重组乳酸菌抗原递送方式及黏膜免疫效果研究进展

 乳酸菌是一类能利用碳水化合物产生乳酸的革兰阳性菌的统称.作为重要的益生菌,乳酸菌已被广泛应用到食品工业、生物制药、疫苗研制等各个领域,是公认安全的食品级微生物.随着基因工程技术的发展,乳酸菌已被广泛用作基因工程宿主菌,用于表达多种生物多肽和蛋白,并成为向黏膜免疫系统递送抗原的理想菌株.抗原的细胞定位可影响其免疫原性,近年来一系列递送系统的建立,可将异源蛋白和抗原靶向表达于乳酸菌胞质、细胞壁和胞外培养基.本文就重组乳酸菌这三种抗原递送方式及其对蛋白表达量和免疫原性的影响作一综述. 更多     

日本的乳酸菌制剂

日本的乳酸菌制剂应用较早,在30年代日本武田药厂即生产乳酸菌制剂“表飞鸣”,用于肠内异常发酵、消化不良与腹泻。1961年日本药局方第七版收载了乳酸菌制剂“牛酪乳粉”(Puluis Lactis Deflo-ratus)作为药典品用于医疗。“表飞鸣”是乳酸菌的     

内皮型一氧化氮合酶在2型糖尿病大鼠主动脉的表达及罗格列酮干预效果

目的：观察2型糖尿病(T2DM)大鼠血清内皮素、一氧化氮水平，主动脉病理变化、内皮型一氧化氮合酶( eNOS)蛋白及其mRNA的表达，探讨噻唑烷二酮类药物的干预作用。方法雄性Wistar大鼠80只，随机分为对照组、高脂组、模型组、治疗组各20只，制备T2DM大鼠模型后，治疗组予罗格列酮灌胃治疗，分别于给药6和12周时取样进行血糖、血脂、内皮素、一氧化氮的检测，观察光镜下主动脉的病理变化，主动脉eNOS蛋白及其mRNA表达情况。结果给药6、12周高脂组、模型组及治疗组较对照组内皮素升高，一氧化氮降低( P<0．01)。给药12周时模型组较高脂组和治疗组内皮素升高，一氧化氮降低( P<0．05)。给药12周时模型组一氧化氮水平较给药6周时下降( P<0．05)。给药12周时高脂组、模型组和治疗组大鼠主动脉出现不同程度病理改变。给药6和12周时，主动脉eNOS的蛋白和mRNA表达高脂组、模型组和治疗组较对照组下调，且12周时模型组较高脂组下调( P<0．01)。结论高血糖、高血脂可导致内皮素升高和一氧化氮下降，以及eNOS蛋白及其mRNA表达异常，从而引起动脉硬化，噻唑烷二酮类药物对血管内皮功能、病理形态、eNOS蛋白及其mRNA表达异常有一定逆转作用。     

蛋白和多肽药物的透粘膜吸收

随着蛋白和多肽药物的增多，其非注射给药剂型的研究受到了越来越多的重视，本文综述了国外对鼻腔、口腔、口服、直肠和阴道几种主要透粘膜吸收给药途径的研究。蛋白和多肽药物的透粘膜给药剂型的研究推进了蛋白和多肽药物的临床应用。     

促红细胞生成素对大鼠局灶性脑缺血再灌注损伤的影响

目的：观察脑缺血再灌注大鼠腹腔注射人重组促红细胞生成素（Epo）后，脑组织切片的病理形态及细胞凋亡指标caspase-3蛋白的变化。方法：应用线栓法制备大鼠大脑中动脉缺彬再灌注模型，应用免疫组化染色、原位末端标记法（TUNEL），检测脑缺血再灌注后各组大鼠凋亡细胞数和caspase-3蛋白的表达，经图像分析仪计算凋亡细胞数及测量阳性反应细胞光密度。结果：①治疗组海马CAl区和皮层神经细胞较缺血再灌组数目明显减少；②TUNEL法检测凋亡细胞数与caspase-3蛋白阳性反应细胞数统计学分析显示：再灌注组与假手术组比较、给药组与缺血再灌注组比较均有极显著性差异（P〈0．01），多次给药组与一次给药组比较差异显著（P〈0．05）。结论：促红细胞生成素对局灶性脑缺血再灌注大鼠具有神经保护作用，可能与其明显减少海马及皮层神经元细胞的凋亡数量以及降低caspase-3蛋白的表达有关。     

多西环素对乳腺癌小鼠移植瘤生长及转移的影响

【摘要】目的 通过检测多西环素对小鼠乳腺癌肺转移模型（BCML-TAⅡ99）中基质金属蛋白酶-9(MMP-9)及金属蛋白酶组织抑制因子1(TIMP-1)蛋白表达水平,揭示其对该肿瘤生长及转移的影响。方法建立BCML-TAⅡ99 模型,应用免疫组织化学SP法检测MMP-9和TIMP-1蛋白在多西环素给药组与对照组肿瘤组织中的表达情况,并进行比较。结果与对照组相比,给药组肿瘤转移率明显低于对照组;MMP-9蛋白在给药组表达下调,而TIMP-1蛋白表达上调,差异具有统计学意义（P<0.05）,且MMP-9与TIMP-1表达呈负相关（P<0.05）。结论多西环素可通过调节MMP-9及TIMP-1蛋白表达水平,对BCML-TAⅡ99模型中肿瘤生长及转移发挥抑制作用。     

红花注射液对肺纤维化小鼠的干预作用

目的探究红花注射液对肺纤维化小鼠的影响及其作用机制。方法 120只ICR小鼠随机分为4组,空白组、模型组、地塞米松(3 mg·kg~(-1))组、红花注射液(50 mg·kg~(-1))组,每组30只,其中10只用于观察存活率。空白组除外,其余3组鼻腔滴注博来霉素建立肺纤维化模型,造模后第1日起,各组给予相应的药物腹腔注射治疗。于给药7、 28 d取材,检测肺系数、肺组织羟脯氨酸(HYP)含量, ELISA和Western blot法检测肺组织中转化生长因子β1 (TGF-β1)蛋白表达水平。结果给药7、 28 d,与空白组比较,模型组肺系数、肺组织HYP含量、 TGF-β1蛋白表达水平均升高(P <0.01)。给药7、 28 d,与模型组比较,红花注射液组肺系数、 HYP含量、 TGF-β1蛋白表达水平均降低(P <0.01)。给药28 d,与地塞米松组比较,红花注射液组肺系数、 HYP含量、 TGF-β1蛋白表达水平均降低(P <0.01)。结论红花注射液能抑制博来霉素诱导的肺纤维化,其作用机制可能与降低肺组织TGF-β1蛋白表达水平有关。     

靶向给药大分子载体系统：关于生物分布中的药动学问题

本文从药动学角度述了大分子载体系统用于普通药物，蛋白及基因药物定位给药的现状及未来发展趋势。     

鼻腔给药治疗上呼吸道感染的进展

目的：介绍鼻腔给药治疗上呼吸道感染的临床应用情况。方法：通过查阅近年国内医药学刊物，总结鼻腔给药用于呼吸道感染治疗的献。结果：多种化学药物、中药以及生物缺点品都被用于鼻腔给药治疗上呼吸道感染，分别从缓解症状、抗感染等途径发挥局部或全身疗效。结论：鼻腔给药是治疗上呼吸道感染的一种新给药途径，具有作用迅速、应用方便及用药量小等特点，但尚需进一步深入研究。     

盐酸氨溴索、地塞米松对大鼠胎肺TGF-β3表达的影响

目的 比较产前给药盐酸氨溴索、地塞米松对大鼠胎肺转化生长因子-β3(TGF-β3)基因和蛋白表达的影响.方法 9只孕鼠随机分为生理盐水对照组、地塞米松组、盐酸氨溴索组,每组3只.孕16、17、18 d腹腔注射给药3剂.孕19 d取胎鼠肺, RT-PCR法检测TGF-β3基因表达水平;免疫组织化学法检测TGF-β3蛋白表达水平.结果 地塞米松组和盐酸氨溴索组TGF-β3基因和蛋白的表达均明显高于对照组 (P＜0.05).地塞米松组TGF-β3基因和蛋白的表达高于盐酸氨溴索组(P＜0.05).结论 产前给药盐酸氨溴索、地塞米松均能促进胎肺发育相关因子TGF-β3基因及蛋白的表达,从而促进胎肺功能的发育.     

乳酸菌作为载体的异源蛋白表达及应用研究进展

乳酸菌是公认的食品级安全微生物,利用乳酸菌作为表达载体进行抗原表达,能诱导机体产生有效的免疫应答。经口服和滴鼻免疫还可诱导机体的黏膜免疫应答,且接种方便,免疫效果和依从性均优于传统的注射途径。此文就乳酸菌作为表达载体的优势、表达系统的组成、目前成功表达的异源蛋白以及新型传送系统革兰阳性菌增强基质颗粒的研究进展做一综述。     

Platelets as delivery systems for disease treatments

Platelets are small, anucleate, discoid shaped blood cells that play a fundamental role in hemostasis. Platelets contain a large number of biologically active molecules within cytoplasmic granules that are critical to normal platelet function. Because platelets circulate in blood through out the body, release biological molecules and mediators on demand and participate in hemostasis as well as many other pathophysiologic processes, targeting expression of proteins of interest to platelets and utilizing platelets as delivery systems for disease treatment would be a logical approach. This paper reviews the genetic therapy for inherited bleeding disorders utilizing platelets as delivery system, with a particular focus on platelet-derived FVIII for hemophilia A treatment.     

Biodegradable dextran hydrogels crosslinked by stereocomplex formation for the controlled release of pharmaceutical proteins

Hydrogels are based on hydrophilic polymers, which are crosslinked to prevent dissolution in water. Because hydrogels can contain large amounts of water, they are interesting devices for the delivery of proteins. In this contribution a biodegradable dextran hydrogel is described which is based on physical interactions and is particularly suitable for the controlled delivery of pharmaceutically active proteins. The unique feature of our system is that the preparation of the hydrogels takes place in an all-aqueous solution, by which the use of organic solvents is avoided. Furthermore, chemical crosslinking agents are not needed to create the hydrogels, since crosslinking is established physically by stereocomplex formation between enantiomeric oligomeric lactic acid chains. The hydrogel is simply obtained after mixing aqueous solutions of dextran(l)-lactate and dextran(d)-lactate. In this contribution, the formation of the hydrogels as well as their protein release properties and degradation behavior are discussed.     

Application of sialic acid/polysialic acid in the drug delivery systems

The properties of modified biomaterial are gaining more and more importance in drug delivery systems. Sialic acid (SA) and polysialic acid (PSA) serve as endogenous substances, which are non-immunogenic and biodegradable. At the same time, SA modification of the drugs/carriers can enhance the uptake of tumor cell and retention in brain; PSA modification can reduce the immunogenicity of the proteins or polypeptides and increase circulation time of the modified drugs/carriers in the blood, thus achieving active targeting effect. These properties offer a variety of opportunities for applications in drug delivery systems. This article summarizes the biological functions of SA and PSA and presents the technologies of SA/PSA modified small molecule drugs, proteins and carriers in drug delivery systems.     

Localized,non-viral delivery of nucleic acids: Opportunities,challenges and current strategies

Localized delivery of drugs is an emerging field both with regards to drug delivery during disease as well as in tissue engineering. Despite significant achievements made in the last decades, the efficient delivery of proteins and peptides remains challenging, especially in cases requiring long-term release of proteins after application. The localized delivery of nucleic acids (NA) represents an interesting alternative due to higher physicochemical stability of NA, increased efficiency by harnessing cells as bioreactors for the production of required proteins and improved versatility with regards to expression of specific proteins through plasmid DNA or repression of gene products through siRNA. However, unlike most proteins and peptides, NA must be delivered to the cytoplasm or nucleus to be efficacious, resulting in significant delivery challenges. We herein describe frequently used non-viral vectors for the delivery of NA including polyplexes, lipoplexes and lipopolyplexes and summarize recent developments in the field of nucleic acid delivery systems for local application based on hydrogels, solid scaffolds and physical delivery methods. The challenges associated with the different approaches are identified and options to address these challenges are discussed.     

Clostridium spores for tumor-specific drug delivery

Insufficient blood supply of rapidly growing tumors leads to the presence of hypoxia, a well-known feature in solid tumors. Hypoxia is known to decrease the efficiency of currently used anti-cancer modalities like surgery, chemotherapy and radiotherapy. Therefore, hypoxia seems to be a major limitation in current anti-cancer therapy. The use of non-pathogenic clostridia to deliver toxic agents to the tumor cells takes advantage of this unique physiology. These strictly anaerobic, Gram-positive, spore-forming bacteria give, after systemic administration, a selective colonization of hypoxic/necrotic areas within the tumor. Moreover, they can be genetically modified to secrete therapeutic proteins like cytosine deaminase or tumor necrosis factor-alpha. The specificity of this protein delivery system can be further increased when expression is controlled by the use of a radio-inducible promoter, leading to increased spatial and temporal regulation of protein expression. This approach of bacterial vector systems to target protein expression to the tumor can be considered very safe since bacteria can be eliminated at any moment by the addition of proper antibiotics. The Clostridium-based delivery system thus presents an alternative therapeutic modality to deliver anti-tumor agents specifically to the tumor site. This high selectivity offers a major advantage in comparison with the classical gene therapy systems.     

Rumen-stable delivery systems

Ruminants have a distinct digestive system which serves a unique symbiotic relationship between the host animal and predominantly anaerobic rumen bacteria and protozoa. Rumen fermentation can be both beneficial by enabling utilization of cellulose and non-protein nitrogen and detrimental by reducing the nutritive value of some carbohydrates, high biological value proteins and by hydrogenating unsaturated lipids. In addition it can also result in the modification and inactivation of many pharmacologically active ingredients administered to the host animal via the oral route. The advances in ruminant nutrition and health demand a rumen-stable delivery system which can deliver the active ingredient post-ruminally while simultaneously meet efficacy, safety and cost criteria. In contrast to drug delivery systems for humans, the demand for low-cost has hindered the development of effective rumen-stable delivery systems. Historically, heat and chemical treatment of feed components, low solubility analogues or lipid-based formulations have been used to achieve some degree of rumen-stability, and products have been developed accordingly. Recently, a polymeric pH-dependent rumen-stable delivery system has been developed and commercialized. The rationale of this delivery system is based on the pH difference between ruminal and abomasal fluids. The delivery system is composed of a basic polymer, a hydrophobic substance and a pigment material. It can be applied as a coating to solid particles via a common encapsulation method such as air-suspension coating. In the future, the delivery system could be used to deliver micronutrients and pharmaceuticals post-ruminally to ruminant animals. A further possible application of the delivery system is that it could also be combined with other controlled delivery devices/systems in order to enhance slow release or to achieve targeted delivery needs for ruminants. This paper discusses the rumen protection and the abomasal release mechanism of the polymeric coating. It also reviews other rumen stable delivery systems and methods for evaluating their in vitro and in vivo performance.     

菌影在蛋白亚单位疫苗和DNA疫苗递送中的应用

菌影（bacterialghost,BG）是由噬菌体ФX174E基因表达蛋白介导革兰阴性菌裂解所形成的空胞,无胞质内容物,但保留了天然的胞壁结构。由于BG具有颗粒特性并含有天然表面抗原组分,因此易于被抗原呈递细胞识别并激发体液和细胞免疫应答。BG可负载外源蛋白和DNA,并可通过各种免疫途径进行接种。因此,BG是一种有潜力的疫苗递送系统。     

Microbially controlled drug delivery to the colon

The human gastrointestinal tract consists of a highly complex ecosystem of aerobic and anaerobic microorganisms that plays a significant role in the metabolism of nutrients as well as drugs. In the colon, bacteria ferment various types of substrates that are not susceptible to digestion in the small intestine. This arouses interest in specific drugs, drug delivery systems, and prodrugs that escape small bowel digestion, arrive intact, and are absorbed or degraded in the large bowel. For the past forty years, experience has been gained with the azo prodrug of 5-amino salicylic acid, salazopyrine, which is cleaved by colonic bacteria to its parent drug. Some laxative drugs were also reported to degrade into active metabolites in the colon. Lately equally interesting and more sophisticated microbial controlled delivery systems, have been developed based on similar principles.     

Lung surfactant as a drug delivery system

Lung surfactant is a complex mixture of mainly phospholipids and proteins. The composition leads to a unique spreading effect of the surfactant as well as spontaneous vesicle formation, which may be favourable characteristics of a drug delivery system for pulmonary delivery. The aim of study was to investigate the potential use of the surfactant extract, HL10 (LeoPharma, DK) as a drug delivery system. Studies involved incorporation of hydrophilic- and amphipathic model drugs (sucrose and acylated peptides) into HL10 and elucidation of the influence of surfactant proteins on the HL10 behaviour. Results showed that HL10 vesicles did not retain sucrose indicating formation of leaky vesicles. Studying the influence of surfactant proteins on release from DPPC-liposomes showed tendencies toward a protein-induced release. Hence, the surfactant proteins may influence the membrane lipid packing and characteristics resulting in leakiness of the membranes. Incorporation of acylated peptides into HL10 depended on the chain length rendering a successful incorporation of the peptide acylated with C14-acyl chains. This study suggests that HL10 may be a promising drug delivery system for the pulmonary delivery of amphipathic drug substances, e.g. therapeutically active acylated peptides (e.g. acylated insulin).