Up-regulation of inducible heat shock protein-70 expression in multiple sclerosis patients

Abstract

Inducible heat shock protein (HSP)70 (HSP70-1A and HSP70-1B proteins) is a chaperone responsible for assisting proper protein folding. Following stress conditions, HSP70 is highly up-regulated to mediate cytoprotective functions. In addition, HSP70 is able to trigger innate and adaptive immune responses that promote the immune recognition of antigens and to act as a cytokine when it is released. The data in the literature are controversial with regard to expression studies in peripheral blood mononuclear cells (PBMCs). In the present study, we aimed to examine if alterations of HSP70-1A/B expression are involved in the autoimmune pathogenesis of multiple sclerosis (MS). We determined both mRNA and protein expression in PBMCs of MS patients and healthy donors (HDs). We found a baseline increased expression of the HSPA1A gene in PBMCs from MS patients compared with HDs. Gene expression findings were associated with an increased protein expression of HSP70-1A/B in T lymphocytes (CD4+ and CD8+) and monocytes from MS patients under basal conditions that may reflect the immunological activation occurring in MS patients. We also provided evidence that heat shock (HS) stimulus induced HSP70-1A/B protein expression in HDs and MS patients, and that?HS-induced HSP70-1A/B protein expression in monocytes correlated with the number of T2 lesions at baseline in MS patients. However, after lipopolysaccharide inflammatory stimulus, monocytes from MS patients failed to induce HSP70-1A/B protein expression. Our data hint at altered immune responses in MS and may indicate either a state of chronic stress or increased vulnerability to physiological immune responses in MS patients.     

Humoral immune response to the chlamydial heat shock proteins hsp60 and hsp70 in Chlamydia -associated chronic salpingitis with tubal occlusion

The aim of this study was to evaluate the prevalence of serumimmunoglobulin (Ig) G and IgA antibodies to recom-binant chlamydial60 kDa heat shock protein (C-hsp60) and to assess the prevalenceof serum IgG antibodies to recombinant chlamydial 70 kDa heatshock protein (C-hsp70) in Chiamydia-associated chronic salpingitisand/or salpingitis isthmica nodosa with tubal occlusion. Infertilepatients (n = 34) with Chlamydia-associated, histologicallydocumented chronic salpingitis and/or salpingitis isthmica nodosaand bilateral tubal occlusions (group I) were compared withinfertile patients (n = 19) without tubal occlusions (groupII). The prevalence of chlamydial antigen in endocervical, urethraland urine samples was low in both groups. The median chlamydialserum IgG and IgA antibody titres were significantly higherin group I than in group II (P < 0.0001 and P = 0.0002 respectively).Serum IgG antibodies to C-hsp60 and C-hsp70 were detected in24 out of 34 patients (71%) in group I compared with 10 outof 19 (53%) and nine out of 19 (47%) patients in group II (notsignificantly different). There was a significant difference(P = 0.035) between the prevalences of serum IgA antibodiesto C-hsp60 in groups I (seven out of 34 patients; 21%) and II(none of the 19 patients). The association between the presenceof serum IgA antibodies to C-hsp60 and Chlamydia-ossocmted chronicsalpingitis and/or salpingitis isthmica nodosa with tubal occlusionunderlines the significance of chlamydial 60 kDa heat shockprotein in the pathogenesis of tubal infertility.     

Localization of heat shock protein 27 (hsp27) in the rat gingiva and its changes with tooth eruption

Heat shock protein 27 kDa (Hsp27) functions as a molecular chaperon to prevent apoptosis as well as to contribute to the regulation of cell proliferation and differentiation during development. In the present study, the localization of Hsp27 in the oral epithelium of rats and its expression change during formation of the gingiva with the tooth eruption were examined immunohistochemically to elucidate the roles of Hsp27 in the oral mucosa.In adult rats, Hsp27-immunoreactivity was localized in the prickle and granular layers but absent in the basal and horny layers of the oral epithelium. On the other hand, in the outer and sulcular epithelia of the free gingival, Hsp27-immunoreactivity was detected in the whole layers, while it was not found in the proliferation zone of the junctional epithelium immunoreactive for Ki67. In immature rats on 10th postnatal day, Hsp27-immunoreactivity was intense in the prickle and granular layers of the oral epithelium, but was not detected in its basal layer. In rats at the eruptive phase on 15th postnatal day, Hsp27-immunoreactivity was detected in sites of the basal layer adjacent to where the dental cusps penetrated through the oral epithelium. Although the immunoreactivity for Ki67 was found in the basal layer of the oral epithelium, it was not localized in the Hsp27-immunopositive sites of tooth-penetration in the basal layer. Just after the tooth-eruption on 20th postnatal day, Hsp27-immunoreactivity was not found in the stratified squamous epithelium at the dentogingival junction, whereas it was intense in a single layer of cuboidal epithelial cells attached to the tooth neck. Ki67-positive cells were scattered in the stratified squamous epithelium at the dentogingival junction, whereas no positive cells were found in the portion of a single layer of cuboidal epithelial cells.These findings suggest that the outer and sulcular epithelia of the free gingiva have a relatively slower rate of proliferation than other gingival and oral epithelia, and that Hsp27 might inhibit the proliferation of the basal cells. Such specific phenomenon in the free gingiva occurred immediately after the dental cusps were exposed to the oral cavity.     

细胞外热休克蛋白70的释放机制及免疫功能

热休克蛋白（heat shock protein，HSP）是机体细胞在受到热应激或其他应激状态下合成增多的一类蛋白质，在细胞的多种功能中起十分重要的作用。最近研究发现，正常人和多种疾病的循环血液和体液中存在热休克蛋白70。而且还发现这种存在于细胞外的热休克蛋白70（extracellular heat shock protein 70，eHSP70）可能作为免疫系统的“危险信号”（danger signal），调节免疫细胞的功能。     

热体克蛋白70在肝细胞癌中的表达及其意义

目的　探讨热休克蛋白 70 (HSP70 )在肝细胞癌 (HCC)中的表达及其意义。方法　采用免疫组化技术对 4 4例HCC和癌旁组织中HSP70的表达进行检测。结果　HCC中HSP70阳性率明显高于癌旁组织 (阳性率分别为 6 8.2 %和2 7.3% ,χ2 =7.3,P <0 .0 1)。HSP70表达与癌周淋巴细胞浸润 (χ2 =3.2 ,P >0 .0 5 )和转移 (χ2 =2 .3,P >0 .0 5 )无关 ,但与癌组织分化程度有关 (χ2 =4 .5 ,P <0 .0 5 )。结论　HSP70的异常表达与HCC的发生、发展有关 ,且可能是HCC发展、恶化的重要标志     

HSP70在慢性乙型肝炎患者肝组织中的表达及意义

目的探讨热休克蛋白 70 ( HSP70 )在慢性乙型肝炎肝组织中的表达及意义。方法采用免疫组化技术对 3 6例慢性乙型肝炎和 2 0例正常肝组织中 HSP70的表达进行检测。结果慢性乙型肝炎和正常肝组织中肝细胞 HSP70表达阳性率分别为 4 5 %和 15 % ,两者相比差异显著 ( χ2 =6.3 ,P<0 .0 5 ) ;中度和重度肝炎 HSP70阳性率明显高于轻度肝炎 (阳性率分别为 62 .5 %和 3 0 % ,χ2 =3 .9,P<0 .0 5 ) ;HSP70阳性细胞多位于灶性和碎屑样坏死区。结论肝细胞 HSP70的异常表达在慢性乙型肝炎免疫保护中起重要作用 ,可作为肝组织损伤的一种标志。     

热休克蛋白70对荷瘤鼠的治疗作用

目的研究肿瘤来源的热休克蛋白70(heat shock protein 70,HSP70)对荷瘤小鼠的治疗作用,为其应用提供参考依据.方法细胞培养、蛋白质提纯技术、western-blot法、毛细管电泳技术和动物实验等.结果应用5μgHSP70能延长小鼠的生存期限,平均生存(28.7±4.5)d,与对照组(18.5±3.9)d相比差异显著(P＜0.05);而10μg治疗组的小鼠生存期为＞(62.3±29.1)d,40%获长期生存(＞90d),所接种肿瘤完全消退.与其他组相比,差异具显著性(P＜0.01).结论肿瘤来源的HSP70具有明确的治疗作用,本研究对于研究应用肿瘤来源的热休克蛋白70治疗人类恶性肿瘤具有较重要的参考意义.     

热休克蛋白70、90在子宫内膜癌中的表达

目的:探讨子宫内膜癌组织中热休克蛋白(HSP)70、90的表达及意义。方法:用免疫组化Envision法及图象分析仪,检测 30例正常子宫内膜、30例增生过长子宫内膜和53例子宫内膜癌中HSP70、HSP90的表达。结果:子宫内膜癌中HSP70表达的灰度值为(209.06±5.36),明显高于正常内膜[(145.21±4.09),P<0.01]和增生过长内膜[(148.59±4.23),P<0.01];子宫内膜癌中HSP90表达的灰度值为(166.98±5.71),明显低于正常子宫内膜[(208.57±31.14),P<0.05]和增生过长子宫内膜[(249.73±4.94),P<0.01]。子宫内膜癌中HSP70的表达随肿瘤病理分级的增加而增强(P<0.01),非内膜样癌(229.90±3.77)较内膜样癌表达强[(198.37±3.15),P<0.01];子宫内膜癌中HSP90表达随肿瘤病理分级的升高而表达减弱,非内膜样癌(140.21±3.22)较内膜样癌表达减弱[(176.59±2.79),P<0.01]。子宫内膜癌中HSP70、90的表达与肌层浸润深度、临床分期及淋巴结转移未见显著相关性(P>0.05)。结论:HSP70、90可能与子宫内膜癌的发生及预后有关。     

Bleomycin induces the hsp 70 heat shock promoter in cultured cells

Bleomycin-induced lung disease is characterized by cell injury followed by fibroblast proliferation. Cells respond to injury by synthesizing a family of heat shock proteins. These proteins are critical to cell survival, and those of the 70,000 MW group (hsp 70) are essential for cell division and proliferation. To evaluate the effect of bleomycin on heat shock gene expression, we transfected a gene construct containing the hsp 70 heat shock gene promoter into fibroblasts. Doses of bleomycin, which have previously been shown to augment lung fibroblast proliferation, induce the hsp 70 heat shock promoter in the transfected cells. Bleomycin did not induce the expression of a non-hsp promoter placed in cells as a control of nonspecific gene activation. These observations suggest that bleomycin exposure may cause significant alterations in important DNA promoter regions such as the hsp 70 promoter and point to new ways to assess bleomycin-induced changes in cells.     

妊娠早期小鼠子宫内膜热休克蛋白70的免疫组化研究

目的 :研究妊娠早期小鼠子宫内膜热休克蛋白 70的表达。方法 :采用免疫组织化学方法。结果 :热休克蛋白 70主要存在于子宫内膜固有层的基质细胞及蜕膜细胞 ,内膜上皮、腺上皮中未见表达。与未孕小鼠相比 ,孕鼠热休克蛋白 70免疫反应阳性细胞显著增多 (P <0 .0 1 )且随妊娠日龄的增加而增加 (P <0 .0 1 )。结论 :热休克蛋白70可能参与了子宫内膜蜕膜反应中基质细胞的增殖 ,与蜕膜反应密切相关     

肿瘤热休克蛋白70联合IL-2对荷瘤小鼠的治疗作用

目的:研究肿瘤热休克蛋白70(HSP70)与IL-2联合应用对荷瘤小鼠的治疗作用。方法:用液相色谱法纯化小鼠肿瘤细胞 株中的HSP70。对纯化产物用SDS-PAGE及Western blot进行定性分析,再用毛细管电泳鉴定其纯度。通过动物实验,观察HSP70与IL-2单独应用及联合应用的抗肿瘤作用。结果:HSP70与IL-2联合应用较单独应用的治疗效果更明显,但治疗作用仍以HSP70为主。单独应用IL-2只能延长小鼠的生存期限[平均为(36.6±13.0)d],而HSP70 10μg组可使40％荷瘤小鼠的肿瘤最终全部消退,生存期最长者>90[平均生存期(>59.2±29.6)d],与对照组相比较差异显著(P<0.01)。HSP70与IL-2联合应用组可使60％荷瘤小鼠的肿瘤完全消退,平均生存期(>70.8±26.5)d,与对照组相比差异十分显著(P<0.01)。结论:合适剂量的HSP70与IL-2联合应用,对荷瘤小鼠有明显的治疗作用,可明显抑制肿瘤进展,提高生存率。以上结果对研究人类恶性肿瘤的免疫治疗具有较大的参考价值。     

Lack of cytotoxic activity against Mycobacterium leprae 65-kD heat shock protein (hsp) in multibacillary leprosy patients.

Cytotoxic T cells play an important role in host defence mechanisms, as well as in the immunopathology of leprosy. In this study, we evaluated whether Mycobacterium leprae hsp18, hsp65 and Myco. tuberculosis hsp71 could induce cytotoxic T cell activity against autologous macrophages pulsed with these hsp. Paucibacillary (PB) patients and normal controls generated more effector cells than multibacillary (MB) patients with all three hsp tested. There was no cross-reactivity between any of the hsp tested. Mycobacterium leprae hsp65 induced cytotoxic responses only in those MB patients undergoing an erythema nodosum leprosum (ENL) episode. Although hsp65 and hsp18 induced similar proliferation in MB patients, a high proportion of these patients did not generate cytotoxic effector cells in response to hsp65. Hence, those T cells reacting to hsp65 may play an important role in the control of Myco. leprae infection.     

结核杆菌热休克蛋白70基因的克隆与原核表达

目的：克隆结核杆菌热休克蛋白70(TBhsp70)基因，并在大肠杆菌中表达。方法：利用PCR技术从结核杆菌H37Rv中扩增Hsp70基因，并将其克隆到pUC19中，进行测序。将得到的Hsp70基因克隆到表达载体pGEX-4T-1中，构建重组表达质粒pGEX—TBhsp70，在大肠杆菌DH5α中进行表达。结果：成功地克隆了TBhsp70基因。DNA测序证实，与GenBank公布的序列一致。含pGEX-TBhsp70基因表达质粒的大肠杆菌经IPTG诱导后，能够表达相对分子质量(MR)约为96000的融合蛋白。结论：获得了TBhsp70基因，成功地构建了原核表达质粒pGEX-TBhsp70，并在大肠杆菌得到表达，为其相关研究奠定了一定的基础。     

Circulating antibodies to the 60-kD heat shock protein (hsp) family in patients with Helicobacter pylori infection

Whilst the mechanism by which Helicobacter pylori causes different gastroduodenal diseases is uncertain, strains producing the cytotoxin-associated protein (CagA) have greater pathogenicity. Hsps are immunogenic molecules induced by inflammatory mediators. The aim of this study was to assess pathogenicity of hsp antibodies in H. pylori-infected patients. ELISA techniques were used to assay sera of H. pylori-positive patients with gastritis, gastric atrophy, duodenal or gastric ulcer, and H. pylori-negative controls, for antibodies to CagA and to human, mycobacterial, and in 20 sera, H. pylori (hspB) 60-kD hsp. IgA antibodies to mycobacterial hsp60 in atrophy patients were elevated compared with patients with gastritis (P < 0.05) and with H. pylori-negative controls (P < 0.0005). IgA antibodies to human hsp60 in gastric atrophy patients were elevated compared with H. pylori-negative controls (P < 0.05). Patients with atrophy (P < 0.0005) and gastritis (P < 0.05) who were CagA-positive had raised titres of anti-mycobacterial hsp60 IgA antibodies compared with controls. IgA antibody levels to hspB were positively correlated with those to mycobacterial hsp60 (mhsp60) (P < 0.05) and human hsp60 (hhsp60) (P < 0.005). IgA antibodies to hsp60 are associated with gastroduodenal disease, particularly gastric atrophy, in H. pylori-infected patients. Increased humoral responses to hsp60 could either contribute to gastric atrophy or result from greater gastric mucosal damage induced by CagA-positive strains of H. pylori.     

热休克蛋白47和热休克蛋白60在小鼠胚胎器官形成过程中的表达

目的：研究小鼠胚胎器官形成过程中热休克蛋白47（Hsp47）和热休克蛋白60（Hsp60）的表达情况。方法：于GD11～GD18,取得小鼠胚胎脑、眼、心、肺、胃、肝、四肢,于GD13-GD18取得肾,利用RT-PCR方法半定量检测Hsp47和Hsp60在各器官的表达丰度。结果：Hsp47在GD11～GD12和GD18的脑、GD11～GD12和GD17～GD18的眼、GD11～GD12和GD16～GD18的肺、GD11-GD18的肝脏不表达,在其余时间和其余器官均有表达;Hsp60在CD11～GD18时段胚胎的脑、眼、心、肺、胃、肝、肾（GD13～GD18）、四肢中均有表达。结论：Hsp47和Hsp60在小鼠胚胎器官形成过程中有不同的表达丰度,其表达模式也不一致;Hsp47在小鼠胚胎发育过程中可能有重要功能,Hsp60则具有广泛表达的特点。     

牛珀至宝微丸对内毒素休克肺组织核转录因子-κB表达的调控

目的观察牛珀至宝微丸对内毒素休克肺损伤时核转录因子-κB表达的影响。方法静脉注射内毒素(LPS)1.5mg/kg、腹腔注射D-氨基半乳糖(D-GalN)100mg/kg造成内毒素休克模型,用牛珀至宝微丸作预治疗处理,免疫组化方法检测NF-κB在肺组织内的表达。结果LPS组阳性表达部位在细胞核,牛珀至宝微丸组表达部位主要在细胞质。牛珀至宝微丸降低NF-κB表达,肺损伤减轻。结论证实牛珀至宝微丸能降低内毒素休克时肺组织NF-κB表达,改变其表达部位,提示牛珀至宝微丸对内毒素休克造成的肺损伤的保护作用可能是通过调控肺组织NF-κB而产生的。     

eHSP的研究进展

热休克蛋白(heat shock protein,HSP)一直被认为是细胞内蛋白质,在胞内通过帮助蛋白质正确折叠、组装以及促进变性蛋白质降解而发挥生物学功能.HSP能被主动地分泌到胞外,成为细胞外HSP(extracellular HSP,eHSP),但其分泌及作用机制尚未定论.HSP偶然释放到外周循环是通过细胞坏死,...