Blood Plasma Calorimetric Profiles of Women with Preeclampsia: Effect of Oxidative Stress.

Preeclampsia is a pregnancy-related disease with poor placentation and presents itself through hypertension and proteinuria. The disease is also associated with the oxidative modification of proteins in maternal blood plasma. In this work, we combine differential scanning calorimetry (DSC), capillary electrophoresis, and atomic force microscopy (AFM) to evaluate the changes in the plasma denaturation profiles of patients with preeclampsia (PE) as compared with those of pregnant controls. Our results demonstrate that the last trimester of pregnancy substantially affects the main calorimetric characteristics of blood plasma from pregnant controls relative to nonpregnant women. These variations correlate well with the changes in protein levels determined by electrophoresis. DSC analysis revealed significant deviations in the plasma heat capacity profiles of preeclamptic patients from those of pregnant controls. These alterations are expressed mainly in a substantial reduction in albumin-assigned transitions and an upward shift in its denaturation temperature, lower calorimetric enthalpy changes, and a reduced ratio of heat capacity in the albumin/globulin-assigned thermal transitions, which are more pronounced in severe PE cases. The in vitro oxidation model shows that the alteration of PE thermograms is partly related to protein oxidation. AFM data detected numerous aggregate formations in the plasma of PE samples and fewer small ones in the pregnant controls, which are not found in healthy nonpregnant samples. These findings could serve as a basis for further investigations to reveal the possible relationship between albumin thermal stabilization, the increased inflammatory state and oxidative stress, and protein misfolding in preeclampsia.

Thermodynamic Signatures of Blood Plasma Proteome in Neurodegenerative Pathologies.

Discovery of diagnostic biomarkers for age-related neurodegenerative pathologies (NDDs) is essential for accurate diagnosis, following disease progression and drug development. Blood plasma and blood cells are important peripheral sources for NDDs' biomarkers that, although present in lower concentrations than in cerebrospinal fluid, would allow noninvasive diagnostics. To identify new biomarkers for Parkinson's disease (PD) and amyotrophic lateral sclerosis (ALS), in this work we have evaluated the modifications in the thermodynamic behavior of blood plasma proteome exploring differential scanning calorimetry. The plasma thermodynamics reflected the complexity and heterogeneity of the two pathologies. The unfolding temperature of the most abundant plasma protein albumin and the weighted average center of the calorimetric profile appeared as the two thermodynamic signatures that reflected modifications of the plasma proteome, i.e., strong thermal stabilization of albumin and plasma proteins' interaction network, related to both pathologies. Based on those two signatures, both PD and ALS patients were stratified in two sets, except several cases with thermodynamic parameters that strongly differed from those of the calorimetric sets. Along with modifications of the plasma thermodynamic behavior, we found altered globulin levels in all PD and ALS patients' plasma (higher level of α- and ß-globulin fractions and lower level of γ-globulin fraction than the respective reference values) employing capillary electrophoresis. The presented results reveal the potential of calorimetry to indirectly identify NDDs' biomarkers in blood plasma.

Altered Thermal Behavior of Blood Plasma Proteome Related to Inflammatory Cytokines in Early Pregnancy Loss.

Early pregnancy loss (EPL) is a relatively common pathology of which almost 50% of cases remain idiopathic. In the search for novel biomarkers, differential scanning calorimetry (DSC) is intensively used to characterize the thermodynamic behavior of blood plasma/serum proteome in health and disease. Herein, for the first time, we investigate the DSC denaturation profiles of blood plasma derived from patients suffering EPL compared to healthy pregnant and non-pregnant women. Data analysis reveals that 58% of the EPL thermograms differ significantly from those of healthy pregnant women. Thermal stabilization of a fraction of albumin-assigned transition with concomitant suppression of the major and enhancement of the globulin-assigned transition are characteristic features of EPL calorimetric profiles that could be used as a new indicator of a risk pregnancy. The presented results suggest an altered composition or intermolecular interactions of the plasma proteome of women with EPL. In addition, the alterations of the EPL thermograms correlate with the increased blood levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) and a higher prevalence of the polymorphism in the plasminogen activator inhibitor type-1 (PAI-1) gene, suggesting an expression of an overall enhanced immune response. The concomitant changes in plasma thermograms confirm the potential of the DSC approach for distinguishing changes in the pathological state of the blood plasma proteome.

Calorimetric markers for monitoring of multiple myeloma and Waldenström's macroglobulinemia patients.

The blood proteome has been studied extensively for identification of novel reliable disease biomarkers. In recent years, differential scanning calorimetry has emerged as a new tool for characterization of the thermodynamic properties of the major serum/plasma proteins and for the establishment of calorimetric markers for a variety of diseases. Here we applied calorimetry to monitor the effect of treatment of patients diagnosed with multiple myeloma and Waldenström's macroglobulinemia on the calorimetric profiles of patients' blood sera. The parameters derived from the calorimetric profiles were compared with the primary serum biomarkers, monoclonal immunoglobulin (M protein) concentration, and κ/λ free light chain ratio. For the secretory cases, the calorimetric parameters thermogram's shape similarity and weighted average center strongly depended on the M protein level but had lower sensitivity and specificity. By contrast, for non-secretory cases, the calorimetric parameters did not depend on the κ/λ free light chains ratio and exhibited significantly higher sensitivity and specificity than M protein levels. A combination of the immunological and calorimetric tests was found to greatly improve the sensitivity and specificity of the clinical status evaluation. The pronounced differences in blood sera thermograms before and during monitoring reflected the individual patients' response to treatment received and showed maintenance of heterogeneity during the disease course.

Calorimetric markers of Bence Jones and nonsecretory multiple myeloma serum proteome.

The present work provides a thermodynamic description of blood serum from patients diagnosed with Bence Jones myeloma (BJMM) and nonsecretory myeloma (NSMM) by means of differential scanning calorimetry (DSC), serum protein electrophoresis, and free light chain assay. Specific alterations in the thermodynamic behavior of both BJMM and NSMM proteome have been revealed. On the basis of the transition temperature of the main transition in the calorimetric profiles and the shape similarity criterion, we defined BJMM and NSMM sets/subsets of thermograms with very similar thermodynamic features. We show that some of the BJMM and NSMM subsets correlate with previously defined secretory myeloma subsets (Todinova et al. Anal. Chem. 2011, 83, 7992). The established analogies strongly suggest that common molecular markers contribute to the calorimetric profiles of the different, secretory and nonsecretory, myeloma types; our data show robust evidence that these are ligands stabilizing the major serum proteins. We demonstrate that the DSC approach might be highly beneficial, especially for NSMM patients, since the characteristic modifications in the DSC profiles might serve as calorimetric markers when no monoclonal proteins can be detected in the bloodstream and the diagnosis heavily relies on invasive methods.

Microcalorimetry of blood serum proteome: a modified interaction network in the multiple myeloma case.

Hereby we report on a novel approach in the study of multiple myeloma (MM), namely, differential scanning calorimetry (DSC) combined with serum protein electrophoresis. Distinct thermodynamic signatures describe the DSC thermograms of MM blood sera, in contrast to the unique profile found for healthy individuals. The thermal behavior of MM sera reflects a complex interplay between the serum concentration and isotype of the M protein and of albumin, and modified ligand- and/or protein-protein interactions, resulting in stabilization of globulins and at least a fraction of albumin. In all MM cases the 85 °C, transferrin-assigned transition is missing. A distinct feature of IgG isotype (κ and λ) DSC profiles only is the presence of a transition at 82 °C. A DSC-based classification of MM depicts two sets of melting patterns (MMt2 and MMt3 with two or three successive thermal transitions), and subsets within each set (MMt2(i) or MMt3(i), the subscript i = 1, 2 or 3 denotes the main transition being one of the three transitions). The results demonstrate the potential of DSC to monitor MM-related modifications of the serum proteome, even at low M protein concentrations, Bence Jones and importantly nonsecretory multiple myeloma cases, and prove DSC as a versatile tool for oncohematology.