Zoltán Prohászka

The 90-kDa Molecular Chaperone Family: Structure, Function, and Clinical Applications. A Comprehensive Review

The 90-kDa molecular chaperone family (which comprises, among other proteins, the 90-kDa heat-shock protein, hsp90 and the 94-kDa glucose-regulated protein, grp94, major molecular chaperones of the cytosol and of the endoplasmic reticulum, respectively) has become an increasingly active subject of research in the past couple of years. These ubiquitous, well-conserved proteins account for 1-2% of all cellular proteins in most cells. However, their precise function is still far from being elucidated. Their involvement in the aetiology of several autoimmune diseases, in various infections, in recognition of malignant cells, and in antigen-presentation already demonstrates the essential role they likely will play in clinical practice of the next decade. The present review summarizes our current knowledge about the cellular functions, expression, and clinical implications of the 90-kDa molecular chaperone family and some approaches for future research.

Red cell distribution width in heart failure: Prediction of clinical events and relationship with markers of ineffective erythropoiesis, inflammation, renal function, and nutritional state

OBJECTIVES The goal of this study was to independently validate the recent observations on the predictive role of red cell distribution width (RDW) for outcomes in chronic heart failure and to provide epidemiologic data on the biological correlates of RDW in heart failure (HF). Understanding the mechanism underlying this observation is unclear, largely hampered by the lack of epidemiologic studies demonstrating factors that are associated with anisocytosis in cardiovascular diseases. METHODS One hundred ninety-five patients (145 men, 50 women) with systolic HF were enrolled and followed up for a median of 14.5 months. Primary end points were all-cause mortality and hospital readmission due to worsening HF symptoms. A total of 19 clinical chemistry, hematology, and biochemical variables were considered for analysis together with clinical parameters in Cox proportional hazards and multiple regression models. RESULTS Red cell distribution width was found to be an N-terminal pro-brain natriuretic peptide independent predictor of all-cause mortality (adjusted HR 1.61 per 1 SD increase) in our study. Multiple correlations between biomarkers of ineffective erythropoiesis (serum iron, ferritin, and soluble transferrin receptor levels), inflammation and acute-phase reaction (interleukin-6, soluble tumor necrosis factor (TNF) receptor I and soluble TNF receptor II, C-reactive protein, and prealbumin concentrations), undernutrition (total cholesterol and albumin levels), and renal function were observed. In the multiple regression model, the strongest relationship for RDW was obtained with soluble transferrin receptor, soluble TNF receptor I, soluble TNF receptor II, and total cholesterol. CONCLUSIONS Here we validate the strong, independent prediction of morbidity and mortality in HF by RDW. The described correlations between RDW and inflammation, ineffective erythropoiesis, undernutrition, and impaired renal function may facilitate the understanding why this marker is associated with adverse outcomes in HF.

Involvement of polymorphisms in the chemokine system in the susceptibility for coronary artery disease (CAD). Coincidence of elevated Lp(a) and MCP-1 -2518 G/G genotype in CAD patients

The central role of chemokines in the pathogenesis of atherosclerosis has been made clear. Recently polymorphisms in the gene regulatory region of MCP-1 and in the promoter region of RANTES have been found, which increase the expression of these chemokines. We investigated the role of these polymorphisms together with the chemokine SDF-1-801A and the chemokine receptors CCR2-64I and CCR5Delta32 mutations in 318 patients with coronary artery disease (CAD) referred to coronary bypass surgery, comparing them with 320 healthy controls. The prevalence of the MCP-1 -2518 G/G homozygotes was significantly higher among CAD patients than among controls (P<0.005; OR=2.2 (95% CI 1.25-3.92). The Lp(a) levels of CAD patients with G/G genotype were significantly higher than those in patients with G/A or A/A genotypes. No CAD patients homozygous for the CCR5Delta32 and CCR2-64I mutations have been found. The genotype distributions of the two alleles deviated from the Hardy Weinberg equilibrium in patients, indicating that the numbers of homozygotes were significantly lower than expected. The MCP-1 -2518G variant in homozygous form appears as a genetic risk factor for severe CAD. This genotype is associated with elevated Lp(a) levels in patients. Individuals homozygous for CCR2-64I or CCR5Delta32 mutations are at reduced risk for severe CAD.

Independent and Joint Effects of Antibodies to Human Heat-Shock Protein 60 and Chlamydia pneumoniae Infection in the Development of Coronary Atherosclerosis

BackgroundStudies have suggested that the prevalence of antibodies against heat-shock proteins (HSPs), Chlamydia pneumoniae (Cpn), and cytomegalovirus (CMV) is associated with coronary artery disease (CAD), but the independent or joint effects of human (h) HSP60 antibodies and these pathogens in patients have not been fully elucidated. Methods and ResultsA total of 405 subjects (276 patients with CAD and 129 control individuals) were tested for serum antibodies to hHSP60, Cpn, and CMV immediate-early-1 (IE1) antigens. Patients were also assessed for serum cholesterol, triglyceride levels, and smoking habit. Significantly elevated levels of antibodies to hHSP60 and Cpn but not to CMV-IE1 antigens were documented in CAD patients. Multiple logistic regression analysis and subanalyses of selected subjects showed that these associations were independent of age, sex, smoking, and serum lipid levels. Antibodies to hHSP60 and Cpn did not correlate quantitatively; however, the relative risk of disease development was substantially increased in subjects with high antibody levels to both hHSP60 and Cpn, reaching an odds ratio of 82.0 (95% CI 10.6 to 625.0). ConclusionsHigh levels of antibodies to hHSP60 and Cpn are independent risk factors for coronary atherosclerosis, but their simultaneous presence substantially increases the risk for disease development.

Activation of the complement system in normal pregnancy and preeclampsia.

The purpose of this study was to explore the role of the complement system in normal human pregnancy and preeclampsia in a comprehensive manner, measuring circulating levels of complement proteins, their activation fragments and regulatory factors, as well as those of C-reactive protein (CRP). Sixty preeclamptic patients, 60 healthy pregnant women and 59 healthy non-pregnant women were involved in this case-control study. Circulating levels of complement components and CRP were determined with ELISA, radial immunodiffusion and particle enhanced immunoturbidimetric assay. Levels of CRP, C4d, C3a, SC5b9, C3, C9 and factor H antigen were significantly higher, while those of C1-inhibitor were significantly lower in healthy pregnant than non-pregnant women. In addition, preeclamptic patients had significantly higher CRP, C4d, C3a, SC5b9 levels and significantly lower C3 concentrations as compared to healthy pregnant women. Their CRP, C4d, C3a, SC5b9, C4, C3, C9 and factor H antigen levels were significantly higher, while C1-inhibitor concentrations were significantly lower compared with healthy non-pregnant women. However, no significant difference was found in Bb and C4b-binding protein levels among the three study groups. Preeclamptic patients with fetal growth restriction had significantly higher plasma SC5b9 levels than those without IUGR. There was a relative deficiency of C1-inhibitor and C4b-binding protein, and a relative abundance of factor H both in normal pregnancy and preeclampsia. Activation of the classical or lectin pathway (C4d) showed significant positive correlation to C3 activation (C3a) both in healthy pregnant women and preeclamptic patients. However, the correlation between C3 and terminal pathway activation was dominating only in patients with preeclampsia, but not in healthy pregnant women. In conclusion, the complement system is activated through the classical and/or lectin pathways with increased terminal complex formation in the third trimester of normal human pregnancy, and further in preeclampsia, as shown by the elevated amounts of activation markers in the systemic circulation. Excessive activation of the terminal pathway is associated with fetal growth restriction in preeclamptic women. However, additional studies are required to determine the cause and consequence of systemic complement activation in this pregnancy-specific disorder.

Heat shock protein 70 is a potent activator of the human complement system

Abstract According to new hypotheses, extracellular heat shock proteins (Hsps) may represent an ancestral danger signal of cellular death or lysis-activating innate immunity. Recent studies demonstrating a dual role for Hsp70 as both a chaperone and cytokine, inducing potent proinflammatory response in human monocytes, provided support for the hypothesis that extracellular Hsp is a messenger of stress. Our previous work focused on the complement-activating ability of human Hsp60. We demonstrated that Hsp60 complexed with specific antibodies induces a strong classical pathway (CP) activation. Here, we show that another chaperone molecule also possesses complement-activating ability. Solid-phase enzyme-linked immunosorbent assay was applied for the experiments. Human Hsp70 activated the CP independently of antibodies. No complement activation was found in the case of human Hsp90. Our data further support the hypothesis that chaperones may messenger stress to other cells. Complement-like molecules and primitive immune cells appeared together early in evolution. A joint action of these arms of innate immunity in response to free chaperones, the most abundant cellular proteins displaying a stress signal, may further strengthen the effectiveness of immune reactions.

Complement activation in thrombotic thrombocytopenic purpura

Summary.  Background:  Ultra‐large von Willebrand factor and deficiency of its cleaving protease are important factors in the events leading to thrombotic microangiopathy; however, the mechanisms involved are only partly understood. Whereas pathological activation of the alternative complement pathway is linked to atypical hemolytic uremic syndrome, the role of complement activation in thrombotic thrombocytopenic purpura (TTP) is unknown. The aim of this study was to investigate whether signs of complement activation are characteristic of TTP.

Serum leptin levels in relation to circulating cytokines, chemokines, adhesion molecules and angiogenic factors in normal pregnancy and preeclampsia.

ObjectiveIn this study, we determined circulating levels of C-reactive protein, several cytokines, chemokines, adhesion molecules and angiogenic factors along with those of leptin in healthy non-pregnant and pregnant women and preeclamptic patients, and investigated whether serum leptin levels were related to the clinical characteristics and measured laboratory parameters of the study participants.MethodsSixty preeclamptic patients, 60 healthy pregnant women and 59 healthy non-pregnant women were involved in this case-control study. Levels of leptin and transforming growth factor (TGF)-beta1 in maternal sera were assessed by ELISA. Serum levels of interleukin (IL)-1beta, IL-1 receptor antagonist (IL-1ra), IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p40, IL-12p70, IL-18, interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, interferon-gamma-inducible protein (IP)-10, monocyte chemotactic protein (MCP)-1, intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 were determined by multiplex suspension array. Serum C-reactive protein (CRP) concentrations were measured by an autoanalyzer. Serum total soluble fms-like tyrosine kinase-1 (sFlt-1) and biologically active placental growth factor (PlGF) levels were determined by electrochemiluminescence immunoassay. For statistical analyses, non-parametric methods were applied.ResultsThere were significant differences in most of the measured laboratory parameters among the three study groups except for serum IL-1beta and TGF-beta1 levels. Serum leptin levels were significantly higher in preeclamptic patients and healthy pregnant women than in healthy non-pregnant women. Additionally, preeclamptic patients had significantly higher leptin levels as compared to healthy pregnant women. Serum leptin levels were independently associated with BMI in healthy non-pregnant women. In healthy pregnant women, both BMI and serum CRP concentrations showed significant positive linear association with leptin levels. There were significant positive correlations between serum leptin concentrations of healthy pregnant women and systolic blood pressure, as well as serum levels of IP-10, while their serum leptin levels correlated inversely with fetal birth weight. In preeclamptic patients, a significant positive correlation was observed between serum concentrations of leptin and IP-10. Furthermore, elevated serum leptin level and sFlt-1/PlGF ratio had an additive (joint) effect in the risk of preeclampsia, as shown by the substantially higher odds ratios of their combination than of either alone.ConclusionsSimultaneous measurement of leptin with several inflammatory molecules and angiogenic factors in this study enabled us to investigate their relationship, which can help to understand the role of circulating leptin in normal pregnancy and preeclampsia.

Complement Protease MASP-1 Activates Human Endothelial Cells: PAR4 Activation Is a Link between Complement and Endothelial Function

Activation of the complement system can induce and enhance inflammatory reaction. Mannose-binding lectin-associated serine protease-1 (MASP-1) is an abundant protease of the complement lectin pathway; however, its physiological function is unclear. In this study, we demonstrate for the first time that MASP-1 is able to activate Ca2+ signaling, NF-κB, and p38 MAPK pathways in cultured HUVECs. Activation was initiated by MASP-1 only; the related protease, MASP-2, had no such effect. The phenomenon was dependent on the proteolytic activity of MASP-1, suggesting modulation of endothelial cell function through a protease-activated receptor (PAR). Using synthetic peptide substrates representing the protease-sensitive regions of PARs, we were able to demonstrate that PAR4 is a target of MASP-1. The presence of functionally active PAR4 in HUVECs was demonstrated using PAR4 agonist peptide and mRNA quantification. Finally, we showed that the amount of membrane-bound intact PAR4 decreases after MASP-1 treatment. All of these results provide a novel link between the regulation of endothelial cell function and complement system activation, and they suggest that MASP-1-induced PAR4 activation could contribute to the development of the inflammatory reaction.

Association of Chlamydia pneumoniae with coronary artery disease and its progression is dependent on the modifying effect of mannose-binding lectin

Background—The possible association between coronary artery disease (CAD) and Chlamydia pneumoniae (C pneumoniae) infection is controversial. On the basis of the recent suggestion that mannose-binding lectin (MBL) variant alleles are related to an increased risk of severe atherosclerosis, and on the in vitro interaction of MBL with C pneumoniae, we asked whether MBL might contribute to CAD in conjunction with C pneumoniae. Methods and Results—Antibodies to C pneumoniae were measured by immunofluorescence and MBL alleles were determined by polymerase chain reaction technique in samples from 210 patients with CAD and 257 healthy subjects from Hungary collected between 1995 and 1996. A higher percentage of patients with CAD were anti-C pneumoniae positive as compared with the control group (P =0.058). However, at logistic regression analysis adjusted to age, sex, and serum lipid levels, this difference was confined only to subjects carrying MBL variant alleles (P =0.035, odds ratio 2.63, [95% CI: 1.07 to 6.45]). In contrast, no significant difference was seen in those homozygous for the normal MBL allele (P =0.412). During a 65±5.8-month follow-up period, major outcomes (new myocardial infarction, and/or bypass operation or cardiovascular death) occurred in 11 C pneumoniae positive and 3 C pneumoniae negative patients. In the C pneumoniae positive group, the odds ratio of development of outcomes was 3.27 (95% CI: 1.10 to 9.71, P =0.033) in the carriers of the MBL variant alleles compared with the homozygous carriers of the normal MBL allele. Conclusions—These results indicate that infection with C pneumoniae leads mainly to the development and progression of severe CAD in patients with variation in the MBL gene.