Astrocyte-derived soluble factors promoting neuronal differentiation of adult neural progenitor cells

Adult neurogenesis occurs in two regions of the brain: the subventricular zone of lateral ventricles and the subgranular zone of hippocampus. Neural progenitor cells found in these neurogenic regions are capable of self-renewing and generating mature neurons and glia. It is known that reciprocal interactions with the local environment can regulate cellular maintenance and plasticity of neural progenitor cells. Astrocytes in the microenvironment surrounding neural progenitor cells have a profound influence on a variety of inter- and intra-cellular processes during adult neurogenesis. Thus, we have investigated the roles of astrocyte-derived factors which influence neural progenitor cell differentiation in vitro. The neural progenitor cells used in our studies were hippocampal progenitor cells isolated from adult rat hippocampus (a gift from F. Gage, Salk Institute, La Jolla, CA). Previous immunocytochemical results showed that co-culture with enriched neonatal astrocytes significantly and selectively increased neuronal differentiation of adult hippocampal progenitor cells. We proposed that the astrocytes present discrete modulators to the overlying adult hippocampal progenitor cells involving contact-mediated or release of soluble factors, or a combination of both. From the first investigation in this dissertation, we found that astrocyte-derived soluble factors specifically promote neuronal differentiation of neural progenitor cells. In the second investigation, we tested a hypothesis that a candidate neurogenic factor produced from astrocytes, interleukin-6, is a key regulator that stimulates functional differentiation of neural progenitor cells to a neuronal fate. Using immunocytochemical analysis and whole-cell recording, we have demonstrated that astrocyte-derived interleukin-6 enhances neuronal differentiation of AHPCs by presenting three main observations: (a) neuronal marker expression was increased, (b) the average length of neurites from neuronal-restricted AHPCs was increased and (c) voltage-gated inward current density was increased with no significant differences in voltage-gated outward current density, apparent resting membrane potential, or cell capacitance. In the third investigation, we examined the influence of extracellular calcium and voltage-gated calcium channel activity on interleukin-6-mediated neuronal differentiation of neural progenitor cells. We observed that interleukin-6-enhanced neuronal differentiation was reduced when cultured in low calcium-containing culture medium or with L-type voltage-gated calcium channel antagonists. Interleukin-6 treatment also increased the fraction of neural progenitor cells immunoreactive for a neuronal marker and for cAMP response element binding protein. Overall, we conclude that astrocyte-derived soluble factors promote neuronal differentiation of adult neural progenitor cells, and astrocyte-derived interleukin-6 is a neurogenic factor able to induce neural progenitor cells to differentiate into neurons. These findings may provide important insights into mechanisms for controlling neural progenitor cell differentiation and facilitate development of cell-based therapeutic strategies using adult neural progenitor cells

Soluble factors from neocortical astrocytes enhance neuronal differentiation of neural progenitor cells from adult rat hippocampus on micropatterned polymer substrates

Rat adult hippocampal progenitor cells (AHPCs) are self-renewing, multipotent neural progenitors that have the ability to differentiate into neurons and glia. Previously, we demonstrated that coculture of AHPCs with postnatal day two, type 1 cortical astrocytes on laminin-coated micropatterned polymer substrates facilitates selective neuronal differentiation of the AHPCs 1. Under this condition, multi-dimensional cell-cell and/or cell-extracellular matrix interactions, as well as possible soluble factors released from astrocytes provided spatial and temporal control selectively enhancing neuronal differentiation and neurite alignment on topographically different regions of the same substrate. To investigate the potential role of astrocyte-derived soluble factors as cues involved in neuronal differentiation, a non-contact co-culture system was used. Under control conditions, approximately 14% of the AHPCs were immunoreactive (IR) for the neuronal marker, class III β-tubulin (TUJ1-IR). When co-cultured in physical contact with astrocytes, neuronal differentiation increased significantly to about 25%, consistent with our previous results. Moreover, under non-contact co-culture conditions using Transwell insert cultures, neuronal differentiation was dramatically increased to approximately 64%. Furthermore, neurite outgrowth from neuronal cell bodies was considerably greater on the patterned substrate, compared to the non-patterned planar substrate under non-contact co-culture conditions. Taken together, our results demonstrate that astrocyte-derived soluble factors provide cues for specific neuronal differentiation of AHPCs cultured on micropatterned substrates. In addition, a suppressive influence on neuronal differentiation appears to be mediated by contact with co-cultured astrocytes. These results provide important insights into mechanisms for controlling neural progenitor/stem cell differentiation and facilitate development of strategies for CNS repair

Genetic determinants of statin intolerance

BACKGROUND: Statin-related skeletal muscle disorders range from benign myalgias – such as non-specific muscle aches or joint pains without elevated serum creatinine kinase (CK) concentration – to true myositis with >10-fold elevation of serum CK, to rhabdomyolysis and myoglobinuria. The genetic basis of statin-related muscle disorders is largely unknown. Because mutations in the COQ2 gene are associated with severe inherited myopathy, we hypothesized that common, mild genetic variation in COQ2 would be associated with inter-individual variation in statin intolerance. We studied 133 subjects who developed myopathy on statin monotherapy and 158 matched controls who tolerated statins without incident or complaint. RESULTS: COQ2 genotypes, based on two single nucleotide polymorphisms (SNP1 and SNP2) and a 2-SNP haplotype, all showed significant associations with statin intolerance. Specifically, the odds ratios (with 95% confidence intervals) for increased risk of statin intolerance among homozygotes for the rare alleles were 2.42 (0.99 to 5.89), 2.33 (1.13 to 4.81) and 2.58 (1.26 to 5.28) for SNP1 and SNP2 genotypes, and the 2-SNP haplotype, respectively. CONCLUSION: These preliminary pharmacogenetic results, if confirmed, are consistent with the idea that statin intolerance which is manifested primarily through muscle symptoms is associated with genomic variation in COQ2 and thus perhaps with the CoQ10 pathway

Multipotent adult hippocampal progenitor cells maintained as neurospheres favor differentiation toward glial lineages

Adult hippocampal progenitor cells (AHPCs) are generally maintained as a dispersed monolayer population of multipotent neural progenitors. To better understand cell-cell interactions among neural progenitors and their influences on cellular characteristics, we generated free-floating cellular aggregates, or neurospheres, from the adherent monolayer population of AHPCs. Results from in vitro analyses demonstrated that both populations of AHPCs were highly proliferative under maintenance conditions, but AHPCs formed in neurospheres favored differentiation along a glial lineage and displayed greater migrational activity, than the traditionally cultured AHPCs. To study the plasticity of AHPCs from both populations in vivo, we transplanted GFP-expressing AHPCs via intraocular injection into the developing rat eyes. Both AHPC populations were capable of surviving and integrating into the developing host central nervous system, but considerably more GFP-positive cells were observed in the retinas transplanted with neurosphere AHPCs, compared to adherent AHPCs. These results suggest that the culture configuration during maintenance for neural progenitor cells (NPCs) influences cell fate and motility in vitro as well as in vivo. Our findings have implication for understanding different cellular characteristics of NPCs according to distinct intercellular architectures and for developing cell-based therapeutic strategies using lineage-committed NPCs

Syndecan transmembrane domain specifically regulates downstream signaling events of the transmembrane receptor cytoplasmic domain

Despite the known importance of the transmembrane domain (TMD) of syndecan receptors in cell adhesion and signaling, the molecular basis for syndecan TMD function remains un-known. Using in vivo invertebrate models, we found that mammalian syndecan-2 rescued both the guidance defects in C. elegans hermaphrodite-specific neurons and the impaired development of the midline axons of Drosophila caused by the loss of endogenous syndecan. These compensatory ef-fects, however, were reduced significantly when syndecan-2 dimerization-defective TMD mutants were introduced. To further investigate the role of the TMD, we generated a chimera, 2eTPC, com-prising the TMD of syndecan-2 linked to the cytoplasmic domain of platelet-derived growth factor receptor (PDGFR). This chimera exhibited SDS-resistant dimer formation that was lost in the corre-sponding dimerization-defective syndecan-2 TMD mutant, 2eT(GL)PC. Moreover, 2eTPC specifically enhanced Tyr 579 and Tyr 857 phosphorylation in the PDGFR cytoplasmic domain, while the TMD mutant failed to support such phosphorylation. Finally, 2eTPC, but not 2eT(GL)PC, induced phosphorylation of Src and PI3 kinase (known downstream effectors of Tyr 579 phosphorylation) and promoted Src-mediated migration of NIH3T3 cells. Taken together, these data suggest that the TMD of a syndecan-2 specifically regulates receptor cytoplasmic domain function and subsequent downstream signaling events controlling cell behavior. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.1

Unintended knowledge learnt in primary science practical lessons

This study explored the different kinds of unintended learning in primary school practical science lessons. In this study, unintended learning has been defined as student learning that was found to occur that was not included in the teachers learning objectives for that specific lesson. A total of 22 lessons, taught by five teachers in Korean primary schools with 10- to 12-year-old students, were audio-and video recorded. Pre-lesson interviews with the teachers were conducted to ascertain their intended learning objectives. Students were asked to write short memos after the lesson about what they learnt. Post-lesson interviews with students and teachers were undertaken. What emerged was that there were three types of knowledge that students learnt unintentionally: factual knowledge gained by phenomenon-based reasoning, conceptual knowledge gained by relation- or model-based reasoning, and procedural knowledge acquired by practice. Most unintended learning found in this study fell into the factual knowledge and only a few cases of conceptual knowledge were found. Cases of both explicit procedural knowledge and implicit procedural knowledge were found. This study is significant in that it suggests how unintended learning in practical work can be facilitated as an educative opportunity for meaningful learning by exploring what and how students learnt

Astrocyte-derived soluble factors promoting neuronal differentiation of adult neural progenitor cells

Adult neurogenesis occurs in two regions of the brain: the subventricular zone of lateral ventricles and the subgranular zone of hippocampus. Neural progenitor cells found in these neurogenic regions are capable of self-renewing and generating mature neurons and glia. It is known that reciprocal interactions with the local environment can regulate cellular maintenance and plasticity of neural progenitor cells. Astrocytes in the microenvironment surrounding neural progenitor cells have a profound influence on a variety of inter- and intra-cellular processes during adult neurogenesis. Thus, we have investigated the roles of astrocyte-derived factors which influence neural progenitor cell differentiation in vitro. The neural progenitor cells used in our studies were hippocampal progenitor cells isolated from adult rat hippocampus (a gift from F. Gage, Salk Institute, La Jolla, CA). Previous immunocytochemical results showed that co-culture with enriched neonatal astrocytes significantly and selectively increased neuronal differentiation of adult hippocampal progenitor cells. We proposed that the astrocytes present discrete modulators to the overlying adult hippocampal progenitor cells involving contact-mediated or release of soluble factors, or a combination of both. From the first investigation in this dissertation, we found that astrocyte-derived soluble factors specifically promote neuronal differentiation of neural progenitor cells. In the second investigation, we tested a hypothesis that a candidate neurogenic factor produced from astrocytes, interleukin-6, is a key regulator that stimulates functional differentiation of neural progenitor cells to a neuronal fate. Using immunocytochemical analysis and whole-cell recording, we have demonstrated that astrocyte-derived interleukin-6 enhances neuronal differentiation of AHPCs by presenting three main observations: (a) neuronal marker expression was increased, (b) the average length of neurites from neuronal-restricted AHPCs was increased and (c) voltage-gated inward current density was increased with no significant differences in voltage-gated outward current density, apparent resting membrane potential, or cell capacitance. In the third investigation, we examined the influence of extracellular calcium and voltage-gated calcium channel activity on interleukin-6-mediated neuronal differentiation of neural progenitor cells. We observed that interleukin-6-enhanced neuronal differentiation was reduced when cultured in low calcium-containing culture medium or with L-type voltage-gated calcium channel antagonists. Interleukin-6 treatment also increased the fraction of neural progenitor cells immunoreactive for a neuronal marker and for cAMP response element binding protein. Overall, we conclude that astrocyte-derived soluble factors promote neuronal differentiation of adult neural progenitor cells, and astrocyte-derived interleukin-6 is a neurogenic factor able to induce neural progenitor cells to differentiate into neurons. These findings may provide important insights into mechanisms for controlling neural progenitor cell differentiation and facilitate development of cell-based therapeutic strategies using adult neural progenitor cells.</p