Optical Study of Lysozyme Molecules in Aqueous Solutions after Exposure to Laser-Induced Breakdown.

The properties of a lysozyme solution under laser-induced breakdown were studied. An optical breakdown under laser action in protein solutions proceeds with high efficiency: the formation of plasma and acoustic oscillations is observed. The concentration of protein molecules has very little effect on the physicochemical characteristics of optical breakdown. After exposure to optical breakdown, changes were observed in the enzymatic activity of lysozyme, absorption and fluorescence spectra, viscosity, and the sizes of molecules and aggregates of lysozyme measured by dynamic light scattering. However, the refractive index of the solution and the Raman spectrum did not change. The appearance of a new fluorescence peak was observed upon excitation at 350 nm and emission at 434 nm at exposure for 30 min. Previously, a peak in this range was associated with the fluorescence of amyloid fibrils. However, neither the ThT assay nor the circular dichroism dispersion confirmed the formation of amyloid fibrils. Probably, under the influence of optical breakdown, a small part of the protein degraded, and a part changed its native state and aggregated, forming functional dimers or "native aggregates".

Selenium Nanoparticles Can Influence the Immune Response Due to Interactions with Antibodies and Modulation of the Physiological State of Granulocytes.

Currently, selenium nanoparticles (SeNPs) are considered potential immunomodulatory agents and as targets for activity modulation are granulocytes, which have the most abundant population of immune blood cells. The present study aims to evaluate the cytotoxic effect and its effect on the functional responses of granulocytes. In addition to the intrinsic activity of SeNPs, we studied the activity of the combination of SeNPs and IgG antibodies. Using laser ablation and fragmentation, we obtained nanoparticles with an average size of 100 nm and a rather narrow size evolution. The resulting nanoparticles do not show acute toxicity to primary cultures of fibroblasts and hepatocytes, epithelial-like cell line L-929 and granulocyte-like culture of HL-60 at a concentration of 109 NPs/mL. SeNPs at a concentration of 1010 NPs/mL reduced the viability of HL-60 cells by no more than 10% and did not affect the viability of the primary culture of mouse granulocytes, and did not have a genotoxic effect on progenitor cells. The addition of SeNPs can affect the production of reactive oxygen species (ROS) by mouse bone marrow granulocytes, modulate the proportion of granulocytes with calcium spikes and enhance fMLF-induced granulocytes degranulation. SeNPs can modulate the effect of IgG on the physiological responses of granulocytes. We studied the expression level of genes associated with inflammation and cell stress. SeNPs increase the expression of catalase, NF-κB, Xrcc5 and some others; antibodies enhance the effect of SeNPs, but IgG without SeNPs decreases the expression level of these genes. This fact can be explained by the interaction between SeNPs and IgG. It has been established that antibodies interact with SeNPs. We showed that antibodies bind to the surface of selenium nanoparticles and are present in aqueous solutions in a bound form from DLS methods, ultraviolet-visible spectroscopy, vibrational-rotational spectrometry, fluorescence spectrometry, and refractometry. At the same time, in a significant part of the antibodies, a partial change in the tertiary and secondary structure is observed. The data obtained will allow a better understanding of the principles of the interaction of immune cells with antibodies and SeNPs and, in the future, may serve to create a new generation of immunomodulators.

Investigation of Aggregation and Disaggregation of Self-Assembling Nano-Sized Clusters Consisting of Individual Iron Oxide Nanoparticles upon Interaction with HEWL Protein Molecules.

In this paper, iron oxide nanoparticles coated with trisodium citrate were obtained. Nanoparticles self-assembling stable clusters were ~10 and 50-80 nm in size, consisting of NPs 3 nm in size. The stability was controlled by using multi-angle dynamic light scattering and the zeta potential, which was -32 ± 2 mV. Clusters from TSC-IONPs can be destroyed when interacting with a hen egg-white lysozyme. After the destruction of the nanoparticles and proteins, aggregates are formed quickly, within 5-10 min. Their sizes depend on the concentration of the lysozyme and nanoparticles and can reach micron sizes. It is shown that individual protein molecules can be isolated from the formed aggregates under shaking. Such aggregation was observed by several methods: multi-angle dynamic light scattering, optical absorption, fluorescence spectroscopy, TEM, and optical microscopy. It is important to note that the concentrations of NPs at which the protein aggregation took place were also toxic to cells. There was a sharp decrease in the survival of mouse fibroblasts (Fe concentration ~75-100 µM), while the ratio of apoptotic to all dead cells increased. Additionally, at low concentrations of NPs, an increase in cell size was observed.