Lyudmila Novosadova

Background: Neuroinflammation and neurodegeneration are closely linked in Parkinson’s disease (PD). Specialized pro-resolving mediators (SPMs) are of interest as potential therapeutic agents for PD because they promote the resolution of inflammation. Methods: We investigated whether PARK2 mutations are associated with cell-type-specific patterns of SPM receptor expression by comparing neuronal and glial cultures derived from induced pluripotent stem cells (iPSCs) of PD patients carrying different PARK2 mutations with corresponding cultures from healthy donors (HDs), using real-time PCR and Western blotting. Results: The resolvin E1 receptors, BLT1 and CHEMR23, and the resolvin D1 (RvD1) receptors, GPR32 and FPR2, were detected at both the mRNA and protein levels in PARK2-mutant neurons and glia. Compared with HD cultures, mutant neurons had significantly higher CHEMR23 protein levels despite similar CHEMR23 mRNA levels, whereas no statistically detectable differences in the expression of any of the four receptors were found in glia. Receptor profiles also appeared to differ between cells carrying PARK2 mutations and those carrying mutations in other genes (GBA1, LRRK2). PARK2-mutant neurons showed lower levels of several inflammatory markers than HD neurons, in contrast to the pro-inflammatory profile previously reported in PARK2-mutant glia. A pilot experiment with glial cultures derived from one cell line per group suggested that in the presence of TNFα, RvD1 may reduce IL-6 secretion and increase cellular metabolic activity in PARK2-mutant glia, with responses that appeared to differ from those observed in HD glia. Discussion: The presence of resolvin receptors in PARK2-mutant neurons and glia, together with elevated CHEMR23 protein levels in mutant neurons compared with HD neurons, provides a basis for further investigation of resolvin signaling in PARK2-associated PD. However, the relationship between receptor expression and functional responses to resolvins remains to be established. The contrasting inflammatory profiles of neurons and glia suggest that PARK2-mutant glial cultures may be particularly relevant for investigating the anti-inflammatory effects of resolvins. Conclusions: Cell-type- and genetic-background-associated differences in receptor expression should be considered when developing resolvin-based therapeutic approaches for PD. Further studies are needed to assess the potential anti-inflammatory effects of RvD1 on glial cells in genetic forms of the disease.

It was shown earlier that induced pluripotent stem cells (iPSCs) differentiate into a heterogeneous cell population enriched in dopaminergic neurons on a m

Neuroinflammation is considered to be one of the driving factors in Parkinson’s disease (PD). This study was conducted using neuronal and glial cell cultures differentiated from induced pluripotent stem cells (iPSC) of healthy donors (HD) and PD patients with different PARK2 mutations (PD). Based on the results of RNA sequencing, qPCR and ELISA, we revealed transcriptional and post-transcriptional changes in HD and PD neurons cultivated in HD and PD glial-conditioned medium. We demonstrated that if one or both of the components of the system, neurons or glia, is Parkin-deficient, the interaction resulted in the down-regulation of a number of key genes related to inflammatory intracellular pathways and negative regulation of apoptosis in neurons, which might be neuroprotective. In PD neurons, the stress-induced up-regulation of APLNR was significantly stronger compared to HD neurons and was diminished by glial soluble factors, both HD and PD. PD neurons in PD glial conditioned medium increased APLN expression and also up-regulated apelin synthesis and release into intracellular fluid, which represented another compensatory action. Overall, the reported results indicate that neuronal self-defense mechanisms contribute to cell survival, which might be characteristic of PD patients with Parkin-deficiency.

Introduction. It was previously shown that induced pluripotent stem cells (iPSCs) differentiate into a heterogeneous cell population enriched in dopaminergic neurons on a microgel matrix based on recombinant spidroin rS1/9. To further study the functional activity of the resulting neuronal culture, it was used to administer it to mice with a toxic model of Parkinson’s disease (PD) in order to assess the possibility of modulating motor behavior impaired by the introduction of MPTP(1-methyl-4-phenyl-1.2,3.6-tetrahydropyridine). Material and methods. Mice with a toxic model of early symptomatic PD were used for stereotaxic transplantation of neuronal cells. Motor behavior was assessed using the open field test. Results. It has been shown that in vitro cultivation of neuronal cells on a microgel-free medium has a pronounced effect on the motor activity of mice 2 weeks after their introduction into the mouse brain and 3 weeks after toxic damage to neurons of the substantia nigra. The effect of short-term 3-day cultivation turned out to be negative — the motor activity of mice decreased. With longer cultivation, on the contrary, it increases — which is manifested primarily in an increase in the duration of the active period and the distance traveled. This is especially clearly manifested when comparing the motor activity of mice that were injected with neuronal cells cultured on a microgel-free medium for 2 weeks or 3 days. These effects are modulated by cell cultivation on a microgel — and this effect is different for neuronal cultures with different periods of in vitro cultivation. Conclusion. Neuronal cell cultures derived from iPSCs and differentiated in vitro can increase motor activity when stereotaxically introduced into the brain of mice with a toxic model of PD. This effect depends on the duration of cell culture in vitro and on the matrices used in the cultivation process.

Parkinson’s disease (PD) is the most serious movement disorder, but the actual cause of this disease is still unknown. Induced pluripotent stem cell-derived neural cultures from PD patients carry the potential for experimental modeling of underlying molecular events. We analyzed the RNA-seq data of iPSC-derived neural precursor cells (NPCs) and terminally differentiated neurons (TDNs) from healthy donors (HD) and PD patients with mutations in PARK2 published previously. The high level of transcription of HOX family protein-coding genes and lncRNA transcribed from the HOX clusters was revealed in the neural cultures from PD patients, while in HD NPCs and TDNs, the majority of these genes were not expressed or slightly transcribed. The results of this analysis were generally confirmed by qPCR. The HOX paralogs in the 3′ clusters were activated more strongly than the genes of the 5′ cluster. The abnormal activation of the HOX gene program upon neuronal differentiation in the cells of PD patients raises the possibility that the abnormal expression of these key regulators of neuronal development impacts PD pathology. Further research is needed to investigate this hypothesis.

The production and transplantation of functionally active human neurons is a promising approach to cell therapy. Biocompatible and biodegradable matrices that effectively promote the growth and directed differentiation of neural precursor cells (NPCs) into the desired neuronal types are very important. The aim of this study was to evaluate the suitability of novel composite coatings (CCs) containing recombinant spidroins (RSs) rS1/9 and rS2/12 in combination with recombinant fused proteins (FP) carrying bioactive motifs (BAP) of the extracellular matrix (ECM) proteins for the growth of NPCs derived from human induced pluripotent stem cells (iPSC) and their differentiation into neurons. NPCs were produced by the directed differentiation of human iPSCs. The growth and differentiation of NPCs cultured on different CC variants were compared with a Matrigel (MG) coating using qPCR analysis, immunocytochemical staining, and ELISA. An investigation revealed that the use of CCs consisting of a mixture of two RSs and FPs with different peptide motifs of ECMs increased the efficiency of obtaining neurons differentiated from iPSCs compared to Matrigel. CC consisting of two RSs and FPs with Arg–Gly–Asp–Ser (RGDS) and heparin binding peptide (HBP) is the most effective for the support of NPCs and their neuronal differentiation.

Parkinson’s disease (PD) is the second most common neurodegenerative diseases characterized by progressive loss of midbrain dopaminergic neurons in the substantia nigra. Mutations in the PARK2 gene are a frequent cause of familial forms of PD. Sustained chronic neuroinflammation in the central nervous system makes a significant contribution to neurodegeneration events. In response to inflammatory factors produced by activated microglia, astrocytes change their transcriptional programs and secretion profiles, thus acting as immunocompetent cells. Here, we investigated iPSC-derived glial cell cultures obtained from healthy donors (HD) and from PD patients with PARK2 mutations in resting state and upon stimulation by TNFα. The non-stimulated glia of PD patients demonstrated higher IL1B and IL6 expression levels and increased IL6 protein synthesis, while BDNF and GDNF expression was down-regulated when compared to that of the glial cells of HDs. In the presence of TNFα, all of the glial cultures displayed a multiplied expression of genes encoding inflammatory cytokines: TNFA, IL1B, and IL6, as well as IL6 protein synthesis, although PD glia responded to TNFα stimulation less strongly than HD glia. Our results demonstrated a pro-inflammatory shift, a suppression of the neuroprotective gene program, and some depletion of reactivity to TNFα in PARK2-deficient glia compared to glial cells of HDs.

Oxidative stress (OS) is implicated in the pathogenesis of several neurodegenerative diseases. We have previously shown that N-acyl dopamines (N-ADA and N-DDA) protect the neural cells of healthy donors and patients with Parkinson’s disease from OS. In this study, we assessed the effects of N-acyl dopamines on the expression of neurotrophic factors in human-induced pluripotent stem cell-derived neuronal cultures enriched with dopaminergic neurons under conditions of OS induced by hydrogen peroxide. We showed that hydrogen peroxide treatment increased BDNF but not GDNF mRNA levels, while it did not affect the secretion of corresponding proteins into the culture medium of these cells. Application of N-acyl dopamines promoted BDNF release into the culture medium. Under conditions of OS, N-DDA also increased TRKB, TRKC and RET mRNA levels. Furthermore, N-acyl dopamines prevented cell death 24 h after OS induction and promoted the expression of antioxidant enzymes GPX1, GPX7, SOD1, SOD2 and CAT, as well as reduced the BAX/BCL2 mRNA ratio. These findings indicate that stimulation of the expression of neurotrophic factors and their receptors may underlie the neuroprotective effects of N-acyl dopamines in human neurons.