As a result of the implementation of the project, we expect to achieve breakthrough research results determining a significant scientific groundwork for the Russian researchers in the field of the development of unique materials that, by virtue of direct physical impacts or their combination or a combination of those with targeted drug delivery, can be used for the control of the growth and differentiation of various types of cells;
The obtained fundamental knowledge will, at the next stage, allow to use these effects for the treatment of diseases that require the recovery of nervous and smooth muscle tissue and the inhibition of tumour development.
As a result, we will produce new hybrid piezoactive materials with enhance electrophysical and physico-mechanical characteristics for stem cell differentiation control and/or somatic cell transdifferentiation (for instance, into nerve or soft muscle cells) using physical effects (piezoelectric charge, ultrasound impact, temperature) generated by developed materials.
The impact of an external magnetic field on piezoelectric hybrid materials in the form of nano-particles and scaffolds will be studied. We will also identify the laws of the impact of the surface charge and polarity on cell differentiation and the delivery of medications to cells in in vitro experiments;
We will conduct a genome-wide study of the influence of physical impact on the gene expression profile, which will allow not only to describe the effects observed in biochemical systems in detail but also to determine the mechanisms of those effects.
We will propose new types of materials and structures for targeted drug delivery to cells, organs, and tissues of the organism as well as for controlled dosage drug delivery.
It is expected that the laboratory will patent the developed technologies as well as register know-hows not only for the methods of their production but also for means of targeted drug delivery and controlled release of pharmaceuticals with parameters exceeding the known counterparts.
The creation of the laboratory will aid the formation of a cluster that is currently non-existent in the Siberian Federal District whose main objective is conducting world-class studies in the domain of the development of piezo- and magnetoelectric materials for biomedical applications.
Scientific results:
Biocompatible magnetoelectric (ME) core-shell nanoparticles (NPs) based on manganese ferrite (MFO) and perovskite (BCZT) were obtained for the first time using the microwave hydrothermal method. Compared with the classical hydrothermal method, the use of microwave radiation allows to significantly reduce the growth time of magnetic cores and the perovskite shell on their surface. It was found that in-situ functionalization of the surface of MFO cores leads to the formation of a covalent bond between the oxygen atoms of the carboxyl groups of citric acid and the Fe and Mn atoms of manganese ferrite. Epitaxial formation of the BCZT shell on the surface of the MFO cores was revealed. A correlation of the crystal structure parameters is observed for these compounds: MFO ((220) – 3.08 Å, (400) – 2.12 Å) and BCZT ((110) – 2.99 Å, (204) – 2.14 Å). Formation of the BCZT shell with a thickness of 10-20 nm leads to a decrease in the saturation magnetization and an increase in the coercive force of the MFO cores with a diameter of 43 nm from (41.40±1.20) emu/g to (6.10±0.20) emu/g and from (46±3) Oe to (69±5) Oe, respectively. The efficiency of decomposition of the model pollutant RhB of 95% under the influence of a magnetic field (150 mT, 100 Hz) for 2.5 h was achieved due to the catalytic activity of the developed ME NPs added to the solution at a concentration of 4 mg/ml. The absence of a cytotoxic effect of ME NPs at their concentration up to 6 mg/ml on stem cells in vitro was shown. Magnetic nanoparticles (MNPs) MFO and MFO@BCZT do not have a cytotoxic effect on normal human fibroblasts. MFO nanoparticles induce apoptosis in tumor cells and suppress their viability. At the same time, MFO@BCZT core-shell structure MNPs suppress metabolic activity but do not cause cell death by necrosis or apoptosis.
The results of the study of the interaction of magnetic and magnetoelectric nanoparticles based on manganese ferrite, scaffolds based on biodegradable PLLA and non-biodegradable PVDF-TrFE polymers with cells showed that the obtained magnetoactive nanomaterials demonstrate high biocompatibility with respect to normal human cells and allow controlling the functional activity of cells when an external magnetic field is applied. The ability to influence the proliferation and differentiation of human cells adsorbed on the surface of magnetic matrices when applying external magnetic fields, demonstrated as a result of the study, determines high prospects for using these materials to solve the most important problems in the field of regenerative medicine. The selectivity of the cytotoxic effect on cancer cells observed for magnetoelectric nanoparticles with a core-shell structure and a BCZT coating, combined with the established fact of the presence of an operating range of the long-range effect of a constant magnetic field, will allow us to offer promising solutions for personalizing anti-cancer therapy.
It is shown that, in comparison with magnetic NPs, the application of AMF (7-50 mT) leads to a decrease in the viability of cancer cells incubated with ME NPs, which is due to their ME properties. AMF (100 mT) leads to an increase in the cytotoxic effect of the studied ME nanoparticles. It was found that incubation with magnetic and ME NPs leads to an increase in the ROS content in MDA-MB231 and U87 cells for 8 hours. It is shown that the formation of intracellular ROS in U87 cells does not increase with the application of AMF in the presence of magnetic nanoparticles, whereas ME nanoparticles in combination with the application of AMF (100 mT, 100 Hz) increase the production of ROS in U87 cells. It was also found that long-term storage of nanoparticles (in solution and as powders) can negatively affect their biological properties due to deterioration of their stability. In this regard, it is recommended to use freshly prepared nanoparticles for conducting biological experiments.
The effect of an alternating magnetic field on cell growth in vitro was established when studying nanoparticles with the core-shell structure MFO@BCZT. Together with the low toxicity of MFO@BCZT, this allowed the use of these particles in experiments on neurons ex vivo and in vivo. Ex vivo on vital hippocampal slices, it was possible to demonstrate the specific ability of MFO@BCZT to affect the functional activity of cells in the presence of an alternating magnetic field, which is specific to MFO. It was also shown that MFO@BCZT can be captured in vivo by mouse olfactory neurons and move from the nasal cavity to the brain mainly inside neurons, while preserving their shell. Thus, there is a possibility of using MFO@BCZT in vivo for deep brain stimulation of mammals. Human monocytes and fibroblasts adhere to the fibers of magnetically active PLLA-Fe3O4 and PVDF-TrFE-Fe3O4 scaffolds, retaining normal morphology and functional activity. The model of human peripheral blood monocytes demonstrated the possibility of their differentiation into both M1 and M2 macrophages. The effect of AMF on monocyte-macrophage cells attached to scaffolds leads to activation of their polarization without adding chemical stimuli. In the case of normal human fibroblasts, their adhesion to scaffolds affects biological processes involved in tissue regeneration, and the use of AMF triggers a number of molecular events through the activation of G-protein-coupled receptors and ion channels. This indicates the possibility of modulating their behavior in the body and controlling them in the future using MP. The obtained data prove that stimulation of AMF is an ideal biocompatible signal, on the one hand, it is favorable for differentiation of stem cells and, on the other hand, can be successfully used for remote control of the properties of their microenvironment.
A change in cytokine expression was detected in fibroblasts adhered to scaffolds. An increase in the expression of the CXCL5 and CXCL8 genes, the products of which promote the infiltration and activation of neutrophils, indicates that fibroblasts in this state rather favor inflammation and hinder the regeneration process. In turn, exposure to AMF leads to a further increase in the expression of the IL24 gene, which is known to coordinate the proangiogenic program of repair and proliferation to restore tissue integrity and homeostasis, thus promoting regeneration.
Transcriptomic data showed that adhesion of normal human fibroblasts to scaffolds activated pathways associated with important biological processes such as coagulation and wound healing, which are the first reactions in the cascade of events that restore damaged tissue after injury, as well as key signaling pathways regulating the regeneration process, such as Wnt, BMP and TGFβ. Exposure of cells on scaffolds to AMF can trigger extracellular (via G protein-coupled receptors) and intracellular (via calcium ion channel activation) signaling pathways, the obtained experimental data allow us to consider hybrid magnetically active scaffolds as promising materials for solving problems of regenerative medicine.
Methods for culturing stem cells on the surface of scaffolds, as well as samples impregnated with 20 wt. % magnetite nanoparticles coated with citric acid, have been developed. Methods of cell stimulation using a magnetic field have been developed. A method of targeted neural differentiation has been developed and a primary assessment of the degree of cell differentiation on the scaffold surface in the presence of a magnetic field has been carried out. The effect of physical effects (magnetic field) on the viability of the cell population on the scaffold surface has been studied. The biocompatibility of hybrid scaffolds has been established in in vitro experiments on multipotent mesenchymal stromal cells. The effect of a magnetic field on the proliferative and differentiating activity of multipotent mesenchymal stromal cells has been established.
Implementation of research results:
The research results have been implemented into the educational process within the framework of the discipline Materials and coatings for biomedical purposes for the training of master's students in the field of study 18.04.01 - Chemical technologies.
Organizational and infrastructural changes:
The International Research Center “Piezo- and Magnetoelectric Materials” is a structural division of the research school of chemical and biomedical technologies.
Education and personnel occupational retraining:
Retraining of personnel participating in project 11. As a result of the project implementation, the implementers defended 3 dissertations for the degree of candidate of sciences, as well as 1 dissertation for the degree of doctor of sciences.
Cooperation:
- Ural Federal University (Yekaterinburg).
- Siberian Federal University (Krasnoyarsk).
- Institute of Cytology and Genetics SB RAS (Novosibirsk).
- Siberian State Medical University (Tomsk).
- Lomonosov Moscow State University (Moscow).
- Institute of Chemical Biology and Fundamental Medicine SB RAS (Novosibirsk).
