Circulating antibodies to inducible heat shock protein 70 in patients with uveitis

Heat shock proteins with molecular weight 70 kDa (hsp70) are highly conserved immunogenic intracellular molecules. There are two main subtypes: one is expressed constitutively (hsc70), while the other is induced under stressful conditions (ihsp70). Using an ELISA directed against recombinant human ihsp70, antibody titers were determined in patients with defined ocular inflammatory conditions (Behçet's disease, Vogt-Koyanagi-Harada (VKH), pars planitis, and sarcoidosis) as well as in a group of age-matched normal volunteers. In comparison to healthy controls (n = 14, absorbance 0.269), levels were significantly elevated in Behçet's disease (n = 18; 0.412), sarcoidosis (n = 15; 0.432), and pars planitis (n = 13; 0.346), but not in VKH (n = 10; 0.263). A correlation was also noted for treatment versus no treatment in pars planitis (p = 0.028), but not in other inflammatory conditions. There was no correlation with the level of intraocular disease activity as defined by vitreous haze and vision drop. Since pars planitis is a purely ocular condition, circulating levels of ihsp antibodies likely reflect the extent of disease involvement within the eye.     

Immunoreactivity of constitutive and inducible heat shock protein 70 in human fetal retina

The purpose of this study was to measure the level of expression of the inducible heat shock protein70 (Hsp70), the constitutive heat shock protein70 (Hsc70) in the outer nuclear layer and the photoreceptors in the human fetal retina. Fetal eyeballs were selected from fetal autopsy specimens of 12 and 17 to 40 week old fetuses, with no history of congenital anomalies. The retinas had differentiated from neuroblastic cells, into matured sensory retinas, from a gestational age (GA) from 12 to 36 weeks. The Hsp70 and Hsc70 were prominently expressed in the nuclear layers. The Hsc70 was expressed at all GAs and the Hsp70 was expressed from 20 to 33 weeks GA. This period is in accordance with the maturation of the sensory retina. The expression of heat shock protein may be regulated by the development of the fetus, and play an important role in the ocular development.     

Hsp25 and -90 immunoreactivity in the normal rat eye.

PURPOSE: The distributions of heat shock protein (Hsp)25 and -90 in various regions of the rat eye are described to provide a basis for understanding their roles in normal, damaged, and diseased ocular tissues. This work complements the earlier examination of Hsp70 and Hsc70 (the constitutive form). METHODS: Eyes of adult male Sprague-Dawley rats were fixed in methacarn and embedded in paraffin. Sagittal sections (10 microm) through the optic nerve were stained with hematoxylin and eosin, or incubated with anti-Hsp90, anti-Hsp25, or control IgG. Bound antibody was visualized using an avidin-biotin-horseradish peroxidase detection system. RESULTS: Hsp90 immunoreactivity was abundant in the retina, whereas only low levels of Hsp25 were detected there. In the optic nerve, the relative difference in immunoreactivity for the two Hsps was reversed, with Hsp25 being considerably greater than Hsp90. Both Hsps were detected at low levels in the retinal pigment epithelium (RPE), except for that portion within 250 microm of the optic disc, where Hsp25 and -90 immunoreactivities were increased. Similar to the optic nerve, the corneal epithelium showed greater staining for Hsp25 than for Hsp90, and basal cells contained the highest levels of immunoreactivity for both Hsps. In the ciliary body and iris, Hsp25 and -90 were abundant and similarly distributed in the epithelial and stromal layers. CONCLUSIONS: Each of the ocular tissues had distinctive patterns of Hsp25 and -90 immunostaining. These results suggest that the various structures of the eye have unique requirements for the particular chaperoning and supportive functions of these two Hsp families.     

热休克蛋白27和热休克蛋白70与视网膜视神经疾病关系的研究进展

热休克蛋白(heat shock protein,HSP)是细胞受到不同刺激时产生的一组高度表达的蛋白。这种蛋白在结构上高度保守,在功能上常被认为具有保护细胞的作用。近年来,有研究表明HSP27和HSP70具有保护视网膜视神经细胞的作用,而其相关抗体具有促进这些细胞凋亡的作用。在细胞培养和动物实验中,我们观察到阻断HSP27抗体和HSP70抗体的表达有利于保护视网膜视神经细胞。本文就HSP27和HSP70与视网膜视神经疾病的关系作一综述。     

大鼠钝挫性眼外伤动物模型中晶状体上皮细胞热休克蛋白70和27的表达及其热耐受的调节作用

目的探讨大鼠眼钝挫伤后晶状体上皮细胞HSP70和HSP27的表达,并给予预先的热刺激(热休克),观察热休克对晶状体上皮细胞HSP70和HSP27表达的调节。方法48只Sprague-Dawley(SD)大鼠,应用随机数字表法随机分成扑打组和水浴组,每组24只鼠,右眼为实验眼。扑打组:20g铁球20cm高度拍打眼球100次。水浴组:给大鼠体温提高至40.5~41.5℃8min诱导出热耐受,2~3h后再给予20g铁球20cm高度拍打眼球100次。逆转录聚合酶链反应检测晶状体上皮细胞HSP70和HSP27基因的表达丰度。结果钝挫性眼外伤可造成晶状体上皮细胞HSP70基因表达的增强,扑打眼球后1h HSP70表达升高至4.59±0.12,3h后达到高峰7.72±0.27,24h后降至正常1.32±0.14。预先的热刺激导致HSP70基因表达增高至1.83±0.30。两组HSP27基因表达丰度均无明显改变。结论钝挫性眼外伤中晶状体上皮细胞HSP70表达的增强提示HSP70可能在钝挫性外伤过程中对晶状体变性蛋白起保护作用,给予预先的热刺激可能通过提高HSP70的表达保护晶状体。(中华眼科杂志,2006,42:241-245)     

恒河猴经瞳孔温热疗法与热休克蛋白相关研究

目的探讨恒河猴经瞳孔温热疗法（transpupillary thermotherapy，TTT）治疗后视网膜和脉络膜组织热休克蛋白70（heat shock protein 70，HSPT0）表达。方法利用恒河猴做动物模型。于1TIrr治疗后不同时间点（1h、1d、1周、2周、1月、4月）摘除双眼固定。应用免疫组化技术，分析TTT治疗对猴眼视网膜和脉络膜组织HSP70的影响。结果TTT治疗后，于1d起，在强反应光斑边缘视网膜和脉络膜有HSP70的表达，4个月时仍有弱表达。弱反应光斑1d时，在视网膜全层及脉络膜均有HSP70表达，4个月时消失。同时,TTT治疗可以引起视网膜不同程度组织病理学的损害。结论TTT可以引起视网膜、脉络膜组织病理学损害。局部温度的升高会诱导视网膜、脉络膜内源性HSP70的产生，并且这种表达在激光治疗后1～4个月的猴眼中仍可以存在。     

CO2激光对豚鼠耳蜗Hsp70和诱生型一氧化氮合酶表达特点的研究

目的探讨CO2激光照射豚鼠耳蜗后Hsp70（heat shock proteins 70）和诱生型一氧化氮合酶（inducible nitric oxide synthetase，iNOS）表达的改变及与耳蜗听觉功能变化的关系。方法红目豚鼠36只，随机分为3组（A，B和C组），分别以功率为1，2，3W的CO2激光在豚鼠左耳耳蜗底周打孔，此为实验耳，右耳为对照耳。于术前1d及处死前检测听性脑干反应（auditory brainstem response，ABR）。术后即刻和术后3周分批断头处死豚鼠，以链霉亲和素-生物素-过氧化物酶复合物免疫组织化学法及图像分析技术观察激光照射后耳蜗Hsp70和iNOS的表达情况。结果术后即刻3组ABR反应阈值较术前明显升高（P均〈0．05），照射后3周A组ABR反应阈值基本恢复（P〉0．05），B，C组有所下降，但仍高于术前（P均〈0．05）；术后即刻3组耳蜗中Corti器和螺旋神经节Hsp70和iNOS表达较对照组明显增强（P均〈0．01），术后3周A组Hsp70和iNOS表达与对照组比较差异无统计学意义（P〉0．05），B，C组两者表达较对照组有所增强（P均〈0．05）。A组和B组术后即刻和术后3周、C组术后即刻ABR阈值的变化和Hsp70和iNOS阳性反应平均灰度值的变化呈负相关（P均〈0．05），而C组术后3周三者变化无相关性（P〉0．05）。结论CO2激光照射豚鼠耳蜗后Hsp70和iNOS表达增强，其表达程度与听觉功能改变密切相关。     

小分子热休克蛋白在眼部表达及其影响因素的研究进展

小分子热休克蛋白(small heat shock proteins,sHSPs)是热休克蛋白的亚单位,在各种应激状态下,它在组织中迅速表达增强,参与对组织的保护和损伤作用。sHSPs的研究已经受到国内外学者的广泛关注,认为其在作为分子伴侣、协同免疫、抑制凋亡、抗原提呈以及甾体激素受体功能等方面具有重要作用。充分认识sHSPs在眼部疾病中的功能及其作用机制,能进一步了解眼部疾病的发病机制,为疾病的治疗研究提供新的分子生物学依据。     

HSP70在STZ-糖尿病性白内障发病机制中的作用

目的：探讨HSP70在链脲佐菌素－糖尿病性白内障发生、发展中的作用。方法：将60只SD大鼠随机分为两组，正常对照组与白内障组。用链脲佐菌素（STZ）诱发糖性白内障，每周观察晶状体的变化，在实验开始后2w末、4w末、8w末，分别摘取眼球，检测热休克蛋白－70（HSP70）在晶状体上皮细胞（LECs）中的表达情况。结果：对照组晶状体一直保持透明，白内障组晶状在2w末出现空泡，8w末全部混浊。HSP70在对照组中未见表达，在白内障组中表达明显，并随着白内障的发展而增加，结论：HSP70可能通过调节LECs的生产在糖性白内障的发生，发展中起重要作用。     

实验性大鼠青光眼模型诱导热休克蛋白27的表达

目的 测定热休克蛋白（HSP）家族中HSP27、HSP70和HSP90在正常和实验性大鼠青光眼模型视网膜中的蛋白质水平，探索其与青光眼视神经病变的潜在关系。设计 实验性研究。研究对象 60只Wistar大鼠。方法 将60只大鼠随机分为高眼压组（30只），假手术对照组（30只），右眼为实验眼，左眼为对照眼。采用双极电凝器电凝巩膜表面3组静脉，建立高眼压模型，用Tonopen眼压计检测眼压，每周2次，取眼压在27～35mmHg的大鼠进行研究。在眼压升高第10、20和60天处死大鼠，匀浆提取视网膜蛋白质用于Western印迹分析，并用β-actin校正点样误差。主要指标 眼压、Western印迹分析。结果 Western印迹分析显示。在眼压升高第10、20及60天，视网膜HSP27的蛋白质水平平均为对照组的197％。在实验眼视网膜内未观察到HSP70、HSP90蛋白质水平的变化。结论 在高眼压状态下大鼠视网膜HSP27上调，而HSP70和HSP90无变化。HSP27为眼压升高导致的特定基因改变，其持续过度表达可能与青光眼视神经病变有关。     

热休克蛋白70的主要功能及与白内障发病的相关研究

热休克蛋白70(heat shock protein70,HSP70)在正常细胞中水平较低,而在应激后表达显著增高,是HSP中重要的成员。HSP的功能涉及细胞的结构维持、更新、修复、免疫等,但其基本功能为帮助蛋白质的正确折叠、移位、维持和降解。我们就热休克蛋白70的主要功能及其与白内障发病的相关研究进展作一简要概述。     

Association between heat shock protein 70/Hom genetic polymorphisms and uveitis in patients with sarcoidosis

PURPOSE: The predisposition to sarcoidosis, a systemic granulomatous disorder of unknown etiology, is genetically determined, and genetics appears also to drive the disease down distinct phenotypic pathways. This study was undertaken to test the hypothesis that sarcoidosis-related uveitis represents a genetically distinct disease subset, by investigating single nucleotide polymorphisms (SNPs) in the HSP-70/1 and HSP-70/Hom genes. HSP70 molecules play a key role in the immune response by functioning both as chaperones and as inducers of proinflammatory cytokine secretion. METHODS: By sequence-specific primers-polymerase chain reaction (SSP-PCR) five SNPs were evaluated in 270 white patients with sarcoidosis, including 88 with sarcoid-related uveitis, and in 347 matched control subjects. One hundred twenty-five patients with idiopathic anterior uveitis (IAU) and 56 with idiopathic intermediate uveitis (IIU) were also included in the study as disease control subjects. RESULTS: The HSP-70/Hom rs2075800 G allele frequency was higher in the sarcoid-uveitis group than in both the sarcoid-non-uveitis and control groups (83% vs. 71%, OR = 2.00, P(c) = 0.01; and 83% vs. 66%, OR = 2.45, P(c) = 0.00005, respectively). Similar results were observed when considering the carriage frequency of the associated haplotype (HSP-70 haplotype 2) across the three study groups (47% vs. 29%, OR = 2.17, P(c) = 0.03; and 47% vs. 21%, OR = 3.26, P(c) = 0.0003, respectively). In addition, the carriage frequency of the HSP-70 haplotype 2 discriminated among sarcoid-related uveitis, IAU, and IIU (47% vs. 19%, OR = 3.26, P(c) = 0.001; and 47% vs. 23%, OR = 2.81, P(c) = 0.04, respectively). CONCLUSIONS: A strong association was found between HSP-70/Hom rs2075800 G and uveitis in patients with sarcoidosis. Further studies are needed to understand the molecular mechanisms underlying this association.     

Protein s-glutathionylation in retinal pigment epithelium converts heat shock protein 70 to an active chaperone

A disulfide bond between key redox-sensitive cysteine residues and glutathione is one mechanism by which redox related allosteric effectors can regulate protein structure and function. Here we test the hypothesis that glutaredoxin-1 (Grx-1), a member of the oxidoreductase family of enzymes, may be a critical component of redox-sensitive molecular switches by mediating reversible protein S-glutathionylation and enzymatic catalysis of thiol/disulfide exchange. Deglutathionylation of a 70 kDa protein by Grx-1 was detected using a monoclonal antibody specific to protein S-glutathionylation. Heat shock cognate protein 70 (Hsc70) was identified as a substrate of Grx-1 through mass spectrometry. Recombinant Hsc70 was glutathionylated in vitro, and protein S-glutathionylation reversed by Grx-1. Glutathionylated Hsc70 was more effective in preventing luciferase aggregation at 43 degrees C than reduced Hsc70 in a dose dependent fashion. ATP did not effect the chaperone activity of Hsc70-SG but did increase the activity of reduced Hsc70-SG. Reversible glutathionylation of Hsc70 may provide a mechanism for post-translation regulation of chaperone activity.     

突发性聋临床特征与血清热休克蛋白70水平相关性的研究

目的检测突发性聋患者血清热休克蛋白70（HSP70）含量,并初步探讨它与突发性聋患者临床特征之间的关系。方法本研究包括54例突发性聋患者（患者组）和54例正常人（对照组）。用高敏的酶联免疫吸附测定法检测血清HSP70,并观察血清HSP70与突发性聋患者临床特征的相关性,这些临床变量包括年龄,性别,病耳,发病时程,伴随症状（有无眩晕、耳鸣、脑梗死）,发病初听力,治疗后听力,听力曲线类型,听力损失程度,听力恢复程度。结果突发性聋患者血清HSP70明显高于对照组,差异有统计学意义（P〈0．05）。血清HSP70水平与突发性聋患者听力恢复程度有相关性（P〈0．05）。结论血清HSP70可作为突发性聋患者发病初期的血浆标记物,推测HSP70可作为突发性聋患者的一个预后指标。     

三种热休克蛋白mRNA在高眼压大鼠视网膜中的表达

目的 探讨热休克蛋白家族(HSPs)中HSP27、HSP70和HSPg0 mRNA在高眼压大鼠视网膜中的表达水平,及其与青光眼视神经病变可能的潜在关系.方法 将30只Wistar大鼠随机分为高眼压组和伪手术对照组(各15只),右眼为实验眼,左眼为对照眼.采用双极水下电凝器电凝大鼠右眼巩膜表面3组静脉的方法建立高眼压动物模型,用Tonopen眼压计检测眼压,每周2次.高眼压组眼压保持在27-35 mmHg;对照组为12-17 mmHg.分别于第10、20、60 d处死大鼠.摘除眼球,剥离视网膜,提取RNA,Northem印迹分析.结果 Northern印迹分析显示：在眼压升高第10、20、60 d,视网膜HSP27的mRNA水平平均增高达250％,HSP70和HSPg0的mRNA水平平均增高达98％和82.6％,HSP27水平明显增高,HSP70,HSP90变化不大.结论 高眼压大鼠视网膜HSP27 mRNA表达水平持续增高,推测与高眼压状态下视网膜神经节退变有潜在关系.     

热休克蛋白27在实验性青光眼中表达的研究

目的　观察大鼠眼压升高后热休克蛋白 2 7(HSP2 7)在视网膜中的表达。方法　5 0只Wistar大鼠随机分为高眼压组和sham对照 (假手术 )组。采用电凝鼠巩膜表面至少3组静脉及角膜缘周围血管 ,减少房水静脉回流升高眼压。观察手术后 1、2、3、4及 8周大鼠眼压 ,同时免疫组化检测视网膜中HSP2 7的表达及分布情况。结果　高眼压组右眼术后眼压明显升高。术后 1周眼压 :(30 .12± 5 .18)mmHg(1kPa =7.5mmHg) ,1周后眼压基本稳定。术后各时间点高眼压组右眼眼压与术前、左眼及sham对照组右、左眼间比较 ,差异有显著性 (P <0 .0 0 1)。视网膜中HSP2 7阳性表达主要表现在视网膜神经节细胞的胞浆内及神经纤维层中 ,HSP2 7阳性表达率在术后 1、2、3、4及 8周 ,高眼压组右眼与左眼、sham对照组右、左眼间比较 ,差异有显著性 (P <0 .0 0 1) ,同时发现视网膜中HSP2 7阳性表达随着眼压升高及高眼压持续时间延长逐渐增强。结论　内源性HSP2 7在青光眼视网膜神经节细胞中表达增强可能在青光眼视神经保护中具有重要作用。     

Heat shock protein expression in the eye and in uveal melanoma

PURPOSE: Expression of heat shock proteins (HSPs) is of prognostic significance in several tumor types, whereas HSPs may also have clinical use as stimulators in tumor vaccination. HSP expression levels were determined in normal eyes and in uveal melanoma and tested whether HSPs expression was associated with prognostic parameters in the uveal melanoma. METHODS: Expression of HSP27, HSP70, HSP90, and glycoprotein96 (GP96) were determined on paraffin-embedded and frozen sections from seven healthy eyes, 20 primary uveal melanomas without prior treatment, and 18 uveal melanomas after prior treatment. HSP expression was determined by alkaline phosphatase-anti-alkaline phosphatase (APAAP) immunohistochemistry, using appropriate monoclonal antibodies and scored semiquantitatively. Expression of HSPs was validated on retinal tissue of a normal eye and in two uveal melanoma cell lines by Western blot analysis. RESULTS: Expression of HSPs was observed in epithelial and pigment cells of the normal eyes. In uveal melanoma, the level of expression of HSPs varied. Expression of HSP27 and GP96 was noted in more than 30 of 38 uveal melanomas (with, respectively, a mean of 66% and 53% positive cells). HSP70 and HSP90 were expressed in 6% of tumor cells. The amount of expression of any of the HSP types was not significantly associated with known prognostic factors. There was not a significant difference in expression of the HSPs between uveal melanomas with or without any type of prior treatment. CONCLUSIONS: In this study, expression of HSPs in uveal melanoma is not correlated with known histopathologic prognostic factors. The high expression of GP96 indicates that this protein is a potential vector in tumor vaccination in patients with large uveal melanomas.     

大鼠视网膜缺血再灌注损伤后碱性成纤维细胞生长因子对热休克蛋白70表达的影响

目的 观察大鼠视网膜缺血再灌注损伤后热休克蛋白70(HsP70)的表达以及外源性碱性成纤维细胞生长因子(bFGF)对其表达的影响。 方法 采用升高眼内压的方法，制作实验性视网膜缺血再灌注损伤大鼠模型。将24只wistar大鼠随机分为正常组(3只)和手术组(21只)。其中手术组大鼠左眼为缺血再灌注组，右眼为bFGF治疗组(玻璃体腔内注射bFGF 2肛g)，手术组根据再灌注后不同时间分为。、4、8、12、24、48、72 h组。应用免疫组织化学方法观察大鼠视网膜缺血再灌注损伤后视网膜内层组织中HsP70的表达及玻璃体腔内注射外源性bFGF对其表达的影响。 结果 对照组无阳性细胞表达。缺血再灌注组中，缺血再灌注O h后即可见HsP70的表达[(20．8±4．5)个／mm。]，并随时间延长而逐渐增加，至24 h达到高峰[(111．2±4．4)个／mm z]，随后阳性细胞递减，72 h时偶见阳性细胞。bFGF治疗组HsP70的表达变化规律与缺血组基本一致，但在8、12、24、48、72 h时均较缺血再灌注组显著增高(P<0.05).相似文献   

热休克蛋白70在视网膜神经元和Müller细胞 中的诱导及其保护作用 

目的评价大鼠视网膜神经元（RNs）和Müller细胞中热休克蛋白（HSP）70的诱导表达，及其对低糖和谷氨酸损伤的视网膜神经元的保护作用。方法大鼠RNs和Müller细胞体外培养体系分别经过热休克处理（42℃下1 h）及免疫细胞化学法检测HSP70表达的时间经过；并对RNs进行低糖(0.56 mmol/L葡萄糖，作用6 h)和谷氨酸(100 μmol/L,作用 6 h)兴奋毒性损伤，四唑盐（MTT）比色法评价细胞存活能力，同时用HSP70抗体阻断其表达。结果热休克后大鼠RNs和Müller细胞中HSP70高效表达；经热休克预处理，RNs在低糖和谷氨酸盐损伤后的细胞活力明显提高，该现象可被HSP70抗体阻断。结论热休克能够诱导RNs和Müller细胞高效表达HSP70，从而增强RNs对低糖和谷氨酸兴奋毒性损伤的耐受能力。(中华眼底病杂志,2005,21:110-113)     

热休克蛋白70在视网膜神经元和Müller细胞中的诱导及其保护作用

目的评价大鼠视网膜神经元(RNs)和Müller细胞中热休克蛋白(HSP)70的诱导表达,及其对低糖和谷氨酸损伤的视网膜神经元的保护作用. 方法大鼠RNs和Müller细胞体外培养体系分别经过热休克处理(42℃下1 h)及免疫细胞化学法检测HSP70表达的时间经过;并对RNs进行低糖(0.56 mmol/L葡萄糖,作用6 h)和谷氨酸(100 μmol/L,作用 6 h)兴奋毒性损伤,四唑盐(MTT)比色法评价细胞存活能力,同时用HSP70抗体阻断其表达. 结果热休克后大鼠RNs和Müller细胞中HSP70高效表达;经热休克预处理,RNs在低糖和谷氨酸盐损伤后的细胞活力明显提高,该现象可被HSP70抗体阻断. 结论热休克能够诱导RNs和Müller细胞高效表达HSP70,从而增强RNs对低糖和谷氨酸兴奋毒性损伤的耐受能力.