anti-HSP70 Antibody Research Articles

THE ROLE OF THE HEAT SHOCK PROTEIN 70 IN THE PATHOGENESIS OF CARDIOVASCULAR PATHOLOGY

The problem of studying cardiovascular diseases (CVD) for a long time remains extremely important, and, therefore, there are many works that offer new ways to diagnose and treat this group of diseases. Great opportunities are provided by the study of molecular interactions for a more accurate understanding of the pathogenesis of cardiovascular pathology. Many studies have recently been devoted to finding potential markers of CVD risk with the aim of more accurate and early diagnosis. In this review we analyze the latest literature data dedicated to the role of heat shock protein 70 (HSP70) in cardiovascular pathology. HSP70 take part in such processes as arterial hypertension, coronary heart disease, and atherosclerosis. In atherogenesis, serum heat shock proteins 70 play a major role. It has been proven that in patients with a high concentration of heat shock protein molecules circulating in the blood, increased values of the carotid intima-media complex were observed. The important role of antibodies to circulating HSP70 is noted. Found an association of high levels of these antibodies with atherosclerosis in patients with arterial hypertension in history, with myocardial infarction. Low levels of anti-HSP70 antibodies are observed in patients with acute coronary syndrome. This proves the complexity of the mechanism and the dual role of antibodies against serum heat shock proteins 70. Thus, antibodies against heat shock proteins 70 can be assessed as a protective marker, and as a predictor, which requires further study, and the HSP70 molecules themselves can somehow to participate in the development of cardiovascular pathologies. Much attention is paid to the role of the inflammatory process and the mechanisms of innate immunity in CVD. As it is currently believed that Danger-associated molecular patterns (DAMPs) are involved in the pathogenesis of these pathologies in the context of a “hazard/damage” model. According to this model, the triggering factor is stress, leading to the release of DAMPs and their binding to innate immunity receptors - Toll-like receptors (TLRs). Activation of TLRs triggers the signaling cascade in the cell leading to the synthesis of pro-inflammatory cytokines. This contributes to the development of inflammation, which can provoke the emergence of new pathological processes in the body and worsen the course of existing diseases. The identification of new potential markers and knowledge of the molecular mechanisms of the pathogenesis of CVD can play an important role in the development of a new individual approach to the prevention of cardiovascular diseases.

Depression in adolescent girls: Relationship to serum vitamins a and E, immune response to heat shock protein 27 and systemic inflammation

Background: The inflammation and oxidative stress are thought to play an important role in the etiopathogenesis of some psychological disorders. We aimed to assess the potential relationships between serum fat soluble vitamins (Vitamins A and E), antibody titers to Hsp27 (anti-Hsp27) and hematological markers of inflammation, with mood disorders in a population of adolescent girls. Methods: A total of 563 adolescent girls (Age 12–18 years) were included in the study. The presence and severity of depression, insomnia and sleepiness were assessed using validated questionnaires. Serum vitamins A and E, anti-Hsp27 antibody titers, white blood cell, lymphocyte, neutrophil, platelet counts, and red blood cell distribution width (RDW), were also measured. Neutrophil to lymphocyte ratio (NLR), platelet to lymphocyte ratio (PLR), and RDW to platelet ratio (RPR) were calculated. Results: Serum anti-HSP27 antibody titers, PLR, and RPR values was significantly higher in subjects with a high depression score compared to normal individuals (p < 0.05). However, there was no association between serum inflammatory markers concentrations and sleep disorders; although individuals with insomnia had a lower vitamin E/HDL ratio compared to healthy adolescents. In multivariate logistic regression analyses adjusted for potential confounders, anti-HSP was an independent predictor of severe depression (OR = 5.0, 95% CI: 1.6–15.7, p < 0.05). Limitation: The cross-sectional design of study and the inclusion of only female adolescents participants are limitations. Conclusion: Our findings suggest that serum anti-HSP27 antibody titers may be useful biological marker in depressive patients. This finding may support a role of oxidative stress in the etiology of depression, and targeting this pathway may be of value in the treatment of depression.

Is there any association between Serum anti-HSP27 antibody level and the presence of metabolic syndrome; population based case-control study

Abstract Background: Heat shock protein 27 (HSP27) is an intracellular chaperone constitutively expressed in many cell types including cardio myocytes and endothelial cells. Circulating levels of HSP27 and anti-HSP27 antibody are higher in patients with CVD. Anti-HSP27 antibody concentrations were also reported to be increased in atherogenesis. We aimed to evaluate serum anti-HSP27 antibody titers in individuals with, or without, MetS in the MASHAD study cohort with large sample size in 6,568 subjects. Methods: Participants with MetS were identified from MASHAD cohort (n=3358) using the IDF criteria, and the control group were those individuals who did not meet these criteria (n=3210). In-house enzyme-linked immune sorbent assay (ELISA) method was used for measuring Anti-HSP27 antibody levels. The two groups were matched for age, sex and smoking habit. Results: As expected, there were significant differences in height (p= 0.004), waist and hip circumference, weight, BMI, systolic and diastolic blood pressure, TGs, TC, HDL-C, Hs-CRP, glucose, with the presence of diabetes mellitus, hypertension, hyperlipidemia (p&lt;0.001) between the two groups. Serum HSP27 antibody titers did not show significant difference between the groups with and without metabolic syndrome (p= 0.740). Conclusion: In conclusion, our results revealed serum anti-HSP27 antibody titers were not statistically different between individuals with and without MetS. However, it is possible that drug treatment may affect antibody titers and confound our findings in this population sample..

(-)-Epicatechin attenuates hepatic sinusoidal obstruction syndrome by inhibiting liver oxidative and inflammatory injury

Hepatic sinusoidal obstruction syndrome (HSOS) is a rare liver disease with considerable morbidity and mortality. (-)-Epicatechin (EPI) is a natural flavonol. This study aims to investigate the protection of EPI against monocrotaline (MCT)-induced HSOS and its engaged mechanism. Results of serum alanine/aspartate aminotransferases (ALT/AST) activities, total bilirubin (TBil) and bile acids (TBA) amounts, liver histological evaluation, scanning electron microscope observation and hepatic metalloproteinase-9 (MMP-9) expression all demonstrated the protection by EPI against MCT-induced HSOS in rats. EPI attenuated liver oxidative injury induced by MCT. EPI enhanced the nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) and increased the expression of its downstream antioxidant genes in rats. Molecular docking results implied the potential interaction of EPI with the Nrf2 binding site in kelch-like ECH-associated protein-1 (Keap1). The EPI-provided protection against MCT-induced HSOS was diminished in Nrf2 knock-out mice when mice were treated with MCT for 24 h but not for 48 h. However, EPI reduced the increased liver myeloperoxidase (MPO) activity, hepatic infiltration of immune cells, pro-inflammatory cytokines expression and nuclear factor κB (NFκB) activation in both wild-type and Nrf2 knock-out mice when mice were treated with MCT for 48 h. EPI reduced the elevated serum heat shock protein 60 (HSP60) content, and reversed the decreased mitochondria expression of HSP60 and Lon in livers from MCT-treated rats. Furthermore, the MCT-induced HSOS was markedly alleviated in mice treated with anti-HSP60 antibody. Taken together, this study demonstrates that EPI attenuates MCT-induced HSOS by reducing liver oxidative injury via activating Nrf2 antioxidant pathway and inhibiting liver inflammatory injury through abrogating NFκB signaling pathway initiated by HSP60.

Evaluation of the protective effect of immunization spf DBA/2J mice with selected bacterial, recombinant Hsp60 antigens during Salmonella Enteritidis challenge

Salmonella Enteritidis is one of the most common causes of food poisoning in humans. Many attempts have been made to develop an effective vaccine against S. Enteritidis for use in poultry, but experiments aimed at the complete elimination of this pathogen from poultry farms have not provided satisfactory results. The development of new generation vaccines against salmonellosis, such as subunit vaccines based on heat shock proteins (HSPs), is strongly justified. The high immunogenicity of Hsp60 isolated from Procaryota, including Salmonella, has been suggested by the presence of IgG anti-Hsp60 antibodies in mice immunized with these proteins. The aim of the studies was to evaluate the protective effects of immunization with recombinant Hsp60 from selected gram-negative bacteria (S. Enteritidis, Escherichia coli, Pasteurella multocida, Histophilus somni) in spf DBA/2 J mice experimentally infected with S. Enteritidis. The study demonstrated that double subcutaneous immunization of mice with a dose of 10 μg rHsp60 induced a specific immune response of IgG antibodies in tested animals. The median lethal dose (LD50) for the murine model spf DBA/2 J orally infected with S. Enteritidis was estimated at 6.84 × 105 cfu/animal. Mice immunized with rHsp60 from gastrointestinal pathogens (S. Enteritidis and E. coli) showed better survival after experimental infection with a 3 × LD50 dose from S. Enteritidis, compared to animals immunized with proteins obtained from respiratory pathogens (P. multocida and H. somni). However, the log-rank analysis did not show significant differences in the survival rates between rHsp60-immunized mice and controls. S. Enteritidis was not isolated any less frequently from internal organs and faeces of rHsp60-immunized mice than from controls. Nevertheless, the level of haptoglobin (but not IL-6) was increased in all mice in which the presence of the pathogen was observed. Bacterial Hsp60 is an interesting candidate for a subunit vaccine, but its use in livestock animals must be further investigated.

Serum anti-hsp27 antibodies concentration in diabetes mellitus; population based case-control study

Background: Diabetes mellitus is an important risk factor for cardiovascular disease. Different biomarkers have been investigated for the diagnosis of diabetes pathogenesis or its complications. There are also reports regarding an increased level of anti-HSP27 antibodies in atherogenesis. We aimed to evaluate serum anti-heat shock protein 27 antibodies level in subjects with diabetes mellitus and undiagnosed individuals. Materials and Methods: This cross-sectional study was conducted on 6447 MASHAD study subjects, including four groups with diabetes mellitus (n=610), undiagnosed diabetes (n=162), impaired fasting glucose (IFG) (n=619) and normal (n=5056) subjects. Demographic and anthropometric data were obtained from all participants. Fasting serum glucose (FSG) and other parameters were measured. In-house enzyme-linked immune sorbent assay method was used for measuring Anti-HSP27 antibodies levels. Results: There were significant differences in weight (p=0.034), body mass index, waist, and hip circumference, systolic and diastolic blood pressure, fasting serum glucose, lipid profile and high sensitive- C reactive protein (p<0.001) between four groups of diabetes mellitus, undiagnosed diabetes, impaired fasting glucose, and normal subjects. The serum anti-HSP27 antibody titer did not show a significant difference between studied groups. Conclusion: Serum antibody titers to HSP27 were not significantly different between four groups categorized based on their FSG levels in a large population.

CLUES TO THE DEVELOPMENT OF THE ANTI-ATHEROSCLEROTIC HEAT SHOCK PROTEIN 27 IMMUNOTHERAPY FROM THE BIOPHYSICAL CHARACTERIZATION OF THE HSP27 IMMUNE COMPLEX

Heat Shock Protein 27 (HSP27) is a 205-amino acid protein with novel anti-atherosclerotic properties that attenuates atherosclerosis by reducing inflammation and lowering cholesterol levels in serum and arterial plaques. Diminished serum HSP27 levels are associated with increased cardiovascular risk. Recently, we demonstrated the presence of anti-HSP27 auto-antibodies (AAB), that potentiate atheroprotection by attenuating inflammatory signaling. For example, as opposed to HSP27 alone, the combination of HSP27 plus the AAB (i.e., the immune complex) appears to dock at the cell membrane, altering the interaction with extracellular inflammatory signaling pathways (e.g., TLR4 mediated activation of NF-kB). As we are currently focusing on the development of HSP27 immunotherapeutics, this study was performed to characterize the structural features of the HSP27-AAB interaction that may be of importance to peptidomimetic design. Homology modelling and secondary structure prediction tools were used to generate design assorted forms of the HSP27 in E.coli. Various biophysical techniques including circular dichroism (CD), dynamic light scattering (DLS), isothermal titration calorimetry (ITC) were used to assess structure and function of these recombinant protein in the presence and absence of an anti-HSP27 polyclonal antibody (PAb; generated in rabbits) that was used to mimic naturally occurring AABs. In vitro, this PAb showed a high degree of fidelity with AABs. The HSP27 C-terminus includes a key conserved alpha crystalline domain with a primary beta sheet structure. The N-terminus is responsible for the formation of larger oligomers. Using ITC we determined that the interaction of HSP27 with PAB was an exothermic reaction (△H = -3.0 x 10-4 cal/mol) with a Ka of 8.9 x10-4 M-1 with an average N valve of 0.5 suggesting 2:1 binding between the PAb and HSP27. Moreover, this interaction markedly increases the beta-sheet content and complex multi-site binding profile. Our study suggests that the formation of the HSP27 immune complex involves a moderately intensity interaction which results in conformational (secondary structure) changes. These structural changes may be of importance in determining the extracellular signalling (e.g., docking with TLR4), and highlight newly exposed HSP27 motifs that will be soon be explored using candidate peptidomimetics that may be of therapeutic value for the treatment of not only atherosclerosis, but inflammation in general.

Heat shock protein 70 and anti-heat shock protein 70 antibodies in patients with chronic glomerulonephritis.

We evaluated the heat shock system 70 (HSP70) in patients with chronic glomerulonephritis (CGN). Seventy-six patients with CGN patients were included in our study. Ten patients with mild proteinuria (median 0.48 [0.16-0.78] g/24h) and ten healthy subjects served as positive and negative controls, respectively. Urinary levels of HSP70, interleukin-10, and serum levels of anti-HSP70 were measured by ELISA. The immunohistochemical peroxidase method was used to study the expression of HSP70 and Foxp3+ in kidney biopsies. TregFoxP3+ cells in the interstitium were determined morphometrically. Median urinary HSP70 levels in patients with nephrotic syndrome (NS) [6.57 (4.49-8.33) pg/mg] and subnephrotic range proteinuria [5.7 (4.12-6.9) pg/mg] were higher (p < 0.05) than in positive [3.7 (2.5-4.82) pg/mg] and negative [3.78 (2.89-4.84) pg/mg] controls. HSP70 expression index in tubular cells positively correlated with urinary HSP70 (Rs = 0.948, р < 0.05) and proteinuria (Rs = 0.362, p < 0.05). The number of TregFoxp3+ cells in the kidney interstitium and interleukin-10 excretion were lower in patients with NS. Anti-HSP70 antibody serum levels in patients with NS [21.1 (17.47-29.72) pg/ml] and subnephrotic range proteinuria [24.9 (18.86-30.92) pg/ml] were significantly higher than in positive [17.8 (12.95-23.03) pg/ml] and negative [18.9 (13.5-23.9) pg/ml] controls. In patients with CGN, increasing proteinuria was associated with higher HSP70 renal tissue and urinary levels. However, activation of HSP70 in patients with nephrotic syndrome did not lead to an increase in tissue levels of TregFoxp3+ cells or to the release of IL-10.

HSP27-Mediated Extracellular and Intracellular Signaling Pathways Synergistically Confer Chemoresistance in Squamous Cell Carcinoma of Tongue.

Purpose: Squamous cell carcinoma of tongue (SCCT) is the most common type of oral cavity carcinoma. Chemoresistance in SCCT is common, and the underlying mechanism remains largely unknown. We aimed to identify key molecules and signaling pathways mediating chemoresistance in SCCT.Experimental Design: Using a proteomic approach, we found that the HSP27 was a potential mediator for chemoresistance in SCCT cells. To further validate this role of HSP27, we performed various mechanistic studies using in vitro and in vivo models as well as serum and tissue samples from SCCT patients.Results: The HSP27 protein level was significantly increased in the multidrug-resistant SCCT cells and cell culture medium. Both HSP27 knockdown and anti-HSP27 antibody treatment reversed chemoresistance. Inversely, both HSP27 overexpression and recombinant human HSP27 protein treatment enhanced chemoresistance. Moreover, chemotherapy significantly induced HSP27 protein expression in both SCCT cells and their culture medium, as well as in tumor tissues and serum of SCCT patients. HSP27 overexpression predicts a poor outcome for SCCT patients receiving chemotherapy. Mechanically, extracellular HSP27 binds to TLR5 and then activates NF-κB signaling to maintain SCCT cell survival. TLR5 knockdown or restored IκBα protein level disrupts extracellular HSP27-induced NF-κB transactivation and chemoresistance. Moreover, intracellular HSP27 binds to BAX and BIM to repress their translocation to mitochondrion and subsequent cytochrome C release upon chemotherapy, resulting in inhibition of the mitochondrial apoptotic pathway.Conclusions: HSP27 plays a pivotal role in chemoresistance of SCCT cells via a synergistic extracellular and intracellular signaling. HSP27 may represent a potential biomarker and therapeutic target for precision SCCT treatment. Clin Cancer Res; 24(5); 1163-75. ©2017 AACR.

Physical Exercise Enhanced Heat Shock Protein 60 Expression and Attenuated Inflammation in the Adipose Tissue of Human Diabetic Obese.

Heat shock protein 60 (HSP60) is a key protein in the crosstalk between cellular stress and inflammation. However, the status of HSP60 in diabetes and obesity is unclear. In the present study, we investigated the hypothesis that HSP60 expression levels in the adipose tissue of human obese adults with and without diabetes are different and physical exercise might affect these levels. Subcutaneous adipose tissue (SAT) and blood samples were collected from obese adults with and without diabetes (n = 138 and n = 92, respectively, at baseline; n = 43 for both groups after 3 months of physical exercise). Conventional RT-PCR, immunohistochemistry, immunofluorescence, and ELISA were used to assess the expression and secretion of HSP60. Compared with obese adults without diabetes, HSP60 mRNA and protein levels were decreased in SAT in diabetic obese together with increased inflammatory marker expression and glycemic levels but lower VO2 Max. More interestingly, a 3-month physical exercise differentially affected HSP60 expression and the heat shock response but attenuated inflammation in both groups, as reflected by decreased endogenous levels of IL-6 and TNF-α. Indeed, HSP60 expression levels in SAT were significantly increased by exercise in the diabetes group, whereas they were decreased in the non-diabetes group. These results were further confirmed using immunofluorescence microscopy and anti-HSP60 antibody in SAT. Exercise had only marginal effects on HSP60 secretion and HSP60 autoantibody levels in plasma in both obese with and without diabetes. Physical exercise differentially alleviates cellular stress in obese adults with and without diabetes despite concomitant attenuation of the inflammatory response.

Elevated circulating heat shock protein 70 and its antibody concentrations in chronic spontaneous urticaria.

Heat shock proteins (Hsp) play a complex role in cytoprotection, inflammation, and function of the immune system. They may be involved in pathogenesis of various diseases. Our aim was to determine circulating Hsp70 and anti-Hsp70 antibodies concentrations in patients with chronic spontaneous urticaria (CSU). Concentrations of Hsp70 in plasma and anti-Hsp70 antibodies in serum as well as serum C-reactive protein (CRP) were measured in CSU patients and in the controls. Plasma Hsp70 concentrations were significantly higher in CSU (all) and mild CSU patients as compared with the controls. Moderate–severe CSU patients tended to show higher Hsp70 concentration as compared with the controls, but not with mild activity of the disease. There were no significant differences in Hsp70 concentration between moderate–severe and mild CSU patients. Serum anti-Hsp70 antibodies concentrations were significantly higher in CSU (all) and mild CSU in comparison to the controls. Association was observed between anti-Hsp70 antibodies and increased CRP concentration; however, no correlation between anti-Hsp70 and Hsp70 concentrations was seen in the patients. It seems that up-regulation of Hsp70 in CSU may induce marked increase in anti-Hsp70 antibodies production, which are accompanied by parallel changes in CRP concentration. We suggest that Hsp may be released in CSU in response to stressful stimuli, such as inflammation.

Optimal management of Cogan's syndrome: a multidisciplinary approach.

Cogan’s syndrome (CS) is a rare disorder characterized by nonsyphilitic interstitial keratitis (IK) and audio-vestibular symptoms. CS affects mainly young Caucasian adults, mostly during their first three decades of age, and may develop into typical and atypical variants. Typical CS manifests primarily with IK and hearing loss, whereas atypical CS usually presents with inflammatory ocular manifestations in association with audio-vestibular symptoms but mostly different Ménière-like symptoms and, more frequently, with systemic inflammation (70%), of which vasculitis is the pathogenic mechanism. CS is considered as an autoimmune- or immune-mediated disease supported mainly by the beneficial response to corticosteroids. Using well-developed assays, antibodies to inner ear antigens, anti-Hsp70, and antineutrophil cytoplasmic antibodies were found to be associated with CS. Corticosteroids represent the first line of treatment, and multiple immunosuppressive drugs have been tried with variable degrees of success. Tumor necrosis factor-alpha blockers and other biological agents are a recent novel therapeutic option in CS. Cochlear implantation is a valuable rescue surgical strategy in cases with severe sensorineural hearing loss unresponsive to intensive and/or innovative immunosuppressive regimens.

Blocking ATP-sensitive potassium channel alleviates morphine tolerance by inhibiting HSP70-TLR4-NLRP3-mediated neuroinflammation

BackgroundLong-term use of morphine induces analgesic tolerance, which limits its clinical efficacy. Evidence indicated morphine-evoked neuroinflammation mediated by toll-like receptor 4 (TLR4) - NOD-like receptor protein 3 (NLRP3) inflammasome was important for morphine tolerance. In our study, we investigated whether other existing alternative pathways caused morphine-induced activation of TLR4 in microglia. We focused on heat shock protein 70 (HSP70), a damage-associated molecular pattern (DAMP), which was released from various cells upon stimulations under the control of KATP channel and bound with TLR4-inducing inflammation. Glibenclamide, a classic KATP channel blocker, can improve neuroinflammation by inhibiting the activation of NLRP3 inflammasome. Our present study investigated the effect and possible mechanism of glibenclamide in improving morphine tolerance via its specific inhibition on the release of HSP70 and activation of NLRP3 inflammasome induced by morphine.MethodsCD-1 mice were used for tail-flick test to evaluate morphine tolerance. The microglial cell line BV-2 and neural cell line SH-SY5Y were used to investigate the pharmacological effects and the mechanism of glibenclamide on morphine-induced neuroinflammation. The activation of microglia was accessed by immunofluorescence staining. Neuroinflammation-related cytokines were measured by western blot and real-time PCR. The level of HSP70 and related signaling pathway were evaluated by western blot and immunofluorescence staining.ResultsMorphine induced the release of HSP70 from neurons. The released HSP70 activated microglia and triggered TLR4-mediated inflammatory response, leading to the phosphorylation of p38 mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB) p65 and the activation of NLRP3 inflammasome. Moreover, anti-HSP70 neutralizing antibody partly attenuated chronic morphine tolerance. The secretion of HSP70 was under the control of MOR/AKT/KATP/ERK signal pathway. Glibenclamide as a classic KATP channel blocker markedly inhibited the release of HSP70 induced by morphine and suppressed HSP70-TLR4-NLRP3 inflammasome-mediated neuroinflammation, which consequently attenuated morphine tolerance.ConclusionsOur study indicated that morphine-induced extracellular HSP70 was an alternative way for the activation of TLR4-NLRP3 in analgesic tolerance. The release of HSP70 was regulated by MOR/AKT/KATP/ERK pathway. Our study suggested a promising target, KATP channel and a new leading compound, glibenclamide, for treating morphine tolerance.

Simultaneous induction of HSP70 expression, and degranulation, in IgE/Ag-stimulated or extracellular HSP70-stimulated mast cells.

In mast cells, induction of HSP70 expression during antigen stimulation has not been reported. Mouse bone marrow-derived mast cells (BMMC) were stimulated with IgE/Ag or HSP70. Induction of HSP70 expression and signaling protein phosphorylation were evaluated by immunoblotting. HSP70 expression is induced in BMMC at an early stage of IgE/Ag-dependent stimulation, some of which is released from the cells in a granule-associated form. Induction of HSP70 expression was also observed with an IgE/Ag-stimulated human basophilic cell line, indicating that the phenomenon is not restricted to mouse BMMC. The induction of HSP70 expression, and its release, followed a similar time course to that of degranulation. Released HSP70 seems to be responsible for degranulation and production of eicosanoids, at least in part, because a neutralizing anti-HSP70 antibody mitigated these activities and because exogenous HSP70 not only induced immediate degranulation followed by autocrine HSP70 expression but also enhanced degranulation in IgE/Ag-stimulated BMMC. Extracellular HSP70 was found to induce phosphorylation of linker for activation of T cells (LAT) and a series of downstream signaling molecules in BMMC. We further found that Fyn, Lyn, and spleen tyrosine kinase (Syk), which are known to concern LAT phosphorylation in IgE/Ag-stimulated BMMC, were not phosphorylated in HSP70-stimulated BMMC, whereas lymphocyte-specific protein tyrosine kinase (Lck) was phosphorylated. FcεRI stimulation in BMMC and basophils induces HSP70 expression and its release. Extracellular HSP70 induces degranulation and mediator release via phosphorylation of LAT.

Effect of heat stress during in vitro maturation on developmental competence of vitrified bovine oocytes.

Heat-shock protein 70 (HSP70) plays a crucial role as intracellular cytoprotectant and molecular chaperone. A phenomenon of heat stress (HS) leads to production of these proteins that could be beneficial to cells during cryopreservation, which is also a stressful process for the cell. This study aimed to evaluate the protective effect of exposure of bovine oocytes to moderate HS during in vitro maturation (IVM) prior vitrification. First, oocytes were subjected to HS (41.5°C for 1hr) at 0, 4, 8, 12 or 16 of IVM. Oocytes in vitro matured for 20hr served as control group. Presence of HSP70 was detected at 20hr by immunofluorescence. HSP70 expression was significantly higher when oocytes were subjected to HS at 8hr of IVM. Next, oocytes were distributed into four groups: Control: IVM oocytes; VIT: oocytes vitrified/warmed at 20hr of IVM; HS: oocytes subjected to HS at 8hr of IVM; HS-VIT: oocytes subjected to HS at 8hr of IVM and vitrified/warmed at 20hr of IVM. Oocytes were fertilized at 24hr of IVM, and cleavage and blastocyst yield were assessed. No significant differences were observed among treatments when cleavage rate was evaluated. However, fresh control and HS oocytes resulted in a significantly higher (p<.05) blastocyst rate when compared to VIT and HS-VIT groups, although no significant differences within fresh or vitrified groups were observed. In conclusion, HS did not have a negative impact on the oocyte competence but HS applied before vitrification, offered no benefits for embryo development.

Role of Heat Shock Proteins in Atherosclerosis and Atrial Fibrillation

Heat Shock Proteins (HSPs) are a group of highly conserved molecular chaperones that play important roles in homeostasis and the cellular response to stress. In the cardiovascular system, HSPs maintain vessel wall integrity and cardiomyocyte function; they may be upregulated and translocated to the cell surface in response to environmental stressors. HSPs contribute to the inflammatory processes that mediate the development of atherosclerosis by activating the innate and adaptive immune response. Misdirected autoimmune attack of surface HSPs on stressed vascular endothelial cells by circulating anti-Hsp60 antibodies lead to damage of the vessel wall and atherogenesis. Additionally, other HSPs contribute to the pro-inflammatory state in the vessel wall by stimulating macrophages and reactive Tcells to release cytokine and chemotactic factors. Atrial Fibrillation (AF) is closely related to atherosclerotic burden and changes to levels of various HSPs have been associated with increased incidence of AF in stressed cardiomyocytes after ischemia reperfusion injury. Modulation of HSP expression may be a useful therapeutic strategy in the management of atherosclerosis and AF. Keywords: Heat shock proteins, HSP, HSP60, autoimmunity, atherosclerosis, atrial fibrillation.

Increased Anti-HSP60 and Anti-HSP70 Antibodies in Women with Unexplained Recurrent Pregnancy Loss.

Vascular dysfunction has been reported in women with recurrent pregnancy loss (RPL). We investigated the severity of vascular dysfunction in non-pregnant women with RPL and its correlation with anti-heat shock protein (HSP) antibodies that are known to induce arteriosclerosis. We measured the serum anti-HSP60 antibodies, anti-HSP70 antibodies, and anti-phospholipid antibodies (APA) in 68 women with RPL and 29 healthy controls. Among the women with RPL, 14 had a diagnosis of antiphospholipid syndrome (APS), and in the remaining 54, the causes for RPL were unexplained. Compared to the controls, the brachial-ankle pulse wave velocity (baPWV), carotid augmentation index (cAI), and uterine artery pulsatility index (PI) were all significantly higher in the women with both APS and unexplained RPL. Compared to the controls, the anti-HSP60 antibody levels were significantly higher in the APA-positive group of women with unexplained RPL, and the anti-HSP70 antibody levels were significantly higher in APS and APA-positive group of women with unexplained RPL. However, the anti-HSP60 and anti-HSP70 antibody levels did not correlate with the values of baPWV or cAI. Our results demonstrated anti-HSP60 and anti-HSP70 antibodies are increased in women with unexplained RPL. Further studies are needed to elucidate the roles of anti-HSP antibodies and their pathophysiology in unexplained RPL.

Application of antibodies to recombinant heat shock protein 70 in immunohistochemical diagnosis of mycobacterium avium subspecies paratuberculosis in tissues of naturally infected cattle

BackgroundDetection of Mycobacterium avium subspecies paratuberculosis (MAP) infection is key to the control of Johne’s disease. Immunohistochemistry is one of the methods of detection of MAP infection in tissues. However, unavailability of commercial antibodies that can detect the organism is a limiting factor for the use of immunohistochemistry. This study was aimed at developing an immunohistochemistry method to diagnose MAP in infected tissues using antibodies against MAP recombinant heat shock protein 70kd.ResultsMAP Heat shock protein 70 gene was amplified and cloned into an expression vector, Champion pET-SUMO, then expressed in E coli, purified and used to produce polyclonal rabbit antibodies against the Heat shock protein. Immunohistochemistry was performed in 35 MAP infected tissues with anti-HSP70 polyclonal antibodies. All 35 MAP infected tissues were positive for MAP within macrophages, epithelioid cells and giant cells either in clumps or singly as individual bacilli. No positive staining was seen in the three uninfected normal tissues and in MAP infected tissues where primary antibodies were substituted with PBS or pre-immune serum from the same rabbit.ConclusionAnti-HSP70 produced in this study offers an opportunity for improved diagnosis, screening of MAP in animal tissues and in studies on the pathogenesis of MAP

Autoantibodies against heat shock proteins as biomarkers for the diagnosis and prognosis of cancer.

Thanks to their specificity and stability in the sera, autoantibodies (AAbs) against tumor-associated antigens (TAAs) are very attractive biomarkers for the development of less invasive serological tests for the diagnosis and prognosis of cancer. Heat shock proteins (HSP) belong to TAAs and they are over-expressed in various human cancers. Elevated HSP can stimulate the immune system to produce anti-HSP antibodies. So far, AAbs against HSP have been identified in the circulation of various cancer patients. Here we will review current literature on the use of anti-HSP antibodies for cancer diagnosis and prognosis. The challenges as well as future directions of AAbs identification in oncology are also discussed.

Relationship between serum anti-heat shock protein 27 antibody levels and obesity

BackgroundHeat shock protein 27 (HSP27) is an intracellular molecular chaperone that is expressed at high levels following the exposure of cells to environmental stressors such as heat, toxins, and free radicals. High levels of HSP antigens and antibody titers have been reported in several conditions including cardiovascular disease and cancers. We measured serum anti-HSP27 antibody levels in 993 subjects and assessed the associations between serum anti-HSP27 antibody levels and demographic characteristics including coronary risk factors. MethodsA total of 993 subjects were recruited as part of the Mashhad Stroke and Heart Atherosclerotic Disorders (MASHAD) cohort study. Demographic, clinical, and biochemical parameters and serum anti-HSP27 antibody titers were determined in all the subjects. ResultsSerum anti-HSP27 antibody levels increased with increasing age in men. No significant differences in levels were detected between men and women. Serum anti-HSP27 antibody levels were significantly higher in obese subjects than in nonobese subjects (P=0.046); however, no significant influence of smoking status was observed. Moreover, serum anti-HSP27 antibody titers were positively associated with age, body mass index, waist/hip ratio, the presence of diabetes mellitus, nonsmoking habit, serum triglycerides, cholesterol, and high-sensitivity c-reactive protein. ConclusionWe have found that serum anti-HSP27 antibody titers are related to several cardiovascular risk factors, necessitating further studies on the value of this emerging marker for risk stratification.