成人接种重组(酿酒酵母)乙肝疫苗和重组(汉逊酵母)乙肝疫苗后效果观察

目的观察重组(酿酒酵母)乙肝疫苗和重组(汉逊酵母)乙肝疫苗在成人中的免疫效果。方法对HBsAg、抗HBs、抗HBc三项指标全部为阴性、且未接种过乙肝疫苗的162名16岁以上人群,按照0、1、6的免疫程序接种10μg重组(酿酒酵母)乙肝疫苗和重组(汉逊酵母)乙肝疫苗,于全程免疫1个月后采血,测定血清中抗HBs水平。结果接种重组(酿酒酵母)乙肝疫苗的99人,抗HBs阳转率为94.95%,几何平均浓度为331.08mIU/mL;接种重组(汉逊酵母)乙肝疫苗的63人,抗HBs阳转率为98.41%,几何平均浓度为270.79mIU/mL。两组间抗HBs阳转率和几何平均浓度差异均无统计学意义(P>0.05)。接种两种疫苗均未观察到明显的不良反应。结论重组(酿酒酵母)乙肝疫苗和重组(汉逊酵母)乙肝疫苗均具有良好的安全性和免疫原性。     

Dose and Load Studies for Subcutaneous and Oral Delivery of Poly(lactide-co-glycolide) Microspheres Containing Ovalbumin

Poly(lactide-co-glycolide) microspheres containing different loads of OVA (0.05, 0.1, 0.5 and 1.0% w/w) were manufactured by a w/o/w emulsion/solvent evaporation method. Low load efficiencies of less than 20% were observed. Normal size distributions with mean volume diameters ranging from 3.7 to 4.7 µm were obtained for different batches. The in vitro release of OVA from different loaded microspheres showed an expected burst release with all batches. The in vivo dose study (1, 10, 25, 50 µg of OVA) was performed by subcutaneous and oral inoculation in mice by single (0 week) or double (0 and 3 weeks) administration of PLGA 50/50 microspheres containing 0.1% OVA. Subcutaneous administration showed an immune response (serum Ig levels by ELISA) statistically (Fishers paired t-test; P < 0.05) above OVA saline negative controls at 3, 6 and 12 weeks after administration. Oral administration of microspheres produced statistically higher systemic immune responses at the higher doses. Single and double inoculation orally and subcutaneously produced similar serum antibody levels. The in vivo load study was performed by subcutaneous and oral administration to mice of 25 µg OVA contained in various loaded (0.05, 0.1, 0.5 and 1.0% w/w) microspheres. Serum immune responses at 3, 6, and 12 weeks after inoculation were statistically above OVA saline controls and were inversely proportional to the OVA load using either route. This observation suggested a relationship between the number of microspheres delivered and the in vivo serum response. Single subcutaneous administration of 0.05 or 0.1% OVA loaded PLGA 50/50 microspheres induced larger immune responses compared with complete Freunds adjuvant.     

治疗性乙型肝炎疫苗长期毒性试验

目的：根据我国相关法规,开展治疗性乙肝疫苗的长期毒性试验.方法：将治疗性乙肝疫苗以高、中、低三个剂量经皮下注射食蟹猴,另以等体积生理盐水作为空白对照,每天给药1次,连续21 d,停药后继续观察4周.以血常规、尿常规、血生化、血清抗体和补体、病理学等为观察指标.结果：各组动物外观、行为活动未见异常,未见死亡;试验组动物对受试疫苗均产生免疫应答,血清中可检测到相应抗体;高剂量治疗性乙肝疫苗对食蟹猴肝、肾功能指标有一定影响,停药后可恢复;未见明显的迟发性毒性反应.结论：治疗性乙型肝炎疫苗安全性良好,对食蟹猴的最大无毒作用剂量为21μg·kg-1,约为人临床拟用剂量的21倍,以体表面积计,约为单次人临床拟用量的8倍.     

Immune Augmentation of Single Contact Hepatitis B Vaccine by Using PLGA Microspheres as an Adjuvant

The present study was aimed to replace the alum type adjuvant for hepatitis B vaccine. The hepatitis B vaccine was encapsulated in poly (DL-lactide-co-glycolide) microspheres by solvent evaporation technique. The formulated microspheres were characterized in terms of morphology, particle size analysis, in vitro release study and in vivo immune response in male Wistar rats. The FT IR spectrum illustrates the characteristics bands of poly (DL-lactide-co-glycolide) microspheres and hepatitis B vaccine at 1750 cm^-1 and 1650 cm^-1, respectively. The hepatitis B vaccine loaded poly (DL-lactide-co-glycolide) microspheres were able to release antigens till day 42. Significant enhancement of specific antibodies to HBsAg was produced till day 90 after a single administration of HBsAg encapsulated poly (DL-lactide-co-glycolide) microspheres. However, the conventional alum adsorbed hepatitis B vaccine was not found to produce any significant specific antibody levels till day 90 after a single dose. The results showed that poly (DL-lactide-co-glycolide) microspheres show potential as an adjuvant for hepatitis B vaccine.     

乙型肝炎病毒前S1抗原检测方法及其临床意义探讨

用乙型肝炎病毒(HBV)前S1蛋白合成肽(21～47aa)免疫家兔制备多克隆兔抗前S1抗体,建立前S1抗原ELISA检测方法,对前S1蛋白合成肽的最低检出限为0.1μg/L,可测到2.5×10~5个/Dane颗粒。特异性好,用兔抗前S1抗体可以抑制前S1蛋白合成肽、基因表达前S1蛋白、纯化Dane颗粒和前S1抗原阳性血清与包被抗体的结合。本方法精密度高,批内变异和批间变异的变异系数均不超过0.15。在不同人群中,前S1抗原检出率以慢性活动性乙型肝炎最高。急性乙型肝炎时,前S1抗原与HBeAg平行存在,比HBsAg消失早,作为反映病毒清除的指标优于HBsAg。前S1抗原与HBV-DNA的相关性优于HBeAg,可作为病毒复制与传染性的标志。     

慢性乙型肝炎治疗性疫苗研究进展

目前慢性乙型肝炎（CHB）抗病毒治疗的预定目标是“功能性治愈”。用目前可及的治疗药物,只有小部分CHB患者得以实现。针对乙肝病毒（HBV）的特异性免疫反应功能失调被认为是导致HBV持续感染的主要原因,调节宿主免疫系统以加强特定的细胞免疫反应可能有助于清除HBV。治疗性疫苗,包括蛋白质（HBsAg/preS和HBcAg）、DNA和基于病毒载体的疫苗,能够诱导和加强针对HBV的特异性免疫反应。至今已进行50多项临床试验来评估CHB治疗中的治疗性疫苗,其中一些结果显示出免疫治疗的潜力。综述慢性HBV感染治疗性疫苗有关的基础和临床研究进展,讨论目前临床试验不理想的结果和未来策略。     

A review of multiple approaches towards an improved hepatitis B vaccine

Background: Hepatitis B is a DNA virus that can cause liver inflammation, cirrhosis, and cancer in chronically infected and symptomatic carriers. Antiviral treatments are usually limited in their effectiveness in treating the disease states. Vaccination against hepatitis B in pediatric and adolescent populations has proven to be a generally effective means for preventing diseases that could be potentially caused by this virus. Some 5 – 10% of the vaccinees do not develop protective immunity against the virus. Therefore, a significant amount of effort has been made in many research laboratories across the world to increase the potency of the vaccine by various innovative means, e.g., increasing the immunogenicity of the antigen or through introduction of novel adjuvants that elicit strong humoral and cell-mediated immune responses. Objectives/methods: The objective of this review is to highlight publications of significant developments that have been made over the past decade and efforts that are continuing towards producing an improved vaccine. A number of patents that protect novel hepatitis B vaccine formulations, including those claiming novel hepatitis B core antigen formulations and combinations of a vaccine with small molecule therapeutics, are discussed. Conclusion: There have been promising developments in the area of new adjuvants and delivery systems. The practical need for reducing the total number of childhood vaccinations has driven development of, and patent filings on, multivalent and combination vaccine formulations in which the hepatitis B vaccine is included as one component. Efforts and some advances have also been made in the critical area of therapeutic application of the vaccine. The existence of a large population of already infected patients and the inadequacy of most of the current antiviral drugs against hepatitis B diseases have also inspired efforts to produce a vaccine that would be efficacious in clearing an exiting infection.     

电感耦合等离子体原子发射光谱仪（ICP-AES）测定重组酵母乙肝疫苗中硫柳汞的含量

目的 ICP-AES测定重组酵母乙肝疫苗中硫柳汞的含量。方法样品消化后直接用ICP-AES检测。结果线性相关性好,重复性实验变异系数均小于4%,样品回收率超过99%。结论 ICP-AES能够满足重组酵母乙肝疫苗中硫柳汞含量检测的技术要求,是一种快速、有效、准确的分析方法。     

尼索地平微球的制备

采用溶剂挥发法制备了尼索地平微球 ,考察了制备工艺中影响微球质量的 5个主要因素 ,筛选出较理想的处方和工艺。所得微球形态圆整 ,表面光滑 ,粒径 (18.2± 3.8) μm,载药量 2 1.2 % ,包封率 85 .4 %。     

冻干治疗用母牛分枝杆菌菌苗治疗慢性乙型肝炎疗效分析

目的：观察冻干治疗用母牛分枝杆菌菌苗（微卡）治疗慢性乙型肝炎（CHB）中的有效性和安全性。方法：CHB60例分成2组。治疗组：33例，主要用药为母牛分枝杆菌菌苗（微卡）＋乙肝疫苗＋双嘧达莫。对照组：27例，常规保肝治疗，主要用药为肌苷＋葡糖醛酸＋维生素C。6个月为1个疗程。结果：治疗组HBV-DNA、HBeAg阴转12例（36．36％），无不良反应。两组比较差异有显著性（P〈0．05）。结论：冻干治疗用母牛分枝杆菌菌苗治疗CHB安全有效，无不良反应。     

乙肝病毒抗原在肝组织的表达及意义

目的 :了解乙肝病毒感染者肝组织中乙肝病毒(HBV)抗原表达与病理分型及HBV复制的关系。方法 :采用免疫组织化学二步法检测肝组织HBsAg和HBcAg ,用ELISA法检测血清中HBV标志物。结果 :(1)HBV感染者肝组织中HBsAg和HBcAg检出率分别为79.75%和73.01 %。(2)乙肝病毒携带者 (AsC)HBsAg表达以浆型为主 ,膜型检出率较低。 (3)HBcAg浆型表达在重度肝病变高于轻度肝病变 ,而核型表达则相反。除中、重度外 ,各组间比较均有显著性差异 (P<0.01)。 (4)肝组织HBsAg、HBcAg 表达在血清HBeAg阳性组与HBeAb阳性组和HBeAg 和HBeAb均阴性组间有显著性差异 (P<0.01)。结论 :浆型HBsAg和核型HBcAg 多见于病变相对静止的病例 ,而膜型HBsAg 和浆型HBcAg 与病变活动程度明显相关。在血清HBeAg与HBeAb均阴性时 ,HBV仍有可能进行复制。HBcAg 作为主要的靶抗原 ,在肝脏免疫损伤中起重要作用     

乙型肝炎卡介苗联合疫苗的制备

在常规制备乙型肝炎疫苗和卡介苗技术基础上制备联合疫苗,建立了联合疫苗制备及冻干工艺,对冻干工艺的稳定性及其对抗原的影响进行了研究,采用动物实验进行免疫效果比较、安全性、局部毒性、过敏试验。结果表明冻干对两种抗原无影响;两种抗原具有良好的相容性;联合疫苗与同批卡介苗相比,效力无显著性差异(P>0.05);联合疫苗组与同批乙肝疫苗组全程免疫相比,统计学无显著性差异(P>0.05);安全性试验联合疫苗组与卡介苗组反应一致,均未见结核性病变。联合疫苗组与卡介苗组病变性质和病理演变过程相似,HBsAg并不增强卡介苗引起的炎症反应;未见全身过敏反应。连续9批成品检定全部合格,证明冻干工艺稳定,技术可行。     

静注人免疫球蛋白对新生儿乙肝核心抗体的影响

目的:探讨新生儿使用静注人免疫球蛋白(IVIG)后乙肝核心抗体(HBcAb)的变化。方法:采用回顾性研究方法,收集我院2019-2020年使用IVIG的住院新生儿病历,分析使用IVIG前后患儿HBcAb的变化。结果:共收集43例患儿,其中5例使用IVIG前后HBcAb均呈阳性,10例使用IVIG前后HBcAb均呈阴性,18例使用IVIG后HBcAb由阴性转为阳性,10例使用IVIG后因未进行乙肝5项复测未知HBcAb的变化。结论:输注IVIG会引起新生儿HBcAb水平升高,提示对输注IVIG的患儿进行抗体被动转移的排除具有重要意义。     

慢性乙型肝炎患者血清前Pre-S1抗原检测及其与肝功能关系的研究

目的检测慢性乙型肝炎病毒患者前S1(Pre-S1)抗原,探讨其与肝功能的关系。方法收集270例慢性乙型肝炎病毒患者血清,采用ELISA方法检测Pre-S1抗原;采用AU2700全自动生化分析仪检测丙氨酸氨基转移酶(ALT)和天冬氨酸氨基转移酶(AST)。结果 270例慢性乙型肝炎病毒患者中,Pre-S1抗原阳性中ALT异常检出率为91.1%,AST异常检出率为93.0%,Pre-S1抗原中阴性中ALT异常检出率为28.3%,AST异常检出率为35.4%,两组结果有显著性差异(P<0.05)。结论慢性乙型肝炎患者中,Pre-S1抗原可作为判断乙型肝炎病毒(HBV)感染,复制及对肝细胞损伤的另一个指标之一。     

不同剂量重组(酵母)乙肝疫苗对小鼠细胞免疫应答的研究

目的：探索不同剂量重组(酵母)乙肝疫苗对小鼠细胞免疫调节作用。方法：将HBsAg免疫Balb／C小鼠一周后取淋巴结制备淋巴细胞悬液，经HBsAg刺激培养48h后检测其诱导产生的IL-2、IFN-γ的水平，另一份细胞同样经HBsAg刺激培养56h后，用氚-胸腺嘧啶核苷(^3H-TdR)标记16h后检测T细胞增殖的水平。用不同剂量HBsAg免疫小鼠后，检测血清中抗HBsIgG2a的水平。结果：中、高剂量组小鼠产生的IL-2、IFN-γ、抗HBsIgG2a以及T细胞增殖的水平显著高于低剂量组和对照组。结论：一定剂量的重组(酵母)乙肝疫苗可上调小鼠的细胞免疫功能。     

Study of the Influence of Several Stabilizing Agents on the Entrapment and In Vitro Release of pBC 264 from Poly(Lactide-Co-Glycolide) Microspheres Prepared by a W/O/W Solvent Evaporation Method

Pharmaceutical Research -     

新型乙型肝炎病毒表面抗原中蛋白核酸疫苗的免疫原性研究

目的 观察新型乙型肝炎病毒（HBV）表面抗原中蛋白（MHBs)核酸疫苗的免疫原性。方法 以新型人体应用载体质粒pSW3891构建MHBs核酸疫苗（pSW3891/MHBs/adr)，对照组和实验组Balb/c小鼠分别基因枪法免疫对照载体质粒（pSW3891)和MHBs核酸疫苗，采用酶联免疫吸附试验检测抗HBs,LDH释放测定法检测小鼠特异性细胞毒T淋巴细胞（CTL）杀伤活性。结果 MHBs核酸疫苗可在体外高效表达，免疫小鼠后可产生高滴度抗HBs，血清阳性终点滴度达1：97200,小鼠脾细胞HBs特异性细胞毒性T淋巴细胞（CTL）杀伤活性达78.81%。结论 新型MHBs核酸疫苗在Balb/c小鼠实验中表现出良好的体液和细胞免疫原性。     

游离HCV核心抗原检测对丙型肝炎早期筛选价值的研究

目的 分别检测游离丙肝病毒核心抗原(HCV-cAg)、抗体(HCV-Ab)及HCV RNA,研究游离丙肝病毒核心抗原检测早期筛选HCV感染的价值.方法 采用ELISA法测定丙肝病毒核心抗原和抗体,FQ-RT-PCR方法测定HCV RNA,对比300例HCV RNA阴性正常查体人群结果,筛选有危险因素人群HCV阳性者35例,研究丙肝病毒核心抗原测定的特异性和敏感性及缩短检测窗口期的价值.结果 体检人群300例HCV-Ab、HCV RNA均为阴性, HCV-cAg阴性299例(99.67%); 35例随访确诊HCV感染者初次HCV RNA阳性标本, HCV-cAg阳性33例(94.29%), HCV-cAg检测可有效缩短窗口期.结论 游离HCV核心抗原ELISA测定法具高度敏感性及特异性,可以明显缩短HCV窗口期,可作为HCV抗体检测的补充试验.     

Novel Biomimetic Reconstituted Built-in Adjuvanted Hepatitis B Vaccine for Transcutaneous Immunization

Transcutaneous immunization is the administration of a vaccine on the skin to generate efficient systemic and mucosal immune responses against an antigen. In the present study, reconstituted hepatitis B surface antigen vesicles (HBsAg-REVs) integrated with monophosphoryl lipid A were prepared by the delipidation-reconstitution method and tested as built-in adjuvanted vaccine, system for transcutaneous immunization using a combined approach of tape strippings, and enhanced antigen skin contact time. Prepared vesicles were extensively characterized for size, shape, zeta potential, and antigen protein loading efficiency. Following topical application, HBsAg-REVs skin permeation on isolated rat skin and cell uptake by bone marrow–derived dendritic cells were determined by confocal laser scanning microscopy and flow cytometry, respectively. The humoral and cellular immune responses elicited by HBsAg-REVs via transcutaneous immunization were comparable to the marketed intramuscular hepatitis B vaccine formulation with predefined immunization protocols. This study supports that delivery of reconstituted HBsAg vesicles via transcutaneous route may open a new vista for designing topical vaccines with possible immune protection against hepatitis B in future.     

基因重组乙型肝炎疫苗用于成人免疫效果

目的了解基因重组乙型肝炎疫苗用于成人的免疫效果。方法对经筛查乙肝病毒表面抗原(HBsAg)、乙肝病毒表面抗体(抗-HBs)均阴性的一般状况良好的18~60岁成人240例,用基因重组乙肝疫苗20μg按0、1、6个月免疫程序免疫,静脉采血检测乙肝病毒表面抗体(抗-HBs)。结果全程免疫后1个月时,抗-HBs阳性率84.6%,抗-HBs几何平均滴度(GMT)为74.65m IU/m l;6个月时,抗-HBs阳性率68.5%,抗-HBs几何平均滴度(GMT)为34.6 m IU/m l。结论对成人开展乙肝疫苗接种选用基因重组(CHO细胞)乙肝疫苗20μg是安全有效的。