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.

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The thermo-optic mechanism in thylakoid membranes was earlier identified by measuring the thermal and light stabilities of pigment arrays with different levels of structural complexity [Cseh, Z., et al. (2000) Biochemistry 39, 15250-15257]. (According to the thermo-optic mechanism, fast local thermal transients, arising from the dissipation of excess, photosynthetically not used, excitation energy, induce elementary structural changes due to the "built-in" thermal instabilities of the given structural units.) The same mechanism was found to be responsible for the light-induced trimer-to-monomer transition in LHCII, the main chlorophyll a/b light-harvesting antenna of photosystem II (PSII) [Garab, G., et al. (2002) Biochemistry 41, 15121-15129]. In this paper, differential scanning calorimetry (DSC) and circular dichroism (CD) spectroscopy on thylakoid membranes of barley and pea are used to correlate the thermo-optically inducible structural changes with well-discernible calorimetric transitions. The thylakoid membranes exhibited six major DSC bands, with maxima between about 43 and 87 degrees C. The heat sorption curves were analyzed both by mathematical deconvolution of the overall endotherm and by a successive annealing procedure; these yielded similar thermodynamic parameters, transition temperature and calorimetric enthalpy. A systematic comparison of the DSC and CD data on samples with different levels of complexity revealed that the heat-induced disassembly of chirally organized macrodomains contributes profoundly to the first endothermic event, a weak and broad DSC band between 43 and 48 degrees C. Similarly to the main macrodomain-associated CD signals, this low enthalpy band could be diminished by prolonged photoinhibitory preillumination, the extent of which depended on the temperature of preillumination. By means of nondenaturing, "green" gel electrophoresis and CD fingerprinting, it is shown that the second main endotherm, around 60 degrees C, originates to a large extent from the monomerization of LHCII trimers. The main DSC band, around 70 degrees C, which exhibits the highest enthalpy change, and another band around 75-77 degrees C relate to the dismantling of LHCII and other pigment-protein complexes, which under physiologically relevant conditions cannot be induced by light. The currently available data suggest the following sequence of events of thermo-optically inducible changes: (i) unstacking of membranes, followed by (ii) lateral disassembly of the chiral macrodomains and (iii) monomerization of LHCII trimers. We propose that thermo-optical structural reorganizations provide a structural flexibility, which is proportional to the intensity of the excess excitation, while for their localized nature, the structural stability of the system can be retained.

This study describes the effect of collagen type I (Col I) oxidation on its physiological remodeling by adipose tissue-derived mesenchymal stem cells (ADMSCs), both mechanical and proteolytic, as an in vitro model for the acute oxidative stress that may occur in vivo upon distinct environmental changes. Morphologically, remodeling was interpreted as the mechanical rearrangement of adsorbed FITC-labelled Col I into a fibril-like pattern. This process was strongly abrogated in cells cultured on oxidized Col I albeit without visible changes in cell morphology. Proteolytic activity was quantified utilizing fluorescence de-quenching (FRET effect). The presence of ADMSCs caused a significant increase in native FITC-Col I fluorescence, which was almost absent in the oxidized samples. Parallel studies in a cell-free system confirmed the enzymatic de-quenching of native FITC-Col I by Clostridial collagenase with statistically significant inhibition occurring in the oxidized samples. Structural changes to the oxidized Col I were further studied by differential scanning calorimetry. In the oxidized samples, an additional endotherm with sustained enthalpy (∆H) was observed at 33.6 °C along with Col I’s typical one at 40.5 °C. Collectively, these data support that the remodeling of Col I by ADMSCs is altered upon oxidation due to intrinsic changes to the protein’s structure, which represents a novel mechanism for the control of stem cell behavior.

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Erythrocytes (RBCs) constitute a very interesting class of cells both for their physiological function and for a variety of peculiarities. Due to their exceptionally strong relationship with the environment, the morphology and nanoscale characteristics of these cells can reveal their biochemical status and structural integrity. Among the possible subjects of investigations, the RBCs' ageing is of the utmost importance. This is a fundamental phenomenon that, in physiological conditions, triggers the cell turnover and ensures the blood homeostasis. With these premises, in recent years, we have presented an atomic force microscopy-based methodology to characterize the patterns of RBC ageing from the morphological point of view. In the present work, we used an ageing protocol more similar to the physiological conditions and we used differential scanning calorimetry and atomic force microscopy to probe the cross correlation between important structural and functional proteins. We also assessed the role played by fundamental structural and membrane proteins in the development of the most relevant morphological intermediates observed along the ageing. Furthermore, we coupled the morphological ageing patterns to the (bio)chemical alterations detected by Raman spectroscopy. This allowed identifying the chronology of the ageing morphologies and the metabolic pathways most involved in their development. As a whole, the present study provides the base to correlate specific molecular alterations to the development of structural anomalies, and these latter to the functional status of blood cells.

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