The use of efficient and inexpensive substrates (2D matrices) for cultivation and differentiation of nerve cells in vitro is important for the creation of tissue engineering constructs intended for the treatment of nervous system pathologies. Recombinant analogues of the orb-weaver spider dragline-silk proteins spidroins 1 and 2 appear promising in addressing this task. The aim of the study was to evaluate the effect of cell substrates derived from mixtures of recombinant spidroins (RS) rS1/9 and rS2/12 with hybrid proteins (HP) containing rS1/9 monomer fused with biologically active peptides on gene expression levels of key synapse-specific proteins and viability of the human neuroblastoma SH-SY5Y cell line during directed cholinergic differentiation. A two-stage scheme of directed cholinergic differentiation of SH-SY5Y cells using retinoic acid and brain-derived neurotrophic factor (BDNF) was implemented. Cell viability was assessed via MTT assay and crystal violet staining. The mRNA levels of the studied genes were assessed by real-time PCR. Directed differentiation of the SH-SY5Y cells was marked by a significant increase in the gene expression levels of synaptophysin, synapsins I and II, and the postsynaptic protein PSD-95. The highest cell viability and increased PSD-95 expression levels were observed during differentiation on a matrix consisting of RS rS1/9 and rS2/12 mixed with the RGDS peptide (present in extracellular matrix proteins) and heparin-binding peptide (HBP, laminin fragment) containing HPs. The highest efficiency during the differentiation of the SH-SY5Y cells was demonstrated by a matrix consisting of the mixture of RS rS1/9 and rS2/12 and a HP made up by RS rS1/9 monomer fused with RGDS (the ligand of integrins) and HBP (the ligand of growth factors and syndecans). Matrices consisting of RS rS2/12 alone or the mixture of rS2/12 with HP(RGDS) showed lower efficiency, although the use of the GRGGL peptide (which interacts with the neural cell adhesion molecules and is a component of RS rS1/9) led to an increase in efficiency.

Induced pluripotent stem cells (iPSCs) are analogous to embryonic stem cells in their properties, and might be derived from adult somatic cells. There exist two main approaches to practical application of iPSCs technologies they are regenerative medicine and human diseases modeling. Nowadays the process of iPSCs obtaining is being investigated in many world laboratories, but there are no standards in the iPSC obtaining technology. This work was aimed to develop a simple, cheap and practical protocol for obtaining iPSCs and to establish a collection of iPCS lines derived from cells of patients with a Parkinson disease to make a cell model of this disease. The work resulted in a development of the protocol to produce iPSCs based on a lentiviral delivery of transgenes into cells. iPCS lines from 3 patients with inherited Parkinson disease forms have been obtained.

Depression is a complex, heterogeneous and multifactorial disease with poorly understood pathophysiological mechanisms. Outbred Sprague Dawley (SD) and Wistar Han (WH) rats are widely used in biomedical research, in particular, in the model of chronic unpredictable stress (CUS) and in the inflammatory model of depression. However, the differences between SD and WH in these experimental models are not well studied. The aim of the present study was to compare alterations in the hedonic status, food consumption and body weight of male SD and WH rats obtained from Charles River Laboratories (Germany), in the CUS model (7 weeks) and in acute low-dose endotoxemia (single intraperitoneal injection of a low subseptic dose of LPS, 25 mu g/kg). Hedonic status was assessed using the sucrose preference test without prior food and water deprivation. Significant differences were found between SD and WH in the body weight gain, food consumption, locomotor and exploratory activity, and in the mass of the thymus. WH rats demonstrated a lower threshold of sucrose preference (0.3%) compared to SD rats (0.4%). The effects of both CUS and endotoxemia on body weight were more pronounced in SD rats. In endotoxemia, a decrease in hedonic status was more pronounced in WH rats. LPS-induced serum levels of TNF-alpha were higher in SD, and corticosterone was higher in WH (90 min after LPS). In the CUS model, changes in hedonic status and adrenal hypertrophy were detected only in SD rats, but thymic involution was detected only in WH. Significant differences in the physiological characteristics of Sprague Dawley and Wistar Han rats and the different sensitivity of their metabolic and hedonic status to stress and inflammation can have a significant impact on the results obtained in the models of the corresponding pathologies, as well as on their interpretation.

The technology for producing human induced pluripotent stem cells (iPSCs) and the possibility of directed differentiation into specialized cells of all body tissues have opened unique opportunities for studying the molecular genetic basis of the pathogenesis of neurodegenerative diseases in vitro and effective screening for compounds with neuroprotective activity. The aim of this work was to obtain glial cell cultures from iPSCs of a healthy donor and a patient with the familial form of Parkinson's disease (G2019S mutation in the LRRK2) and to characterize them. At the first stage, we compared the three previously described protocols for the differentiation of glial cells from neural precursors of human iPSCs and selected the method most acceptable under our conditions in terms of the quality and time necessary for obtaining the desired cultures. Glial cell cultures obtained by this method were characterized by the levels of expression of a number of neuroglial differentiation genes. Also, in the resulting cultures, we analyzed the expression of genes of some neurotrophic factors (GDNF, BDNF, NGF, NT3). It was shown that the culture medium conditioned by glial cells from a patient with Parkinson's disease had a negative effect on the growth of neurites of dopaminergic neurons in differentiated cultures of a healthy donor, decreasing their length by a factor of 2.