热应激诱导兔脑热休克蛋白70的合成研究

探讨热应激与热休克诱导的脑组织不同细胞热休克蛋白７０合成。     

热应激对A549细胞HSP70、XPA修复酶表达的影响

目的研究热应激作用下A549细胞热休克蛋白70（HSP70）和XPA修复酶的表达规律及其可能的意义。方法利用免疫印迹方法（Western blot）检测A549细胞中HSP70、XPA修复酶的表达水平。结果A549细胞接受不同温度（39、41、42、43℃）处理2h后,HSP70表达水平随温度的增高而逐渐增加,并在42℃应激时表达最高,与对照组相比有统计学意义（P〈0.05）;细胞接受42℃应激1～4h,HSP70表达水平随应激时间的增加而递增,与对照组相比,细胞在42℃应激2～4h时HSP70表达显著上调（P〈0.05）。两处理组细胞XPA修复酶表达水平均无明显变化（P〉0.05）。结论高温能诱导HSP70的表达,并呈一定的温度-效应和时间-效应关系,而对XPA修复酶未见明显影响。     

热休克蛋白70与肝癌

热休克蛋白70(HSP70)广泛存在于细胞中,在应激刺激后过表达,可以参与蛋白质的合成及受损蛋白的修复。HSP70在肝癌治疗中的作用日益受到重视:利用其特异性和免疫原性,靶向治疗肝癌。并且可以利用其分子伴侣的特点,作为肝癌疫苗,诱发机体抗肝癌的免疫反应。HSP70在肝癌治疗中已初显成效,但也存在很多问题,需要更加深入的研究。总之,HSP70在肝癌治疗中有很大的应用前景。     

热应激蛋白70及其抗体在中暑病人血浆中水平改变的观察研究

探讨中暑发生的病理过程中热应激蛋白７０（ＨＳＰ７０）水平及其抗体滴度水平的变化和相互关系。应用Ｗｅｓｔｅｒｎ Ｂｌｏｔ法测定血浆ＨＳＰ７０水平；应用免包被的酶联免疫吸附分析法经倍比稀释测定血浆ＨＳＰ７０抗体滴度。     

热休克蛋白70对肿瘤抑制作用的研究

1993年Ｕｄｏｎｏ从肿瘤组织中获得的热体克蛋白 (ｈｅａｔｓｈｏｃｋｐｒｏｔｅｉｎ ,ＨＳＰ) 70 (相对分子质量 66 0 0 0～ 780 0 0ｕ)具有肿瘤特异的免疫原性 ,ＨＳＰ70可作为提呈分子把肿瘤细胞内的内源性多肽提呈给杀伤细胞。还可通过γδＴ细胞杀伤肿瘤细胞[1 ] 。因此ＨＳＰ70也能通过诱导细胞因子发挥抗肿瘤作用[2 ] 。1　材料与方法1 .1　实验材料　伤寒杆菌 (0 90 1 )本实验室保存的菌种 ;Ｓ1 80及Ｈ2 2肿瘤细胞由大连医科大学提供 ;实验动物为昆明系小白鼠 ,体质量 1 8～ 2 2ｇ ,雄性 ,由本校实验动物中心提供 ;主要试剂为甘氨酸…     

热休克蛋白70融合肿瘤抗原基因的DNA疫苗研究进展

在抗肿瘤免疫治疗中,已经发展了多种形式的热休克蛋白70(HSP70)相关治疗性疫苗,这些疫苗主要包括蛋白疫苗、肽疫苗、细胞疫苗及DNA疫苗等,其中HSP70融合DNA疫苗凭借其自身的优势引起人们的关注。它不但可诱导高效、持久的抗肿瘤免疫应答,而且可以维持免疫记忆。本文就HSP70在肿瘤治疗性DNA疫苗中的作用和应用作一综述。     

热休克同源蛋白70羧基端作用蛋白的功能及其与神经退行性疾病的关系

神经退行性疾病是一组病因不明的慢性进行性神经损害.蛋白质的错误折叠和聚集是这类疾病共同的病理特征.热休克同源蛋白70羧基端作用蛋白(carboxyl-terminus of Hsc70 interacting protein,CHIP)具有双重功能,一方面可作为热休克蛋白的辅助因子,通过氮基端的34肽重复序列结构域与Hsp90和Hsc/Hsp70相互作用,调节热休克蛋白介导的蛋白质异常折叠,同时又可作为E3泛素连接酶,通过羧基端的U-box结构域参与泛素-蛋白酶体系统(ubiqutin-proteasome system,UPS)介导的蛋白质降解过程.CHIP在分子伴侣系统和UPS的交互作用中扮演重要角色,对维持细胞内蛋白质稳态具有重要的作用.本文从CHIP蛋白的分子结构特点、生理功能、在细胞代谢中的作用、与神经退行性疾病的关系等方面进行综述.     

人热休克蛋白70基因的原核表达

目的 表达人热休克蛋白70（HSP70）并进行鉴定。方法 用PCR方法扩增人HSP70基因片段，经T-A克隆法克隆到载体pUCm-T中，并进行DNA测序，将人HSP70基因片段从载体pUCm-T中酶切后，构建重组表达载体pGEX-4T-1-HSP70，并转化大肠杆菌JM109，用IPTG诱导，收集细菌，菌体裂解后进行SDS-PAGE及Western blot检测。结果 人HSP70基因的PCR产物约为1.9kb。序列测定结果证实，所获目的序列与文献[3]报道的相一致，经EcoRⅠ和XhoⅠ酶切鉴定证实。人HSP70基因已成功地克隆到表达载体pGEX-4T-1中，构建的表达载体pGEX-4T-1-HSP70，能很好地在大肠杆菌中表达相对分子质量（Mr)为96000并具有抗原特性的融合蛋白，结论 成功地克隆并表达了HSP70基因，为研究HSP70的结构。功能与临床应用提供了必要条件。     

热休克蛋白70的抗肿瘤免疫效应研究

目的 :研究HSP70诱导的抗肿瘤免疫效应 ,为应用其治疗人类恶性肿瘤提供实验依据。方法 :应用细胞培养、液相色谱法、电泳技术、Western blot法等获得高纯度肿瘤中的HSP70 ,将其应用于同源的 61 5系小鼠 ,观察其抗肿瘤免疫效应。结果 :HSP70免疫后的小鼠能够抵抗同种肿瘤细胞的攻击 ,其中 1 0 μg免疫后的小鼠 1 0 0 %获长期生存 (>90d) ,无一死于肿瘤 ,5μg免疫后的小鼠肿瘤进展受到抑制 (平均生存 2 8 2± 4 8d) ,但无一小鼠获长期生存 ,上述两组与对照组 (平均生存 1 8 3± 3 6d)相比差异均具有显著性 (P <0 0 1 )。HSP70对荷瘤小鼠的治疗研究证明 ,5μgHSP70有一定的治疗作用 ,荷瘤小鼠平均生存 31 3± 2 9d ,而 1 0 μgHSP70治疗组的小鼠生存期 >58 8± 33 7d ,其中 40 %的小鼠所接种肿瘤完全消退 ,获长期生存(>90d) ,与其他组相比 ,差异具有显著性 (P <0 0 1 )。结论 :HSP70可诱发强大的抗肿瘤免疫效应 ,并且有良好的治疗作用 ,对于研究利用HSP70治疗人类的恶性肿瘤有重要的参考意义。     

结核杆菌热休克蛋白肽段对胶原诱导性关节炎保护作用的研究

目的：研究结核杆菌HSP65肽段（P256-270）和结核杆菌HSP70肽段（P111-125）对类风湿关节炎动物模型——胶原诱导性关节炎（CIA）大鼠的保护性作用。方法：结核杆菌HSP肽段滴鼻免疫大鼠，观察其对CIA的防治效果，观察指标包括关节炎指数评分、关节病理学分析、酶联免疫斑点实验（ELISPOT）检测脾脏淋巴细胞分泌细胞因子（IL4、IFN-γ）水平、流式细胞术分析外周血T淋巴细胞亚群及酶联免疫吸附实验（ELISA）测定血清中抗CⅡ抗体水平。结果：分别经结核杆菌HSP65和HSP70肽段滴鼻免疫的实验组大鼠，与阳性对照组比较，关节炎指数明显下降（P〈0．05），病理变化减轻，抑制性细胞因子IL-4水平升高（分别为P〈0．05和P〈O．01），而炎性细胞因子IFN-1水平降低（P〈0．01），CD4^＋／CD8^＋T细胞比值降低（P〈0．05），血清中抗CIⅡ抗体水平也降低（分别为P〈0．05和P〈0．01）。结论：应用结核杆菌HSP65肽段或HSP70肽段能减轻CIA大鼠的症状和体征，其作用机制与调节T细胞的功能有关，此研究可为探讨人类RA新的免疫治疗方法提供依据。     

DNA vaccines encoding membrane‐bound or secreted forms of heat shock protein 70 exhibit improved potency

Traditional vaccine strategies are inefficient against challenge with complex pathogens including HIV; therefore, novel vaccine technologies are required. DNA vaccines are attractive as they are relatively cheap and easy to manufacture, but a major limitation has been their lack of immunogenicity in humans, which may be overcome with the incorporation of an adjuvant. HSP70 is a recognised damage‐associated molecular pattern, which is a potential adjuvant. We investigated the immunogenicity of a DNA vaccine encoding HIV gag and HSP70; the latter was genetically modified to produce cytoplasmic, secreted or membrane‐bound HSP70, the expression of which was controlled by an independent promoter. The DNA was administered to C57BL/6 mice to evaluate gag‐specific T‐cell responses. Our results demonstrated the ability of membrane‐bound and secreted HSP70 to significantly enhance gag‐specific T‐cell responses and increase the breadth of T‐cell responses to include subdominant epitopes. Membrane‐bound or secreted HSP70 also significantly improved the multifunctionality of HIV‐specific T cells and T‐cell proliferation, which is important for maintaining T‐cell integrity. Most importantly, the inclusion of membrane‐bound HSP70, secreted HSP70 or a combination significantly increased protection in mice challenged with EcoHIV, a chimeric virus that replicates in mouse leukocytes in vivo.     

A new role for CCR5 in innate immunity--binding to bacterial heat shock protein 70

CCR5 has been associated with adaptive immune responses, because of its expression on effector and memory T cells, and dentric cells. In this issue of the European Journal of Immunology, Whittall et al. identify CCR5 as a receptor that binds bacterial heat shock protein 70 (HSP70). Thus HSP70 may stimulate responses by CCR5+ T cells or dendritic cells, particularly at epithelial surfaces. These findings imply a role for CCR5 in innate immunity, in addition to its more established role as a chemoattractant receptor for adaptive immune responses.     

Response to hepatocarcinoma Hca-F of mice immunized with heat shock protein 70 from elemene combo tumor cell vaccine

To analyze immune response to murine hepatocarcinoma Hca-F of mice immunized with heat shock protein 70 （HSP70） derived from elemene combo tumor cell vaccine （EC-TCV） of Hca-F, HSP70 was isolated from EC-TCV by ADP affinity chromatography. Mice were immunized with HSP70 intraperitoneally three times and spleen calls were sampled. For cells, their proliferation and cytotoxicity against Hca-F were measured with MTT assay and their phenotypes were analyzed with flow cytometry. Spleen cells of immunized mice with HSP70 exhibited more potent cytotoxicity against Hca-F and proliferation than that of normal control mice, but less potent than that of mice immunized with EC-TCV. Among three groups, the percent of T6 T lymphocytes in the mice immunized with HSP70 （35.5%） was the highest compared with 6.25% in normal mice, and 28.4% in the mice immunized with EC-TCV. Immunization of HSP70 derived from EC-TCV could elicit potent immune response to Hca-F. HSP70 is one of elements inducing anti-tumor immune responses against Hca-F.     

The role of heat shock protein 70 induced by geranylgeranylacetone in carbon tetrachloride-exposed adult rat testes

Carbon tetrachloride (CCl₄) induces testicular damage, through formation of free-radical metabolites. Molecular chaperone heat shock protein of 70 kDa (HSP 70) protects cells from various stresses. This study was designed to investigate the potential role of induction of HSP70 using geranylgeranylacetone (GGA) on testicular damage caused by CCl₄. Rats were divided into group I (control group), Group II (CCl₄ group) received CCl₄ s.c. for 4 weeks, group III received CCl₄ s.c. for 4 weeks simultaneously with daily single oral dose of GGA (GGA – treated CCl₄ group). Serum testosterone, testicular lactate dehydrogenase (LDH) and alkaline phosphatase (ALP), testicular malondialdehyde (MDA), total nitrite and total antioxidant capacity (T-AOC) were measured. Evaluation of histopathological changes and immunohistochemical HSP70 expression for testicular biopsies were performed. Group II showed lower values of gonado-somatic index, serum testosterone, testicular LDH, ALP, T-AOC and greater values of testicular MDA and total nitrite than in control. Testicular morphology showed widening of seminiferous lumen, less spermatogenesis, vacuolization of germinative epithelium. Group III had higher values of gonado-somatic index, serum testosterone, testicular LDH, ALP, T-AOC with less testicular MDA and total nitrite than in group II. They have less damage and restored the altered testicular morphology. Immunohistochemical HSP70 expression was increased in the testicular spermatogenic and sertoli cells in group II that was significantly accentuated in group III. These findings suggest that GGA-induced activation of HSP 70 significantly alleviate CCl₄ inflicting testicular damage by HSP 70 mediated cytoprotection and antioxidant effects.     

HSP70与肿瘤相关性的研究进展

HSP70是HSP家族中最重要最保守的一类,不仅能够有效维持细胞结构的稳定以及提高细胞对应激原的耐受性,还参与了肿瘤细胞的发生、生长以及凋亡,与肿瘤免疫也有很大的相关性.目前,HSP70在临床上已开始应用于肿瘤的免疫治疗与基因治疗,并取得了一定的成效.本文就近年来HSP70与肿瘤相关性的研究展开综述.     

An enzyme‐linked immunosorbent assay for heat‐shock protein 70 in plants

Procedures are described for the production of antibodies against heat‐shock protein 70 (HSP70) from plants and for the development of an enzyme‐linked immunosorbent assay (ELISA) for the antigen in crude plant extracts. Competitive and two‐site sandwich variants of the ELISA are compared. Results are presented for a preliminary analysis of HSP70 levels in control and heat‐shocked mung bean shoots.