotC

Oral administration of urease-carrying B spores prevents Helicobacter pylori infection

[45]

Protective antigen (PA)

Adsorption

Development of an oral vaccine for the surface presentation of PA for the treatment of anthrax

[46]

PfCSP

Coupling

Oral vaccines for malaria prevention

[47]

TTFC

Adsorption

Effective in inducing an immune response and can be used as an adjuvant for vaccine immunization

[48]

Protein expression mode	Bacterial strain type	Applications	References
Surface display	S. cerevisiae	Expression of avian influenza HA protein for use as an oral vaccine against avian influenza	[91]
	L. plantarum	Expression of Clostridium nucleatum outer membrane FomA protein inhibits IBD	[92]
	L. plantarum	Expression of Staphylococcus aureus non-toxic mutant alpha hemolysin, protective against Staphylococcus aureus lung and skin infections	[93]
	E. faecalis	Separate expression of four Eimeria aphidis proteins of Tennessee and protection against homologous infection in chickens by combined immunization	[94]
	Bifidobacterium	Expression of WT1 protein for use as an oral vaccine against oral cancer	[95]
	L. plantarum	Expression of the influenza virus antigen HA1-DCpep, which can play an immunizing role against influenza virus	[96]
	L. casei	Expression of AHA1-CK6 and VP2 proteins and oral immunization induces prevention of infectious pancreatic necrosis in rainbow trout	[97]
	L. casei	Expression of DCpep, Col, and CoE antigenic fusion proteins effectively induces an immune response against porcine epidemic diarrhea virus	[98]
	L. casei	Expression of an OmpK fusion CTB protein for use as an adjuvant against Vibrio anisopliae infection in crucian carp	[99]
Secrete	P. pentosaceus	Secretion of therapeutic protein P8 for tumor suppression and regulation of intestinal microflora	[100]
	L. paragasseri	Express and efficiently secrete epitope-4	[101]
	B. subtilis	Expression of the multi-epitope protein OmpC-FliC-SopF-SseB-IL-18 induces a strong immune response against Salmonella enteritidis	[102]
	L. lactis	Expression of extracellular TGFβR2 with significant anti-hepatic fibrosis effects	[103]
	L. lactis	Expression of soluble CD80 enhances anti-tumor immunity to inhibit tumor growth	[104]
Intracellular	L. casei	Expression of an antigen directed against SARS-CoV-2 Omicron variant B.1.1.529	[105]
	L. lactis	Expression of heat shock protein 65 and inhibition of atherosclerosis	[106]
	L. lactis	Expression of Helicobacter lipoprotein Lpp20 produces significant immune effects	[107]

Protein expression mode

Bacterial strain type

Applications

References

Surface display

S. cerevisiae

Expression of avian influenza HA protein for use as an oral vaccine against avian influenza

[91]

L. plantarum

Expression of Clostridium nucleatum outer membrane FomA protein inhibits IBD

[92]

L. plantarum

Expression of Staphylococcus aureus non-toxic mutant alpha hemolysin, protective against Staphylococcus aureus lung and skin infections

[93]

E. faecalis

Separate expression of four Eimeria aphidis proteins of Tennessee and protection against homologous infection in chickens by combined immunization

[94]

Bifidobacterium

Expression of WT1 protein for use as an oral vaccine against oral cancer

[95]

L. plantarum

Expression of the influenza virus antigen HA1-DCpep, which can play an immunizing role against influenza virus

[96]

L. casei

Expression of AHA1-CK6 and VP2 proteins and oral immunization induces prevention of infectious pancreatic necrosis in rainbow trout

[97]

L. casei

Expression of DCpep, Col, and CoE antigenic fusion proteins effectively induces an immune response against porcine epidemic diarrhea virus

[98]

L. casei

Expression of an OmpK fusion CTB protein for use as an adjuvant against Vibrio anisopliae infection in crucian carp

[99]

Secrete

P. pentosaceus

Secretion of therapeutic protein P8 for tumor suppression and regulation of intestinal microflora

[100]

L. paragasseri

Express and efficiently secrete epitope-4

[101]

B. subtilis

Expression of the multi-epitope protein OmpC-FliC-SopF-SseB-IL-18 induces a strong immune response against Salmonella enteritidis

[102]

L. lactis

Expression of extracellular TGFβR2 with significant anti-hepatic fibrosis effects

[103]

L. lactis

Expression of soluble CD80 enhances anti-tumor immunity to inhibit tumor growth

[104]

Intracellular

L. casei

Expression of an antigen directed against SARS-CoV-2 Omicron variant B.1.1.529

[105]

L. lactis

Expression of heat shock protein 65 and inhibition of atherosclerosis

[106]

L. lactis

Expression of Helicobacter lipoprotein Lpp20 produces significant immune effects

[107]