Connexin Regulation and Modulation of Neural Stem Cell Differentiation Induced by Cell-Permeable Itaconate.

Robust In Vivo Transduction of Nervous System and Neural Stem Cells by Early Gestational Intra Amniotic Gene Transfer Using Lentiviral Vector

Engineered extracellular vesicles for ischemic stroke treatment

IGFBP2 promotes neural stem cell maintenance and proliferation differentially associated with glioblastoma subtypes

Neural stem cells (NSCs) have the potential to replace the major cell types of the central nervous system (CNS) and may be important in therapies for injuries to and diseases of the CNS. However, for such treatments to be safe and successful, NSCs must survive and differentiate appropriately following transplantation. A number of polymer scaffolds have shown promise in improving the survival and promoting the differentiation of NSCs. To capitalize on the interaction between scaffolds and NSCs, we need to determine the fundamental material properties that influence NSC behavior. To investigate the role of material properties on NSCs, we synthesized a library of 52 hydrogels composed of poly(ethylene glycol) and poly(L-lysine) (PLL). This library of hydrogels allows independent variation of chemical and mechanical properties across a wide range of values. By culturing NSCs on this library, we have identified a subset of gels that promotes NSC migration and a further subset that promotes NSC differentiation. By combining the material properties of these subsets with the cell behavior, we determined that mechanical properties play a critical role in NSC behavior with elastic moduli promoting NSC migration and neuronal differentiation. Amine concentration is less critical, but PLL molecular weight also plays a role in NSC differentiation.

BackgroundThe limited neuronal differentiation of the endogenous or grafted neural stem cells (NSCs) after brain injury hampers the clinic usage of NSCs. Panax notoginseng saponins (PNS) were extensively used for their clinical value, such as in controlling blood pressure, blood glucose, and inhibiting neuronal apoptosis and enhancing neuronal protection, but whether or not it exerts an effect in promoting neuronal differentiation of the endogenous NSCs is completely unclear and the potential underlying mechanism requires further exploration.MethodsFirstly, we determined whether PNS could successfully induce NSCs to differentiate to neurons under the serum condition. Mass spectrometry and quantitative polymerase chain reaction (Q-PCR) were then performed to screen the differentially expressed proteins (genes) between the PNS + serum and serum control group, upon which dihydropyrimidinase-like 2 (DPYSL2), a possible candidate, was then selected for the subsequent research. To further investigate the actual role of DPYSL2 in the NSC differentiation, DPYSL2-expressing lentivirus was employed to obtain DPYSL2 overexpression in NSCs. DPYSL2-knockout rats were constructed to study its effects on hippocampal neural stem cells. Immunofluorescent staining was performed to identify the differentiation direction of NSCs after 7 days from DPYSL2 transfection, as well as those from DPYSL2-knockout rats.ResultsSeven differentially expressed protein spots were detected by PD Quest, and DPYSL2 was found as one of the key factors of NSC differentiation in a PNS-treated condition. The results of immunostaining further showed that mainly Tuj1 and GFAP-positive cells increased in the DPYSL2-overexpressed group, while both were depressed in the hippocampal NSCs in the DPYSL2-knockout rat.ConclusionsThe present study revealed that the differentiation direction of NSCs could be enhanced through PNS administration, and the DPYSL2 is a key regulator in promoting NSC differentiation. These results not only emphasized the effect of PNS but also indicated DPYSL2 could be a novel target to enhance the NSC differentiation in future clinical trials.

During neuroinflammation, monocytes that infiltrate the central nervous system (CNS) may contribute to regenerative processes depending on their activation status. However, the extent and mechanisms of monocyte-induced CNS repair in patients with neuroinflammatory diseases remain largely unknown, partly due to the lack of a fully human assay platform that can recapitulate monocyte-neural stem cell interactions within the CNS microenvironment. We therefore developed a human model system to assess the impact of monocytic factors on neural stem cells, establishing a high-content compatible assay for screening monocyte-induced neural stem cell proliferation and differentiation. The model combined monocytes isolated from healthy donors and human embryonic stem cell derived neural stem cells and integrated both cell-intrinsic and -extrinsic properties. We identified CNS-mimicking culture media options that induced a monocytic phenotype resembling CNS infiltrating monocytes, while allowing adequate monocyte survival. Monocyte-induced proliferation, gliogenic fate and neurogenic fate of neural stem cells were affected by the conditions of monocytic priming and basal neural stem cell culture as extrinsic factors as well as the neural stem cell passage number as an intrinsic neural stem cell property. We developed a high-content compatible human in vitro assay for the integrated analysis of monocyte-derived factors on CNS repair.

The present study comprised a series of experiments to investigate the mechanism underlying the effect of ginsenoside on the self-renewal, proliferation and differentiation of neural stem cells (NSCs) undergoing oxygen-glucose deprivation/reperfusion (OGD/R) in vitro. The NSCs, which were isolated from the hippocampus of embryonic day 17 embryo rats, were subjected to OGD/R to establish an in vitro model of brain ischemia-reperfusion, following which different doses of ginsenoside were administered to the model. The proliferation of the NSCs was determined using MTT colorimetry and nestin/bromodeoxyuridine (BrdU) immunofluorescent double-labeling. The NSCs were identified by measuring the expression of nestin, and the differentiation of NSCs was assessed through the immunofluorescent double-labeling of nestin/vimentin and nestin/neuron-specific class III β-tubulin (tuj-1). The protein levels of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α) were detected to investigate the function and mechanism of ginsenoside on ischemic stroke using an enzyme-linked immunosorbent assay. Marked increases in the optical density, area density and numbers of nestin/BrdU-, nestin/vimentin- and nestin/tuj-1-positive cells were found in the ginsenoside-treated group. Compared with the control group, enhanced expression levels of BrdU, tuj-1 and vimentin were found in the ginsenoside-treated group, suggesting that ginsenoside may significantly promote the proliferation and differentiation of NSCs. The results of the present study also showed that ginsenoside significantly increased the protein level of HIF-1α (P<0.05) in the NSCs exposed to OGD/R. These results indicated that ginsenoside may maintain NSC replication, promote NSC proliferation and promote NSC differentiation into neurons and astrocytes. Ginsenoside may initiate the expression of downstream VEGF, which is involved in promoting the survival, self-renewal and differentiation of NSCs.

Cold atmospheric plasmas (CAPs) have been shown to be capable of enhancing stem cell differentiation, especially directed differentiation of neural stem cells (NSCs). Consequently, one-step CAP treatment shows promise as an aid to tissue transplantation. However, the mechanisms involved in the enhancement of NSCs differentiation by CAP treatment are not yet fully understood. We have previously shown that in atmospheric helium plasma jet treatment, nitric oxide (NO) is the main factor involved in promoting NSC differentiation. This article further investigated the possible signaling pathways stimulated by NO in the neuronal differentiation of C17.2-NSCs after plasma treatment. Extracellular and intracellular NO concentrations were measured at different time points of incubation to monitor NO production. Meanwhile, the expressions of related genes and proteins were detected by quantitative real-time polymerase chain reaction and western blot, respectively. It is found that plasma treatment could both generate extracellular NO and increase extracellular NO concentration by inducing inducible nitric oxide synthase expression. The synergetic effect of extracellular and intracellular NO then downregulated Notch1 and Id2, and upregulated Ngn2 and Ascl1, thereby activating downstream NeuroD expression and finally enhancing and directing differentiation of NSCs into neurons.Cold atmospheric plasmas (CAPs) have been shown to be capable of enhancing stem cell differentiation, especially directed differentiation of neural stem cells (NSCs). Consequently, one-step CAP treatment shows promise as an aid to tissue transplantation. However, the mechanisms involved in the enhancement of NSCs differentiation by CAP treatment are not yet fully understood. We have previously shown that in atmospheric helium plasma jet treatment, nitric oxide (NO) is the main factor involved in promoting NSC differentiation. This article further investigated the possible signaling pathways stimulated by NO in the neuronal differentiation of C17.2-NSCs after plasma treatment. Extracellular and intracellular NO concentrations were measured at different time points of incubation to monitor NO production. Meanwhile, the expressions of related genes and proteins were detected by quantitative real-time polymerase chain reaction and western blot, respectively. It is found that plasma treatment could both genera...

Control of the growth and differentiation of neural stem cells is fundamental to brain development and is largely dependent on the Notch signaling pathway. The mechanism by which the activity of Notch is regulated during brain development has remained unclear, however. Fbxw7 (also known as Fbw7, SEL-10, hCdc4, or hAgo) is the F-box protein subunit of an Skp1-Cul1-F-box protein (SCF)-type ubiquitin ligase complex that plays a central role in the degradation of Notch family members. We now show that mice with brain-specific deletion of Fbxw7 (Nestin-Cre/Fbxw7(F/F) mice) die shortly after birth with morphological abnormalities of the brain and the absence of suckling behavior. The maintenance of neural stem cells was sustained in association with the accumulation of Notch1 and Notch3, as well as up-regulation of Notch target genes in the mutant mice. Astrogenesis was also enhanced in the mutant mice in vivo, and the differentiation of neural progenitor cells was skewed toward astrocytes rather than neurons in vitro, with the latter effect being reversed by treatment of the cells with a pharmacological inhibitor of the Notch signaling pathway. Our results thus implicate Fbxw7 as a key regulator of the maintenance and differentiation of neural stem cells in the brain.

Stage-dependent STAT3 activation is involved in the differentiation of rat hippocampus neural stem cells

Traditional therapeutic strategies for spinal cord injury (SCI) are insufficient to repair locomotor function because of the failure of axonal reconnection and neuronal regeneration in the injured central nervous system (CNS). Neural stem cell (NSC) transplantation has been considered a potential strategy and is generally feasible for repairing the neural circuit after SCI; however, the most formidable problem is that the neuronal differentiation rate of NSCs is quite limited. Therefore, it is essential to induce the neuronal differentiation of NSCs and improve the differentiation rate of NSCs in spinal cord repair. Our results demonstrate that both Wnt5a and miRNA200b-3p could promote NSC differentiation into neurons and that Wnt5a upregulated miRNA200b-3p expression through MAPK/JNK signaling to promote NSC differentiation into neurons. Wnt5a could reduce RhoA expression by upregulating miRNA200b-3p expression to inhibit activation of the RhoA/Rock signaling pathway, which has been reported to suppress neuronal differentiation. Overexpression of RhoA abolished the neurogenic capacity of Wnt5a and miRNA200b-3p. In vivo, miRNA200b-3p was critical for Wnt5a-induced NSC differentiation into neurons to promote motor functional and histological recovery after SCI by suppressing RhoA/Rock signaling. These findings provide more insight into SCI and help with the identification of novel treatment strategies.

BackgroundCircular RNAs (circRNAs) have recently been found to be expressed in human brain tissue, and many lines ofevidence indicate that circRNAs play regulatory roles in neurodevelopment. Proliferation and differentiation of neural stem cells (NSCs) are critical parts during development of central nervous system (CNS).To date, there have been no reports ofcircRNA expression profiles during the differentiation of mouse NSCs. We hypothesizethat circRNAs mayregulate gene expression in the proliferation anddifferentiation of NSCs.ResultsIn this study, we obtained NSCs from the wild-type C57BL/6 J mouse fetal cerebral cortex. We extracted total RNA from NSCs in different differentiation stagesand then performed RNA-seq. By analyzing the RNA-Seq data, we found 37circRNAs and 4182 mRNAs differentially expressedduringthe NSC differentiation. Gene Ontology (GO) enrichment analysis of thecognate linear genes of these circRNAsrevealed that some enriched GO terms were related to neural activity. Furthermore, we performed a co-expression network analysis of these differentially expressed circRNAs and mRNAs. The result suggested a stronger GO enrichmentin neural features for both the cognate linear genes of circRNAs and differentially expressed mRNAs.ConclusionWe performed the first circRNA investigation during the differentiation of mouse NSCs. Wefound that12 circRNAs might have regulatory roles duringthe NSC differentiation, indicating that circRNAs might be modulated during NSC differentiation.Our network analysis suggested the possible complex circRNA-mRNA mechanisms during differentiation, and future experimental workis need to validate these possible mechanisms.

MicroRNA-378 regulates neural stem cell proliferation and differentiation in vitro by modulating Tailless expression

Inflammation is the primary pathological feature of neurodegenerative diseases such as Alzheimer's disease (AD) and Parkinson's disease. Proinflammatory molecules (e.g., S100A9) play important roles during the progression of the diseases by regulating behavior and fate of multiple cell types in the nervous system. Our earlier studies reveal that S100A9 is toxic to neurons, and its interaction with Aβ peptides leads to the formation of large nontoxic amyloidogenic aggregates, suggesting a protective role of coaggregation with Aβ amyloids. We herein demonstrate that S100A9 interacts with neural stem cells (NSCs) and causes NSC differentiation. In the brain of transgenic AD mouse models, we found large quantities of proinflammatory S100A9, which colocalizes with the differentiated NSCs. NSC sphere formation, which is a representative character of NSC stemness, is also substantially inhibited by S100A9. These results suggest that S100A9 is a representative marker for the inflammatory conditions in AD, and it promotes NSC differentiation. Intriguingly, in contrast to the death of both stem and differentiated NSCs caused by high S100A9 doses, S100A9 at a moderate concentration is toxic only to the early differentiated NSCs but not the stem cells. We therefore postulate that, at the early stage of AD, the expression of S100A9 leads to NSC differentiation, which remedies the neuron damage. The application of drugs, which help maintain NSC stemness (e.g., the platelet-derived growth factor, PDGF), may help overcome the acute inflammatory conditions and improve the efficacy of NSC transplantation therapy.

SummaryThe consequences of DNA damage generation in mammalian somatic stem cells, including neural stem cells (NSCs), are poorly understood despite their potential relevance for tissue homeostasis. Here, we show that, following ionizing radiation-induced DNA damage, NSCs enter irreversible proliferative arrest with features of cellular senescence. This is characterized by increased cytokine secretion, loss of stem cell markers, and astrocytic differentiation. We demonstrate that BMP2 is necessary to induce expression of the astrocyte marker GFAP in irradiated NSCs via a noncanonical signaling pathway engaging JAK-STAT. This is promoted by ATM and antagonized by p53. Using a SOX2-Cre reporter mouse model for cell-lineage tracing, we demonstrate irradiation-induced NSC differentiation in vivo. Furthermore, glioblastoma assays reveal that irradiation therapy affects the tumorigenic potential of cancer stem cells by ablating self-renewal and inducing astroglial differentiation.