Heat Shock Protein Antibodies Research Articles

Sensitive detection of HSP70 using a current-amplified biosensor based on antibody-loaded PS-AuNPs@Cys/Au modified ITO chip.

A biosensing electrochemical platform for heat shock protein 70 (HSP70) has been developed by integrating a three-electrode indium tin oxide (ITO) on a chip. The platform includes modifications to the reference electrode and working electrode for the detection of HSP70. The new platform is constructed by assembly of HSP70 antibody on PS-AuNPs@Cys/Au indium tin oxide (ITO) electrode to create a high HSP70 sensitive surface. The PS-AuNPs@Cys/Au indium tin oxide (ITO) electrode is obtained by immersing the ITO electrode into the PS-AuNPs@Cys solution and performing constant potential deposition at -1.4V (Ag/AgCl). The PS-AuNPs@Cys/Au film deposited on ITO glass provides a desirable substrate for the immobilization of the HSP70 antibody and improves the loading of antibody between PS-AuNPs@Cys/Au and the electrode resulting in a significant amplification. Under optimal conditions, the fabricated sensor demonstrates a linear range extending from 0.1 ng mL- 1 to 1000 ng mL- 1, with an impressive detection limit of 25.7 pg mL- 1 (S/N = 3). The developed immunoassay method successfully detected the HSP70 content in normal human blood samples and outperformed the ELISA method commonly used for clinical sample analysis.

Hubungan antara Ekspresi Heat Shock Protein-70 (HSP70) dengan Skor Risk of Ovarian Cancer Relapse (ROVAR) pada Pasien Kanker Ovarium Epitelial yang Dilakukan Operasi dan Dilanjutkan dengan Kemoterapi

Background: Ovarian cancer is one of the leading causes of death in gynecological malignancies which occupies the third place in the world. One of largest challenge in epithelial ovarian cancer therapy is resistance to chemotherapy. Increased of Heat Shock Protein-70 (HSP70) expression associated with the progressivity of tumor cells growth through the resistance process against chemotherapy, whereas Risk Of Ovarian Cancer Relapse (ROVAR) score is a tool that used to predict the ovarian cancer recurrence risk in patient who underwent surgery followed by chemotherapy.Objective: This study is aimed to evaluate the correlation between HSP70 expression and ROVAR score in patient with epithelial ovarian cancer who underwent first line therapy. Method: This is a retrospective cohort study that involve 42 subjects with epithelial ovarian cancer who had been through surgery (optimal or suboptimal surgery) followed by chemotherapy. Immunohistochemical staining of pre chemotherapy paraffin blocks was performed using HSP70 antibody and recurrence risk was calculated using ROVAR scores after surgery followed by chemotherapy. The external variables evaluated were age, parity, Body Mass Index (BMI), menopause history, and surgical type. Data analysis used Chi Square test, spearman test, and logistic regression analysis.Result and Discussion: Of the total 42 subjects who met the inclusion criteria, most found in advanced stage with the age mean is 54,3 years old. In the ROVAR score measurement, in high risk category is found about 32(76.19%), while in the medium risk and low risk category is 10 (23.81%). On the other hand, strong HSP70 expression was found in 36(85,71%), moderate HSP70 expression in 6(14,39%), and weak HSP70 expression was not found. Based on Spearman's correlative test, there was a positive, very strong, and significant correlation between the expression of HSP70 and risk allocation according to ROVAR score (r=0.717, p=0.000). As well as in multivariate analysis, HSP70 expression had a statistically and clinically significant relationship with ROVAR score (p=0.017; RR 2.710; 95% CI 0.366-5.054) which strong HSP70 expression was 2.71 times more likely to have a high risk of recurrence according to ROVAR score.Conclusion: There is a positive, very strong, and significantly meaning between HSP70 expression and ROVAR score. Increased in HSP70 expression may lead to increased recurrence risk according to ROVAR score. Both could be used to predict recurrence in epithelial ovarian cancer post first line therapy.Keywords: Epithelial ovarian cancer, HSP70, ROVAR score, recurrence.

Development of a sensitive electrochemical immunosensor using polyaniline functionalized graphene quantum dots for detecting a depression marker.

As a new type of conductive material, polyaniline functionalized graphene quantum dots (PAGD), which were prepared by in-situ polymerization had been used to construct a novel electrochemical immunosensor for early screening of depression markers-heat shock protein 70 (HSP70). Profiting from the huge specific surface area, good bioactivity and excellent structure of PAGD, a variety of heat shock protein 70 (HSP70) was firmly loaded on the surface of PAGD for successful construction of basic electrode (HSP70/PAGD/GCE), which was characterized by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS), respectively. Due to the HSP70 fixed on the surface of basic electrode and the HSP70 in the samples can competitively combine with the horseradish peroxidase labeled human HSP70 antibody (HRP-Strept-Biotin-Ab). As a result, there is negative correlation between the concentration of HSP70 in samples and the detection signal of the proposed electrochemical immunosensor (HRP-Strept-Biotin-Ab-HSP70/PAGD/GCE) in the test liquid. Under conditions optimized for determining HSP70, wide linearity was obtained in the range of 0.0976-100ng/mL, with a low detection limit of 0.05ng/mL at 3σ. Moreover, the proposed electrochemical immunosensors was successfully applied to detect HSP70 in plasma samples, and exhibited good precision, acceptable stability and reproducibility. Therefore, this study provides a novel and convenient method for early clinical screening of depression markers, and also provides a reliable and objective analysis method for the diagnosis of depression at the molecular level.

Heat Shock Protein 70 (HSP70) Reduces Hepatic Inflammatory and Oxidative Damage in a Rat Model of Liver Ischemia/Reperfusion Injury with Hyperbaric Oxygen Preconditioning

BackgroundSeveral clinical conditions can cause hepatic ischemia/reperfusion (I/R) injury. This study aimed to determine the mechanism of the protective effect of hyperbaric oxygen preconditioning (HBO2P) on hepatic ischemia/reperfusion (I/R) injury in a rat model, and to investigate the effects on HBO2P and I/R injury of blocking HSP70 using antibody (Ab) pretreatment.Material/MethodsMale Sprague-Dawley rats underwent HBO2P for 60 min at 2.0 atmosphere absolute (ATA) pressure for five consecutive days before surgical hepatic I/R injury, performed by clamping the portal vein and hepatic lobe. Four groups studied included: the non-HBO2P+ non-I/R group, which underwent sham surgery (N=10); the non-HBO2P + I/R group (N=10); the HBO2P + I/R group (N=10); and the HBO2P + HSP70-Ab + I/R group (N=10) received one dose of HSP70 antibody one day before hepatic I/R injury. Serum lactate dehydrogenase (LDH), aspartate aminotransferase (AST), alanine aminotransferase (ALT), and pro-inflammatory cytokines, tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), and hepatic malondialdehyde (MDA) and myeloperoxidase (MPO) were measured biochemically. Rat liver tissues were examined histologically.ResultsIn rats with hepatic I/R injury without HSP70 antibody pre-treatment, HBO2P significantly reduced hepatic injury and levels of LDH, AST, ALT, TNF-α, IL-6, MDA, and MPO levels; in comparison, the group pre-treated with an antibody to inhibit HSP70 (the HBO2P + HSP70-Ab + I/R group) showed significant reversal of the beneficial effects of HBO2P on hepatic I/R injury (p<0.05).ConclusionsIn a rat model of hepatic I/R injury with HBO2P, HSP70 reduced hepatic inflammatory and oxidative damage.

CD20+ Tumor Infiltrating B Lymphocyte in Oral Squamous Cell Carcinoma: Correlation with Clinicopathologic Characteristics and Heat Shock Protein 70 Expression.

Objective B lymphocyte infiltration in the tumor microenvironment has been proposed to play pivotal roles in tumor progression. Heat shock protein 70 (HSP70) expressed by tumor cells can induce antitumor immune response. Few studies have examined the clinicopathologic relationship between tumor infiltrating B lymphocyte and HSP70 expression in human cancer. So far, there is no complete knowledge on the relationship in oral squamous cell carcinoma (OSCC). The present study was conducted to evaluate the relationship between tumor infiltrating B lymphocyte and HSP70 expression in OSCC, as well as the clinical outcome. Materials and Methods In this retrospective study, the immunohistochemical analysis of 50 OSCC specimens was performed using CD20 and HSP70 antibodies. The relationship between markers' expression and clinicopathologic data was evaluated using Mann–Whitney test, Chi-square test, logistic regression model, and Spearman's correlation coefficient. Results The data analysis showed significant correlation between peritumoral CD20+ B lymphocyte infiltration and lymph node metastasis (P = 0.047). Furthermore, HSP70 expression was significantly correlated with stage (P = 0.003), lymph node metastasis (P < 0.001), and tumor size (P = 0.044). However, no relationship was observed between B lymphocyte infiltration and HSP70 expression. Conclusion The results suggest that peritumoral B lymphocyte infiltration and HSP70 expression level have significant association with OSCC and may be considered as prognostic indicators in OSCC. Thus, evaluation of B cells as therapeutic targets in OSCC patients is recommended.

Switching assay as a novel approach for specific antigen- antibody interaction analysis using magnetic nanoparticles

Switching assay was applied for the detection of antigen-antibody interaction between 70-kDa heat shock protein (Hsp70) and anti-Hsp70 monoclonal antibodies in water solutions using conjugates with magnetic iron oxide nanoparticles (MNPs). Hsp70 is a ubiquitous intracellular protein that plays a crucial role in cancerogenesis and many other pathologies. Detection of the Hsp70 level in the biological fluids might have a prognostic and diagnostic value in clinic. The developed switch assay for the detection of Hsp70 demonstrated high sensitivity for antigen-antibody interaction analysis thus proving its potential for further preclinical and clinical studies.

Role and mechanism of heat shock protein 70 in airway hypersecretion

To explore the role and mechanism of heat shock protein 70 (HSP70) in airway hypersecretion. After a stimulation of 8% cigarette smoke extract (CSE), airway cells A549 were treated with HSP70 antibody and c-Jun N-terminal kinase (JNK) specific inhibitor SP600125 respectively. And the cells were divided into 4 groups of blank contrast (serum-free medium), CSE stimulation (8% CSE for 24 h), HSP70 antibody (30-min pre-treatment of HSP70 antibody and culturing in 8% CSE for 24 h) and SP600125 (30-min pre-treatment of SP600125 30 µmol/L and culturing in 8% CSE for 24 h). The relative expression levels of MUC5AC protein in various groups were determined by enzyme-linked immunosorbent assay (ELISA). And the relative transcription level of MUC5AC mRNA was detected by reverse transcription-polymerase chain reaction (PCR) while the synthesized levels of HS70 as well as the phosphorylation levels of JNK and activated protein-1 (mostly c-Jun) were measured by Western blot. As compared with those in blank contrast group (0.26 ± 0.10, 0.28 ± 0.06, 0.30 ± 0.05, 0.30 ± 0.08, 0.36 ± 0.08), HSP70 antibody group (0.30 ± 0.12, 0.29 ± 0.09, 0.34 ± 0.06, 0.47 ± 0.19, 0.39 ± 0.13) and SP600125 group (0.38 ± 0.06, 0.31 ± 0.14, 0.39 ± 0.04, 0.44 ± 0.12, 0.48 ± 0.11), the relative expression levels of MUC5AC protein and mRNA, phosphorylation JNK (p-JNK), c-Jun and p-c-Jun (0.52 ± 0.07, 0.64 ± 0.11, 0.73 ± 0.06, 0.67 ± 0.10, 0.67 ± 0.09) significantly increased in CSE stimulation group (all P < 0.05). And the synthesis levels of HSP70 (0.75 ± 0.09) in CSE stimulation group increased than blank contrast group (0.29 ± 0.03) and HSP70 antibody group (0.40 ± 0.11) (all P < 0.05). HSP70 may enhance the expression of MUC5AC in bronchial epithelial A549 cells via a signaling pathway of JNK/AP-1..

167 THE EFFECT OF ELEVATED TEMPERATURE ON THE HEAT SHOCK PROTEIN 70 (HSP70) EXPRESSION IN BOVINE CUMULUS&amp;ndash;OOCYTE COMPLEXES AFTER IN VITRO MATURATION

Elevated temperature during in vitro maturation negatively affects the oocyte's developmental competence, leading to disturbances in nuclear and cytoplasmic maturation. In previous studies, the role of cumulus granulosa cells (CGC) in cumulus-oocyte complexes' (COC) response to elevated temperature was underestimated. However, CGC play an essential role in folliculogenesis, supporting the oocyte's metabolism as well as meiotic progression. Thus, CGC may be engaged in COC response to heat shock. Heat shock protein 70 (HSP70) is the major protein engaged in cellular response to heat stress. The aim of this study was to determine the HSP70 expression in both CGC and oocytes in response to elevated temperature after COC in vitro maturation. COC were collected from bovine ovarian follicles (diameter 2–6 mm) from slaughtered cows and divided into two treatment groups: I (control) – COC were in vitro matured in control temperature (38.5°C); II (experimental) – COC were in vitro matured in elevated temperature (41°C). After in vitro maturation they were mechanically separated into CGC and oocyte. In vitro maturation was conducted in TCM199 25 mM HEPES medium supplemented with 10% FBS, 0.02 IU mL–1 NIH-pFSH, 1 μg mL–1 17β-oestradiol, 22 μg mL–1 Na-pyruvate and 10 μg mL–1 gentamicin, adjusted to pH 7.4 in 5% CO2. The HSP70 expression in CGC and oocytes was performed by real-time PCR and normalized to S18/H2A and S18 gene expression, respectively. In addition, the immunocytofluorescent analysis of HSP70 expression in CGC and oocyte's were performed. The HSP70 was stained using primary monoclonal mouse antibodies raised against bovine HSP70 and secondary antibody raised against mouse conjugated with Alexa 488. Nuclei were stained with Hoechst 33342. Slides were analysed under laser confocal microscope (FV-500, Olympus, Center Valley, PA, USA). The HSP70 expression in CGC was measured as total optical density of HSP70 and normalized to total optical density of nuclei. Statistical analyses were performed by Portable Statgraphics 5.0 Centurion. Mean values of HSP70 expression in CGC and oocytes in real-time PCR and immunofluorescent analysis in CGC were compared using Tukey's HSD test (α = 0.01). After in vitro maturation, the expression of HSP70 in CGC was higher (0.13 ± 0.052 a.u., n = 35) in experimental temperature compared to the control (0.058 ± 0.008 a.u., n = 35) (P &lt; 0.01). In oocytes, HSP70 expression in experimental temperature (32.5 ± 5.2 a.u., n = 12) was similar to control (29.8 ± 5.6 a.u., n = 12). The immunofluorescent analysis confirmed data from real-time PCR analysis, indicating that the HSP70 expression in CGCs in experimental temperature was higher (0.46 ± 0.07 a.u., n = 10) compared to the control (0.12 ± 0.02 a.u., n = 10; P &lt; 0.01). After COC in vitro maturation in stress conditions, the HSP70 expression was up-regulated in CGC, but not in oocytes. That may indicate that CGCs play the major role in COC response to elevated temperature; however, the analysis of other heat shock proteins expression in CGC and oocytes need to be conducted. Research was supported by 505-10-023300-K00169-99.

High-altitude pulmonary edema can be prevented by heat shock protein 70-mediated hyperbaric oxygen preconditioning.

The primary goal of this study was to test whether high-altitude exposure (HAE of 9.7% O2 at 0.47 absolute atmosphere [ATA] for 3 days) was capable of increasing lung edema, neutrophil, and hemorrhage scores as well as decreasing lung levels of both aquaporin 1 (AQP1) and AQP5 proteins and messenger RNA (mRNA) expression in rats, with a secondary goal to test whether a preinduction of heat shock protein 70 (HSP70) by hyperbaric oxygen preconditioning (HBO2P of 100% O2 at 2.0 ATA for 1 hour per day for 5 consecutive days) attenuated the HAE-induced increased lung injury scores and decreased lung AQP1 and AQP5 protein and mRNA expressions. Rats were assigned to (1) non-HBO2P (21% O2 at 1.0 ATA) + non-HAE (21% O2 at 1.0 ATA) group; (2) non-HBO2P + HAE group; (3) HBO2P + HAE group; and HBO2P + HSP70 antibodies (Ab) + HAE group. For the HSP70 Ab group, a neutralizing HSP70 Ab was injected intravenously at 24 hours before HAE. All the physiologic and biochemical parameters were obtained at the end of HAE or the equivalent period of non-HAE. The cardiovascular and blood gas parameters were monitored for all experiments. Bronchoalveolar lavage (BAL) was performed to determine proinflammatory cytokines (interleukin 6, interleukin 1β, and tumor necrosis factor α). Parts of the lung were excised for myeloperoxidase activity measurement, whereas the rest was collected for lung damage score assessments. AQP1 and AQP5 protein and mRAN expressions were also determined in the lung tissues. In the non-HBO2P + HAE group, the animals displayed higher values of lung myeloperoxidase activity, BAL proinflammatory cytokines, lung water weight, and acute lung injury scores compared with those of the non-HBO2P + non-HAE controls. In contrast, the non-HBO2P + HAE group rats had lower values of lung AQP1 and AQP5 protein and mRNA expressions, mean arterial pressure, heart rate, SO2, Paco2, HCO3, and pH compared with those of non-HBO2P + non-HAE group rats. The increased acute lung edema, neutrophil, and hemorrhage scores; increased BAL levels of proinflammatory cytokines; decreased lung AQP1 and AQP5 protein and mRNA expressions; and hypotension, bradycardia, hypoxia, and acidosis caused by HAE were all significantly attenuated by HBO2P. Our data indicate that HBO2P may attenuate high-altitude acute lung injury by a preinduction of lung HSP70 in rats.

The effects of wet cupping on serum high-sensitivity C-reactive protein and heat shock protein 27 antibody titers in patients with metabolic syndrome

It has previously been reported that increased level of serum heat shock proteins (Hsps) antibody in patients with metabolic syndrome. It is possible that the expression of Hsp and inflammatory markers can be affected by cupping and traditional Chinese medicine. There is a little data investigating the effects of cupping on markers of inflammation and Hsp proteins, hence, the objective of this study was evaluation of the effects of wet cupping on serum high-sensitivity C-reactive protein (hs-CRP) and Hsp27 antibody titers in patients with metabolic syndrome. Serum Hs-CRP and Hsp27 antibody titers were assessed in samples from 126 patients with metabolic syndrome (18-65 years of age) at baseline, and after 6 and 12 weeks after treatment. One hundred and twenty-six patients were randomly divided into the experimental group treated with wet cupping combined with dietary advice, and the control group treated with dietary advice alone using a random number table. Eight patients in case group and five subjects in control groups were excluded from the study. Data were analyzed using SPSS 15.0 software and a repeated measure ANCOVA. Serum hs-CRP titers did not change significantly between groups (p>0.05) and times (p=0.27). The same result was found for Hsp27 titers (p>0.05). Wet-cupping on the interscapular region has no effect on serum hs-CRP and Hsp27 patients with metabolic syndrome.

The Effects of Wet Cupping on Serum High-Sensitivity C- Reactive Protein and Heat Shock Protein 27 Antibody Titers in Patients with Metabolic Syndrome

Summary Introduction: It has previously been reported that increased level of serum heat shock proteins (Hsps) antibody in patients with metabolic syndrome. It is possible that the expression of Hsp and inflammatory markers can be affected by cupping and traditional Chinese medicine. There is a little data investigating the effects of cupping on markers of inflammation and Hsp proteins,

Attenuating heatstroke-induced acute lung inflammation, edema, and injury in rats by exercise preconditioning

This study aimed to ascertain whether heat-induced acute lung edema, inflammation, and ischemic damage can be affected by heat shock protein 70 (HSP-70)-mediated exercise preconditioning (EP) in rats. Wistar rats were assigned to one of the following four groups: the non-EP + nonheated group, the non-EP + heated group, the EP + heated group, and the EP + HSP-70 antibodies + heated group. EP groups of animals were subjected to a protocol of running on a treadmill for 30 minutes at 20 m/min, 30 minutes at 30 m/min, and 60 minutes at 30 m/min after 1, 2, and 3 weeks of training, respectively. Heated group of animals, under general anesthesia, were put in a folded heating pad of 43°C for 68 minutes. Then, the heated animals were allowed to recover at room temperature. HSP-70 antibodies were injected intravenously 24 hours before heat exposure. As compared with the non heated + non-EP rats, the heated + non-EP rats had significantly higher scores of alveolar edema, neutrophil infiltration, and hemorrhage, acute pleurisy, and increased bronchoalveolar fluid levels of proinflammatory cytokines and ischemic and oxidative damage markers. EP, in addition to inducing overexpression of HSP-70 in lung tissues, significantly attenuated heat-induced acute pulmonary edema, inflammation, and ischemic and oxidative damage in the lungs. HSP-70 antibodies, in addition to reducing HSP-70 expression in the lungs, significantly attenuated the beneficial effects of EP in reducing acute lung inflammation and injury. EP may attenuate the occurrence of pulmonary edema, inflammation, as well as ischemic and oxidative damage caused by heatstroke by up-regulating HSP-70 in the lungs.

Response of circulating heat shock protein 70 and anti-heat shock protein 70 antibodies to catheter ablation of atrial fibrillation

BackgroundThis pilot study investigated the association between heat shock protein 70 (HSP70) and anti-HSP70 antibodies as well as their changes and rhythm outcome after atrial fibrillation (AF) catheter ablation.MethodsWe studied 67 patients with AF (59±11 years, 66% male, 66% lone AF) undergoing catheter ablation. Circulating HSP70 and anti-HSP70 antibody levels were quantified using commercially available assays before and 6 months after catheter ablation. Serial 7-day Holter ECGs were used to detect AF recurrences.ResultsAt baseline, HSP70 was detectable in 14 patients (21%), but there was no correlation between clinical or echocardiographic variables and the presence or the level of HSP70. In contrast, patients with paroxysmal AF (n=39) showed lower anti-HSP70 antibodies (median [IQR] of 43 [28 – 62] μg/ml) than patients with persistent AF (n=28; 53 [41 – 85] μg/ml, p=0.035). Using multivariable regression analysis, AF type was the only variable associated with anti-HSP70 antibodies (Beta=0.342, p=0.008). At 6 months, HSP70 was present in 27 patients (41%, p<0.001 vs. baseline). Similarly, there was an increase of anti-HSP70 antibodies (48 [36 – 72] vs. 57 [43 – 87] μg/ml, p<0.001). AF recurrence rates were higher in patients with HSP70 increase ≥0.025 ng/ml (32 vs. 11%, p=0.038) or anti-HSP70 increase ≥2.5 μg/ml (26 vs. 4%, p=0.033).ConclusionsHSP70 and anti-HSP70 antibodies may – at least in part – be associated in the progression of AF and AF recurrence after catheter ablation.

Suicidal ligature strangulation without an auxiliary mechanism. Reports of two cases with a cotton rope or a T-shirt and staining results of the brains using anti HSP-70, CIRBP, RBM3, HIF1-alpha, SIRT 1 and p53 antibodies

It is still often described in forensic textbooks that suicidal strangulation by ligature without auxiliary device to remain tightening the ligature after unconsciousness of the victim. We report two cases of suicidal ligature strangulation without auxiliary mechanism. One occurred in a closed room and other was carried out in a police cell. These two cases reveal that the suicidal ligature strangulation is possible without auxiliary mechanism, however it was difficult to distinguish from homicide only from forensic autopsy findings, although death scenes and existence of suicidal notes produced clear verdicts of suicide in two cases. The death mechanism in these cases was discussed. The nucleus of the neuronal cells showed good reactivity with hypoxia or ischemia related antibodies, such as cold inducible RNA-binding protein (CIRBP), RNA-binding motif protein 3(RBM3), heat shock protein 70(HSP-70) and hypoxia-inducible factor 1(HIF-1), however the cells showed no reactivity with anti Sirtuin 1(SIRT 1) and p53 antibodies which are related energy metabolism or apoptosis. This is the first report in which the expression of hypoxia and /or ischemic related antigens described above was confirmed even in post-mortem human brains.

Hypobaric Hypoxia Preconditioning Attenuates Experimental Heatstroke Syndromes via Preinduction of Heat Shock Protein 70

IntroductionHeatstroke has been defined as a form of hyperthermia associated with a systemic inflammatory response that leads to multiple organ dysfunction syndrome (MODS). It has also been documented that heat shock protein 70 (HSP70) preconditioning is able to induce thermotolerance. Here, the authors further investigated whether hypobaric hypoxia preconditioning (HHP) improved the MODS in heatstroke by up-regulation of HSP70. MethodsAnesthetized rats were randomly assigned to (a) non-HHP + nonheated group, (b) non-HHP + heated group, (c) HHP + heated group and (d) HHP + HSP70 antibodies (Abs) + heated groups. All heated groups were exposed to heat stress (43°C, 70 minutes) to induce heatstroke. For HHP, animals were exposed to 0.66 atmosphere absolute (18.3% O2) for 5hours daily for consecutive 5days per week for 2weeks before the start of heat exposure. ResultsHHP significantly (i) attenuated hypotension, (ii) reduced plasma index of the toxic oxidizing radicals and the organ injury indicator, (iii) attenuated plasma systemic inflammatory response molecules, (iv) reduced an index of infiltration of polymorphonuclear neutrophils in the lung like myeloper-oxidase activity, (v) promoted plasma levels of an anti-inflammatory cytokine, interleukin-10, (vi) promoted the survival time to fourfold compared with non-HHP group and (vii) promoted the overexpression of HSP70 in different organs (eg, the lung) during heatstroke. The beneficial effects of HHP could be significantly attenuated by HSP70 Ab preconditioning. ConclusionOur results show that HHP protects rats from heat-induced MODS via up-regulating HSP70. Thus, HHP could be a novel strategy for the prevention of heatstroke animals or patients before heat exposure.

Effects of Heavy Metals on Ultrastructure and HSP70S Induction in the Aquatic Moss Leptodictyum Riparium Hedw

The effects of heavy metals, both toxic (Pb, Cd) and essential (Cu, Zn) on the ultrastructure and the induction of Heat Shock Protein 70 (HSP70) have been studied in the aquatic moss Leptodictyum riparium Hedw. In vitro cultured L. riparium was treated with different heavy metals, both toxic, as cadmium or lead; and essential microelements such as Copper or Zinc concentrations ranging from 10−3 to 10−6 M to investigate both ultrastructural damage and HSP induction. TEM observations showed that sub-lethal concentrations of heavy metals caused only slight changes, largely localized in the chloroplasts. Among all the heavy metals tested, cadmium caused the most severe modifications. Heavy metals caused the decrease of the soluble protein content and the enhancement of proteins reacting versus HSP70 antibodies, suggesting that molecular chaperons might be involved in the resistance to toxic effects of lead, cadmium, copper and zinc. Therefore, the induction of HSP70 in L. riparium would confer a higher resistance to pollutants under stressful conditions lethal for other mosses and higher plant species. These results suggest that the moss L. riparium can tolerate heavy metals stress without incurring severe cellular/subcellular damage. Therefore it can be used as a useful indicator of heavy metals accumulation.

Abstract P172: Circulating Heat Shock Protein 70 and Antiheat Shock Protein 70 Antibodies in Atrial Fibrillation: Relation with Atrial Fibrillation Type and Response to Catheter Ablation

Background: HSP and anti-HSP antibodies have been associated with AF development and progression. This study investigated the possible association between circulating heat shock protein 70 (HSP70) and anti-HSP70 antibodies as well their changes and rhythm outcome after atrial fibrillation (AF) catheter ablation. Methods: In 67 patients with AF (59±11 years, 66 % male, 66 % lone AF) undergoing catheter ablation, circulating HSP70 and anti-HSP70 antibodies levels were quantified before and 6 months after catheter ablation. Serial 7-day Holter ECGs were used to detect AF recurrences. Results: At baseline, HSP70 was detectable in 14 patients (21 %), but there was no correlation between clinical or echocardiographic variables and the presence or the level of HSP70. Patients with paroxysmal AF (n=39) showed lower anti-HSP70 antibodies (median 43, IQR 28 - 62 µg/ml) than patients with persistent AF (n=28; 53, 41 - 85 µg/ml, p=.035). Using multivariable regression analysis, AF type was the only variable associated with anti-HSP70 antibodies (Beta=.342, p=.008). At 6 months, HSP70 was present in 27 patients (41 %, p&lt;.001 vs. baseline) with an overall increase (median 0, IQR 0 - 0 vs. 0, 0 - 0.09 ng/ml, p=.029). Similarly, there was an increase of anti-HSP70 antibodies (48, 36 - 72 vs. 57, 43 - 87 µg/ml, p&lt;.001). AF recurrence rates were higher in patients with HSP70 increase &gt;0.025 ng/ml (32 vs. 11 %, p=.038) or anti-HSP70 antibodies increase &gt;2.5 µg/ml (26 vs. 4 %, p=.033). Conclusion: HSP70 and anti-HSP70 antibodies may be involved in the progression of AF and AF recurrence after catheter ablation.

HSP70 Antibodies in 80 Patients with “Clinically Certain” Meniere's Disease

We tested the claim that a significant proportion of patients with Meniere's disease have antibodies to heat shock protein 70 (HSP70) antigen, which may lead to defective endolymphatic sac function and vertigo attacks. Serum samples were taken from 80 subjects with a "certain" diagnosis of Meniere's disease (American Academy criteria plus electrocochleographic confirmation of endolymphatic hydrops with tone burst stimuli) and were tested for HSP70 antibodies with the OTOblot (hsp70) Western blot assay. The response was recorded as negative, positive, or equivocal. Samples from 80 sex- and age-matched blood donors were used as controls. Of 80 patients with "clinically certain" Meniere's disease, 14 were positive for HSP70 antibodies or equivocal; of 80 controls, 10 were positive or equivocal. There was no significant difference (p=0.239). There was no correlation with bilateral disease, "activity" of Meniere's disease, or stage of Meniere's disease. Patients with an unequivocal diagnosis of Meniere's disease do not have a significantly raised incidence of HSP70 antibodies.

Anti-60-kDa Heat Shock Protein Antibodies in Fetal Serum: A Biomarker for Unexplained Small for Gestational Age Fetuses

Introduction: Small for gestational age (SGA) is an important problem affecting 10% of pregnancies and is associated with significant perinatal morbidity. In about 80% of cases, a probable etiology or a major risk factor can be identified. But almost 20% of SGA cases are considered unexplained. The 60-kDa heat shock protein (HSP60) is a highly immunogenic protein whose synthesis is greatly upregulated under nonphysiological conditions. Bacterial and human HSP60 share a high degree of sequence homology, and immunity to conserved epitopes may result in development of autoimmunity following a bacterial infection. We hypothesized that unexplained SGA could be the consequence of immune sensitization to human HSP60. Methods: Unexplained SGA fetuses were identified by ultrasound biometry with normal Doppler velocimetry and with no detectable maternal or fetal abnormalities. Fetal sera were obtained by cordocentesis performed for a karyotype analysis in cases of unexplained SGA (study group) or for screening of Rhesus incompatibility (control group). Fetal sera were tested for HSP60 antigen and for IgG and IgM anti-HSP60 by ELISA as well as for other immune and hematological parameters. Results: Maternal parameters were similar between the 12 study cases and the 23 control cases. The mean gestational age at cordocentesis was 29 weeks. IgM anti-HSP60 was detected in 12 cases (100%) and in no controls (p < 0.00017), while IgG anti-HSP60 was detected in 7 cases (58%) and only 1 control (p < 0.001). Three of the 4 cases with the highest IgM antibody levels died. There were no differences in fetal serum levels of HSP60 antigen or other immune and hematological markers between the two groups. Conclusion: Fetuses with unexplained SGA are positive for IgM and IgG antibody to human HSP60 and the specific IgM antibody level is predictive of fetal mortality. Detection of these antibodies indicates that a placental perturbation and a fetal autoimmune reaction to HSP60 are associated with this developmental delay.

Simvastatin treatment reduces heat shock protein 60, 65, and 70 antibody titers in dyslipidemic patients: A randomized, double-blind, placebo-controlled, cross-over trial

Objective This study aimed to evaluate the effects of statin therapy on serum levels of antibodies to several specific heat shock proteins (HSPs) in dyslipidemic patients. Design and methods Participants ( n = 102) were treated with simvastatin (40 mg/day), or placebo in a randomized, double-blind, placebo-controlled, cross-over trial. Anti-HSP60, 65, 70, and hs-CRP levels were measured before and after each treatment period. Seventy-seven subjects completed the study. Results Treatment with simvastatin was associated with significant reductions in serum anti-HSP60, 65, and 70 titers in the dyslipidemic patients (10%, 14%, and 15% decrease, respectively) ( p < 0.001). There have been previous reports of reductions in serum CRP with statin treatment, and although median CRP levels were 9% lower on simvastatin treatment, this did not achieve statistical significance. Conclusion While it is unclear whether HSP antibodies are directly involved in atherogenesis, our findings suggest that simvastatin inhibits autoimmune responses that may contribute to the development of cardiovascular disease.