Constructive Empathizing – Educational Competence in the Light of Child’s Play

Pre-school play is an important moment in a child’s development.  Caretakers play an important role during play. This article presents a structure of educational competence called “constructive empathizing” or “invitingaccompanying competence”. A caretaker’s special task during child’s play is conversion between the third-person knowledge and a subjective, autotelic interaction. The caretaker’s constructive empathizing during a child’s play determines the child’s social and moral development, it helps the child to turn play into work, as a sign of higher spontaneous understanding of cultural norms standing behind cultural correctness actions. Finally, it is evidence of dynamic interaction between abstract-concrete thinking, with the child learning from the caretaker

Many facets of CPEB proteins in neurons and beyond: expression, mRNA recognition and phosphorylation

CPEBs are a family of evolutionary conserved auxiliary translational factors. They bind to CPEs in the 3’UTR of target mRNA and, by interacting with other members of the translational machinery, promote or decrease gene expression. Since the first description of CPEB protein as a key orchestrator of oocyte maturation in Xenopus Laevis, key functions of CPEBs in embryonic development, cellular senescence and synaptic plasticity have been described. Local, postsynaptic modulation of synaptic efficacy is the most studied function of CPEBs in the CNS. However, the expression of CPEBs is not restricted to neurons and even 7% of total brain mRNAs possesses CPEs. Moreover, specific mRNAs are enriched in functionally distinct subcellular regions, like dendritic spines of neurons or complex cellular processes of astrocytes, where they might be translationally regulated. Herein, I embarked on further elucidating the role of CPEB translational regulators family in selected cell populations in mouse brain. In the first part of the work, the target mRNA specificity of different CPEBs was tested. I argued with the hypothesis that CPEBs -3 and -4 require mRNA secondary structure (a stem-loop) for target recognition. I showed that despite differences in the RBD primary structure, CPEB-1 shows a considerable sequence specificity overlap with CPEB-3 (and possibly, due to sequence similarity, with CPEBs -2 and -4 as well). This holds true for neuronal (CaMKIIα) and astrocytic(β-catenin) CPEB targets. Additionally, RNAco-immunoprecipitation revealed two novel CPEB target mRNAs, encoding (1) an astrocytic gap junction protein (Connexin43) and (2) CPEB-3 protein itself. In the first part of the work, the target mRNA specificity of different CPEBs was tested. I argued with the hypothesis that CPEBs -3 and -4 require mRNA secondary structure (a stem-loop) for target recognition. I showed that despite differences in the RBD primary structure, CPEB-1 shows a considerable sequence specificity overlap with CPEB-3 (and possibly, due to sequence similarity, with CPEBs -2 and -4 as well). This holds true for neuronal (CaMKIIα) and astrocytic(β-catenin) CPEB targets. Additionally, RNAco-immunoprecipitation revealed two novel CPEB target mRNAs, encoding (1) an astrocytic gap junction protein (Connexin43) and (2) CPEB-3 protein itself. In the second part of the thesis, CPEBs 1-4 gene expression in microglia, NG2 cells and astrocytes was assessed at the mRNA and protein level, confirming the ubiquitous nature of the CPEB expression. A more detailed analysis of CPEB-3 mRNA revealed expression of only certain isoforms of the protein in ESdM cells. Each CPEB family member occurs in several splice variants. Hitherto, only a handful of reports are dealing with the significance of alternative splicing of CPEBs. Interestingly, regulation of expression of CPEBs in neurons may happen in an isoform-specific fashion. Seeing this as a highly intriguing phenomenon, I focused on CPEB-3, containing a putative kinase recognition site on the alternatively spliced exon 5. In a cell-free systems and in cultured cells, I showed that CPEB-3 is a target of PKA and CaMKII – both critical for LTP/LTD, and that exon 5 harbors a consensus sequence required for kinase recognition. In vivo, in a transgenic mouse overexpressing CPEB3 protein in principal neurons of the hippocampus, I founda significant decrease in GluR2 AMPA-R subunit protein levels. In view of the above, the stimulation-induced phosphorylation of CPEB-3 may very well be a novel mechanism of translational modulation of synaptic plasticity. In the final part of the thesis, I aimed at establishing a quality control method for Cre-mediated gene deletions. Using Cx30-/-;Cx43fl/fl mice with hGFAP-Cre recombinase driver, I developed a reliable immunoblot-based tool for post-hoc control of recombination variability in hGFAP-Cre transgenic mice

A NaV1.8FlpO mouse enabling selective intersectional targeting of low threshold C fiber mechanoreceptors and nociceptors

Genetic targeting of select populations of cells in the mouse nervous system is often hampered by a lack of selectivity, as candidate genes for such targeting are commonly expressed by multiple cell populations, also in the same region. Intersectional targeting using two or more genes has been enabled by the development of reporter tools dependent on more than one recombinase or gene regulator. Still, widespread adoption of intersectional tools is complicated by a scarcity of driver mice expressing recombinases other than Cre. Here we report the generation and characterization of a new driver mouse that expresses the FlpO recombinase from the endogenous locus of the Scn10a gene encoding NaV1.8, a voltage-gated sodium channel that is almost exclusively expressed in the afferent limb of the peripheral nervous system. Moreover, among sensory neurons the channel is preferentially expressed in nociceptors and in low-threshold C-fiber mechanoreceptors (C-LTMRs). The mouse showed high recombination efficiency (97%) and selectivity (93%) in dorsal root ganglia. Reporter-expressing fibers were observed in a variety of peripheral tissues, including skin, skeletal muscle, genitalia, bladder and intestines. To validate the suitability of the FlpO mouse line for intersectional targeting, we crossed it with a mouse line expressing CreERT2 from the Th (tyrosine hydroxylase) locus. This approach resulted in strikingly selective and efficient targeting of C-LTMRs, showing robust visualization of nerve endings of these fibers in skin and spinal cord at the light and electron microscopic level. Thus, the NaV1.8Flpo mouse line presented here constitutes a selective and versatile tool for intersectional genetic targeting of NaV1.8 expressing primary afferent neurons

Distinct translatome changes in specific neural populations precede electroencephalographic changes in prion-infected mice

Selective vulnerability is an enigmatic feature of neurodegenerative diseases (NDs), whereby a widely expressed protein causes lesions in specific cell types and brain regions. Using the RiboTag method in mice, translational responses of five neural subtypes to acquired prion disease (PrD) were measured. Pre-onset and disease onset timepoints were chosen based on longitudinal electroencephalography (EEG) that revealed a gradual increase in theta power between 10- and 18-weeks after prion injection, resembling a clinical feature of human PrD. At disease onset, marked by significantly increased theta power and histopathological lesions, mice had pronounced translatome changes in all five cell types despite appearing normal. Remarkably, at a pre-onset stage, prior to EEG and neuropathological changes, we found that 1) translatomes of astrocytes indicated reduced synthesis of ribosomal and mitochondrial components, 2) glutamatergic neurons showed increased expression of cytoskeletal genes, and 3) GABAergic neurons revealed reduced expression of circadian rhythm genes. These data demonstrate that early translatome responses to neurodegeneration emerge prior to conventional markers of disease and are cell type-specific. Therapeutic strategies may need to target multiple pathways in specific populations of cells, early in disease

Anti-prion drugs do not improve survival in novel knock-in models of inherited prion disease.

Prion diseases uniquely manifest in three distinct forms: inherited, sporadic, and infectious. Wild-type prions are responsible for the sporadic and infectious versions, while mutant prions cause inherited variants like fatal familial insomnia (FFI) and familial Creutzfeldt-Jakob disease (fCJD). Although some drugs can prolong prion incubation times up to four-fold in rodent models of infectious prion diseases, no effective treatments for FFI and fCJD have been found. In this study, we evaluated the efficacy of various anti-prion drugs on newly-developed knock-in mouse models for FFI and fCJD. These models express bank vole prion protein (PrP) with the pathogenic D178N and E200K mutations. We applied various drug regimens known to be highly effective against wild-type prions in vivo as well as a brain-penetrant compound that inhibits mutant PrPSc propagation in vitro. None of the regimens tested (Anle138b, IND24, Anle138b + IND24, cellulose ether, and PSCMA) significantly extended disease-free survival or prevented mutant PrPSc accumulation in either knock-in mouse model, despite their ability to induce strain adaptation of mutant prions. Our results show that anti-prion drugs originally developed to treat infectious prion diseases do not necessarily work for inherited prion diseases, and that the recombinant sPMCA is not a reliable platform for identifying compounds that target mutant prions. This work underscores the need to develop therapies and validate screening assays specifically for mutant prions, as well as anti-prion strategies that are not strain-dependent

Analysis of seroconversion following COVID-19 vaccination among multiple sclerosis patients treated with disease-modifying therapies in Poland

Clinical rationale for the study. The rapid spread of SARS-CoV-2 throughout the world has highlighted the importance of vaccinations to control the pandemic and to protect people at risk for severe disease courses. Disease-modifying therapies (DMT) in multiple sclerosis (MS), whether immunomodulatory or immunosuppressive, may affect the immune response. Therefore, the question arose as to whether these vaccinations would be effective.Aim of the study. We planned a study to assess the immune response to SARS-CoV-2 vaccines by type of therapy.Material and methods. Participants were recruited from 14 Polish MS centres. The data was obtained by neurologists using a questionnaire. We collected data on 353 MS patients (269 females, 84 males) who received complete primary SARS-CoV-2 vaccination. All persons with MS (PwMS) were treated with disease-modifying therapies.Results. 305 out of 353 PwMS (86.4%) were positive for IgG Abs against SARS-CoV-2 S domain S1 Ag after vaccination. A strong immune response was noted in 129 PwMS (36.5%). The rate of seroconversion after SARS-CoV-2 vaccination in PwMS who received immunomodulatory DMTs (interferon beta, glatiramer acetate, teriflunomide, dimethyl fumarate, natalizumab) was 91.5%, in PwMS receiving immune reconstruction therapy (alemtuzumab, cladribine) was 92%, and in immunosuppressive DMTs (fingolimod, ocrelizumab), the seroconversion rate was 59%.Conclusions and clinical implications. Our study shows that, in PwMS receiving immunomodulatory therapy, the immune response to vaccination is generally excellent. Even in immunosuppressive patients, seroconversion is satisfactory

Constructive Empathizing – Educational Competence in the Light of Child’s Play

Pre-school play is an important moment in a child’s development.  Caretakers play an important role during play. This article presents a structure of educational competence called “constructive empathizing” or “invitingaccompanying competence”. A caretaker’s special task during child’s play is conversion between the third-person knowledge and a subjective, autotelic interaction. The caretaker’s constructive empathizing during a child’s play determines the child’s social and moral development, it helps the child to turn play into work, as a sign of higher spontaneous understanding of cultural norms standing behind cultural correctness actions. Finally, it is evidence of dynamic interaction between abstract-concrete thinking, with the child learning from the caretaker.</jats:p

Instrumental Value of Knowledge and Knowledge-creation

John Dewey’s pragmatic theory of knowledge supports the assertion that both the instrumental value of knowledge and knowledge-creation have common philosophical underpinnings. An analysis of this issue can help bridge the gap between the typical economic approach to knowledge and those approaches that are represented by contemporary epistemology and philosophy of science. The universal concept of the value of knowledge that is proposed here allows one to carry out further research on using contemporary epistemology in knowledge management practice.3249643Studia Metodologiczn