Capacity Of Neural Stem Cells Research Articles

Effects of prenatal exposure to THC on hippocampal neural development in offspring

Cannabis use is a worldwide issue with the development of legalization. Prenatal exposure to Δ9-tetrahydrocannabinol (THC), the main psychoactive component of cannabis, is related to affect fetal nervous system development. In our present study, we administered THC to pregnant mice from gestational day 5.5–12.5. Differences in neuronal cell composition and organization between the two groups were found by staining sections of the offspring hippocampus at PND21. In addition, RNA-seq of hippocampal tissue also suggested differences in gene expression due to THC treatment, especially significant enrichment to neurogenesis and neural differentiation. Subsequently, the effect of THC treatment on the proliferation and differentiation capacity of neural stem cells (NSCs) was confirmed. Based on the RNA-seq results, we selected the differentially expressed transcription factor MEF2C for validation. The effect of THC treatment on NSCs differentiation was found to be regulated by knocking down the expression of MEF2C in NSCs. Considering that THC is an agonist of cannabinoid receptor (CB1R), the differentiation outcome of NSC after THC treatment was significantly rescued, by pretreating with the CB1R inhibitor Rimonabant. Notably, pretreatment with Rimonabant restored the expression of MEF2C. Taken together, the present results suggested that THC regulated the MEF2C pathway through CB1R and had an impact on hippocampal neurodevelopment.

Microtubule disruption upon CNS damage triggers mitotic entry via TNF signaling activation.

Repair after traumatic injury often starts with mitotic activation around the lesion edges. Early midline cells in the Drosophila embryonic CNS can enter into division following the traumatic disruption of microtubules. We demonstrate that microtubule disruption activates non-canonical TNF signaling by phosphorylation of TGF-β activated kinase 1 (Tak1) and its target IkappaB kinase (Ik2), culminating in Dorsal/NfkappaB nuclear translocation and Jra/Jun expression. Tak1 and Ik2 are necessary for the damaged-induced divisions. Microtubule disruption caused by Tau accumulation is also reported in Alzheimer's disease (AD). Human Tau expression in Drosophila midline cells is sufficient to induce Tak1 phosphorylation, Dorsal and Jra/Jun expression, and entry into mitosis. Interestingly, activation of Tak1 and Tank binding kinase 1 (Tbk1), the human Ik2 ortholog, and NfkappaB upregulation are observed in AD brains.

Nuclear Isoform of FGF13 Regulates Postnatal Neurogenesis in Hippocampus Through Epigenetic Mechanism

The hippocampus is one of the two niches in mammalian brain with persistent neurogenesis into adulthood. The neurogenic capability of hippocampal neural stem cells (NSCs) declines with age but the molecular mechanisms of this process remain unknown. In this study, we found that fibroblast growth factors 13 (FGF13) is essential for the postnatal neurogenesis in mouse hippocampus and FGF13 deficiency impairs learning and memory. In particular, we found that the nuclear isoform of FGF13, FGF13A, is involved in the maintenance of NSCs and the suppression of neuron differentiation during postnatal hippocampal development. Furthermore, we found that FGF13A interacts with ARID1B, a unit of Brahma-associated factor chromatin remodeling complex, and suppresses the expression of neuron differentiation-associated genes through chromatin modification. Our results suggested that FGF13A is an important regulator for maintaining the self-renewal and neurogenic capacity of NSCs in postnatal hippocampus, revealing a novel epigenetic function of FGFs in neurogenesis.

Disruption of the polyubiquitin gene Ubb reduces the self-renewal capacity of neural stem cells

Ubiquitin (Ub) is a highly conserved eukaryotic protein that plays pivotal roles in cellular signal transduction, differentiation, and proteolysis. Although we have previously reported that disruption of the polyubiquitin gene Ubb is associated with the dysregulated differentiation of neural stem cells (NSCs) into neurons, it is unclear how gene expression patterns are altered in Ubb knockout (KO) NSCs, and whether this altered gene expression contributes to Ubb KO neural phenotypes. To answer these questions, we used RNA-Seq to compare the transcriptomes of Ubb KO NSCs and Ubb heterozygous (HT) controls. We found that the expression levels of most proliferation markers were decreased in Ubb KO NSCs. To determine whether the reduced levels of proliferation markers were due to reduced self-renewal of NSCs, such as radial glia, we measured the levels of the radial glia marker, Pax6, in mouse embryonic brains at 14.5 dpc. We found that Pax6 levels were decreased and the ventricular zone was thinner in the embryonic brains of Ubb KO mice compared to those of wild-type (WT) control mice. To determine whether the decreased self-renewal of Ubb KO NSCs was caused by cell-autonomous defects and not due to their microenvironment, we transplanted NSCs into WT mouse brains using a cannula system. In mouse brain sections, immunoreactivity of the NSC marker, nestin, was much lower in Ubb KO NSCs than in Ubb HT controls. Therefore, our data suggest that cell-autonomous defects, due to the disruption of Ubb, lead to a decrease in the self-renewal capacity of NSCs and may contribute to their dysregulated differentiation into neurons.

Efficacy of Minocycline in Neural Stem Cells Proliferation after Traumatic Brain Injury

BACKGROUND: Neuroinflammation is an important secondary injury mechanism that contributes to neurological impairments after traumatic brain injury (TBI). There is a robust evidence that neuroinflammation will diminish neurogenesis after TBI. Therefore, strategies to attenuate the inflammatory responses are potential to increase neurogenesis following TBI. Minocycline, a second-generation tetracycline antibiotic derivate, has potent anti-inflammatory effect by reducing microglial activation and suppressing some pro-inflammatory cytokines.
 AIM: The aim of this study is to investigate if minocycline could enhance neurogenesis after TBI.
 METHODS: Thirty Sprague Dawley rats were randomized into three treatments group, i.e., sham-operated controls, closed head injury (CHI), and CHI with minocycline. We used the modified Feeney’s weight-drop model for making CHI. For the treatment group, we gave minocycline per oral (50 mg/kg) twice daily for the first 2 days followed by 25 mg/kg once daily for 3 consecutive days. Animals were sacrificed on day 5. To assess the proliferation capacity of neural stem cells (NSC), we performed immunohistochemistry staining with SOX2, brain-derived neurotropic factor (BDNF), and NFR. Cell counts were carried out using light microscope with 1000 times magnification in 20 high-power fields.
 RESULTS: SOX2, NF-E2-related factor 2 (NRF-2), and BDNF were upregulated in the CHI group compared to the sham-operated group (p < 0.05). NRF-2, BDNF, and SOX2 were upregulated also significantly in the CHI+ minocycline group compared to the sham-operated group and the CHI group (p < 0.05).
 CONCLUSION: Minocycline increased the proliferation capacity of NSC.

Proliferative capacity of neural stem cells in hippocampus of rats after traumatic brain injury and its relationship with Janus kinase 2/signaling and transcriptional activation factor 3 signaling pathway

Objective To investigate the proliferative capacity of neural stem cells (NSCs) in rat hippocampus after traumatic brain injury (TBI) and its relationship with Janus kinase 2/signaling and transcriptional activation factor 3 (JAK2/STAT3) signaling pathway activity. Methods A total of 108 SD rats were randomly divided into control group (36 rats) and TBI group (72 rats). The TBI model was constructed by PinPoint™ Precision Cortical Impactor. At 1, 3, 7, 14, 21 and 28 days after injury, the brain tissues were taken for immunofluorescence staining to detect the proliferation of NSCs [5-bromodeoxyuridine (BrdU)+ /stem cell key protein-2 (Sox2)+ ] in hippocampus, and phosphorylated JAK2 (p-JAK2) and phosphorylated STAT3 (p-STAT3) were detected by Western blot. The expression level of p-JAK2 and p-STAT3 as well as the changing trend were analyzed. On the basis of preliminary analysis of the proliferation of NSCs and the change of JAK2/STAT3 signaling pathway activity in hippocampus, another 24 SD rats were randomly divided into TBI+ normal saline group and TBI+ AG490 (JAK2 specific inhibitor) group, with 12 rats in each group. At 7 days after injury, the proliferation of NSCs in hippocampus was detected by immunofluorescence staining, and the expression levels of p-JAK2 and p-STAT3 were detected by Western blot, so as to further confirm the correlation between the proliferation ability of NSCs in hippocampus and JAK2/STAT3 signaling pathway. Results Compared with the control group, the number of NSCs in the hippocampus of the TBI group and the expression of p-JAK2 and p-STAT3 increased. And the most significant increase occurred at 7 days after injury [number of NSCs: 31.2±4.7 in the control group, 111.4±8.1 in the TBI group (P<0.01); p-JAK2: 1.11±0.09 in the control group, 2.16±1.01 in the TBI group (P <0.01); p-STAT3: 1.05±0.06 in the control group and 2.06±0.09 in the TBI group (P<0.01)]. The proliferation of NSCs in hippocampus of TBI group was consistent with the change of p-JAK2 and p-STAT3 expression. Seven days after injury, the expression levels of p-JAK2 and p-STAT3 and the proliferation ability of NSCs in the TBI+ AG490 were significantly decreased [p-JAK2: 2.18 ± 0.15 in the TBI + isotonic saline group, 1.24±0.10 in the TBI+ AG490 group (P<0.01); p-STAT3: 2.21±0.12 in the TBI+ isotonic saline group, 1.25±0.11 in the TBI+ AG490 group (P<0.01); NSCs number: 112.8±8.6 in the TBI+ isotonic saline group, 75.5±6.4 in the TBI+ AG490 group (P<0.05)]. Conclusions The proliferation of NSCs in hippocampus of rats increased after TBI, and the activity of JAK2/STAT3 signaling pathway also increased, following the same trend. JAK2 inhibitor AG490 can reduce the activity of JAK2/STAT3 signaling pathway and the proliferation of NSCs. This can provide reference for researches on TBI promoting nerve regeneration and function repair. Key words: Brain injuries; Neural stem cells; Hippocampus; Signal transduction

Ecrg4 deficiency extends the replicative capacity of neural stem cells in a Foxg1-dependent manner.

The self-renewal activity of neural stem cells (NSCs) has been suggested to decrease with aging, resulting in age-dependent declines in brain function, such as presbyopia and memory loss. The molecular mechanisms underlying decreases in NSC proliferation with age need to be elucidated in more detail to develop treatments that promote brain function. We have previously reported that the expression of esophageal cancer-related gene 4 (Ecrg4) was upregulated in aged NSCs, whereas its overexpression decreased NSC proliferation, suggesting a functional relationship between Ecrg4 and NSC aging. Using Ecrg4-deficient mice in which the Ecrg4 locus was replaced with the lacZ gene, we here show that Ecrg4 deficiency recovered the age-dependent decline in NSC proliferation and enhanced spatial learning and memory in the Morris water-maze paradigm. We demonstrate that the proliferation of Ecrg4-deficient NSCs was partly maintained by the increased expression of Foxg1. Collectively, these results determine Ecrg4 as a NSC aging factor.

Ferritin nanoparticles for improved self-renewal and differentiation of human neural stem cells

BackgroundBiomaterials that promote the self-renewal ability and differentiation capacity of neural stem cells (NSCs) are desirable for improving stem cell therapy to treat neurodegenerative diseases. Incorporation of micro- and nanoparticles into stem cell culture has gained great attention for the control of stem cell behaviors, including proliferation and differentiation.MethodIn this study, ferritin, an iron-containing natural protein nanoparticle, was applied as a biomaterial to improve the self-renewal and differentiation of NSCs and neural progenitor cells (NPCs). Ferritin nanoparticles were added to NSC or NPC culture during cell growth, allowing for incorporation of ferritin nanoparticles during neurosphere formation.ResultsCompared to neurospheres without ferritin treatment, neurospheres with ferritin nanoparticles showed significantly promoted self-renewal and cell-cell interactions. When spontaneous differentiation of neurospheres was induced during culture without mitogenic factors, neuronal differentiation was enhanced in the ferritin-treated neurospheres.ConclusionsIn conclusion, we found that natural nanoparticles can be used to improve the self-renewal ability and differentiation potential of NSCs and NPCs, which can be applied in neural tissue engineering and cell therapy for neurodegenerative diseases.

TGFβ1 transduction enhances immunomodulatory capacity of neural stem cells in experimental autoimmune encephalomyelitis

Bone marrow-derived neural stem cells (BM-NSCs) have therapeutic effect on EAE, an animal model of multiple sclerosis. However, the beneficial effect is suboptimal due to the limited immunomodulatory capacity of these cells. In this study, we engineered BM-NSCs with inducible TGFβ1, a potent immunosuppressive cytokine, to enhance their anti-inflammatory capacity. We found that i.v. injected TGFβ1-BM-NSCs more effectively suppressed clinical severity, inflammation and demyelination of the central nervous system of EAE mice. Transduction of TGFβ1 resulted in a higher percentage of Tregs and lower percentage of Th1 and Th17 cells in the periphery, with increased production of IL-10, and reduced production of IFN-γ, IL-17 and GM-CSF. Moreover, myelin-specific splenic proliferation was also inhibited more profoundly by TGFβ1-BM-NSCs. We also found that TGFβ1-BM-NSCs have the capacity to switch microglia from M1 to M2 phenotype. On the other hand, transduction of TGFβ1 did not affect proliferative ability and differentiating potential of BM-NSCs in vitro and in vivo. Together, these findings demonstrate that transduction of TGFβ1 significantly enhanced the immunomodulatory capacity of BM-NSCs for EAE treatment, through inducing Tregs and an M2 phenotype of macrophages/microglia, while retaining their capacity for neural cell differentiation. Thus, our study provides an easily accessible, inducible and effective therapy for CNS inflammatory demyelination.

Temporal changes in the response of SVZ neural stem cells to intraventricular administration of growth factors

In vivo growth factor (GF) treatment is a promising approach to enhance the regenerative capacity of neural stem cells (NSCs) for brain repair. However, how exogenous GFs affect endogenous NSCs is not well understood. This study investigated the impact of intraventricular administration of fibroblast growth factor 2 and epidermal growth factor on NSCs in the subventricular zone of intact adult mice. GFs were administered for various periods (3, 7, 10, and 14 days), and the proliferation and neuronal production of NSCs were assessed during and after GF treatment. We found that proliferation of NSCs and their progeny is markedly augmented during the first 7 days after the initiation of GF treatment. GF treatment for longer periods, however, did not lead to further increases in the NSC pool, but rather attenuated such proliferation and inhibited neurogenesis. As a result, the production of new olfactory bulb neurons was increased in animals treated with GFs for 7 days but decreased in animals treated for 14 days. These results show time-dependent changes in the response of NSCs to exogenous GFs and demonstrate that precise control of the duration of GF treatment is important for significant enhancement of neuronal production by NSCs in vivo for brain repair.

Diabetes and stem cell function.

Diabetes mellitus is one of the most common serious metabolic diseases that results in hyperglycemia due to defects of insulin secretion or insulin action or both. The present review focuses on the alterations to the diabetic neuronal tissues and skeletal muscle, including stem cells in both tissues, and the preventive effects of physical activity on diabetes. Diabetes is associated with various nervous disorders, such as cognitive deficits, depression, and Alzheimer's disease, and that may be caused by neural stem cell dysfunction. Additionally, diabetes induces skeletal muscle atrophy, the impairment of energy metabolism, and muscle weakness. Similar to neural stem cells, the proliferation and differentiation are attenuated in skeletal muscle stem cells, termed satellite cells. However, physical activity is very useful for preventing the diabetic alteration to the neuronal tissues and skeletal muscle. Physical activity improves neurogenic capacity of neural stem cells and the proliferative and differentiative abilities of satellite cells. The present review proposes physical activity as a useful measure for the patients in diabetes to improve the physiological functions and to maintain their quality of life. It further discusses the use of stem cell-based approaches in the context of diabetes treatment.

Self-organization of neural tissue architectures from pluripotent stem cells.

Despite being a subject of intensive research, the mechanisms underlying the formation of neural tissue architectures during development of the central nervous system remain largely enigmatic. So far, studies into neural pattern formation have been restricted mainly to animal experiments. With the advent of pluripotent stem cells it has become possible to explore early steps of nervous system development in vitro. These studies have unraveled a remarkable propensity of primitive neural cells to self-organize into primitive patterns such as neural tube-like rosettes in vitro. Data from more advanced 3D culture systems indicate that this intrinsic propensity for self-organization can even extend to the formation of complex architectures such as a multilayered cortical neuroepithelium or an entire optic cup. These novel experimental paradigms not only demonstrate the enormous self-organization capacity of neural stem cells, they also provide exciting prospects for studying the earliest steps of human neural tissue development and the pathogenesis of brain malformations in reductionist in vitro paradigms.

RETRACTED: Neurotrophin 3 Transduction Augments Remyelinating and Immunomodulatory Capacity of Neural Stem Cells

Neural stem cells (NSCs) have therapeutic potential in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS); however, to date, their use has resulted in only limited clinical and pathological improvement. To enhance their therapeutic capacity, in the present study, we transduced bone marrow-derived NSCs (BM-NSCs) with neurotrophin 3 (NT-3), a potent neurotrophic factor that is both neuroprotective and immunomodulatory. We found that BM-NSCs transduced with NT-3 reduced central nervous system (CNS) inflammation and neurological deficits in ongoing EAE significantly more than conventional NSC therapy, and, in addition, had the following advantages: (i) enhanced BM-NSC proliferation and differentiation into oligodendrocytes and neurons, as well as inhibited differentiation into astrocytes, thus promoting remyelination and neuronal repopulation, and reducing astrogliosis; (ii) enhanced anti-inflammatory capacity of BM-NSCs, thus more effectively suppressing CNS inflammation and accelerating remyelination; (iii) the easy accessibility of BM-NSCs provides another advantage over brain-derived NSCs for MS therapy; and (iv) a novel Tet-on system we used enables efficient control of NT-3 expression. Thus, our study provides a novel approach to break the vicious inflammation-demyelination cycle, and could pave the way to an easily accessible and highly effective therapy for CNS inflammatory demyelination.

LoudNoise Exposure during Activity and Neurogenesis in the Living Rat Brain: Preliminary Study

Objective: The effects of loud noise (80 dB) on neurogenesis in the sub-ventricular zone (SVZ) and the hippocampal sub-granular zone (SGZ) was investigated in vivo in the rat brain using positron emission tomography (PET) with 3’-deoxy-3’- [18F]fluoro-L-thymidine ( [18F]FLT), a marker for thymidine kinase I (TK-1) related to cell proliferation and for neurogenesis in SVZ and SGZ. Methods: Five young Sprague-Dawley rats were exposed to loud noise from two speakers set in front of their cages during the night for 4 nights, and 5 control rats were exposed to normal quiet conditions. PET was performed 4 days after the exposure. Results: The accumulation of [18F] FLT in SVZ was significantly lower in the rats with noise exposure than in the control rats. The level of [18F] FLT uptake was lower in the SVZ than in the SGZ. Conclusion: Our results indicate that loud noise exposure during the awake period may be related to the decrease in proliferative capacity of neural stem cells especially in the SVZ of the living rat brain. This finding may indicate that persistent loud noise during daily life is harmful to brain development in humans

Targeting of the Adenosine Receptors as A Novel Strategy for the Treatment of Arterial Hypertension

Objective: The effects of loud noise (80 dB) on neurogenesis in the sub-ventricular zone (SVZ) and the hippocampal sub-granular zone (SGZ) was investigated in vivo in the rat brain using positron emission tomography (PET) with 3’-deoxy-3’- [18F]fluoro-L-thymidine ( [18F]FLT), a marker for thymidine kinase I (TK-1) related to cell proliferation and for neurogenesis in SVZ and SGZ. Methods: Five young Sprague-Dawley rats were exposed to loud noise from two speakers set in front of their cages during the night for 4 nights, and 5 control rats were exposed to normal quiet conditions. PET was performed 4 days after the exposure. Results: The accumulation of [18F] FLT in SVZ was significantly lower in the rats with noise exposure than in the control rats. The level of [18F] FLT uptake was lower in the SVZ than in the SGZ. Conclusion: Our results indicate that loud noise exposure during the awake period may be related to the decrease in proliferative capacity of neural stem cells especially in the SVZ of the living rat brain. This finding may indicate that persistent loud noise during daily life is harmful to brain development in humans.

Exonuclease 1 is essential for maintaining genomic stability and the proliferative capacity of neural but not hematopoietic stem cells

Exonuclease 1 (Exo1) has been implicated in the regulation of DNA damage responses in stem cells with dysfunctional telomeres. However, it is unclear whether Exo1-mediated DNA maintenance pathways play a role in the maintenance of genomic stability and the self-renewal of tissue stem cells in mice with functional telomeres. Here, we analyzed the proliferative capacity of neural stem cells (NSCs) and hematopoietic stem cells (HSCs) from Exo1−/− mice. Our study shows that Exo1 deficiency impairs the maintenance of genomic stability and proliferative capacity in NSCs but not HSCs. In line with these results, we detected a decrease in proliferation and an up-regulation of p21 expression levels in Exo1-deficient NSCs but not Exo1-deficient HSCs. Our data provide experimental evidence that Exo1 deficiency has a differential impact on the homeostasis and proliferative capacity of tissue stem cells in the brain and bone marrow, suggesting that different tissue stem cells utilize distinct mechanisms for maintaining their genomic stability. Our current study provides important insight into the role of Exo1-mediated DNA maintenance pathways in the maintenance of genomic stability and the proliferative capacity of tissue stem cells.

ID1 affects the efficacy of radiotherapy in glioblastoma through inhibition of DNA repair pathways

Glioblastoma multiforme (GBM) is characterized by poor therapeutic response and poor overall survival. It is crucial that more effective therapies be developed for the treatment of GBM. Inhibitor of DNA binding protein-1 (ID1) has been shown to maintain the self-renewal capacity of neural stem cells and might be involved in the therapeutic resistance of GBM. In the present study, we explored survival data from the The Cancer Genome Atalas database that were based on ID1 expression for patients diagnosed with primary GBMs. Interestingly, patients with high ID1 expression had better survival than patients with low ID1 expression, and a strong correlation was found between radiotherapy efficacy, ID1 expression, and overall survival. We further investigated the relationship between ID1 expression and the radiosensitivity of glioblastoma using glioblastoma cell lines. The clonogenic formation assay showed that U87 ID1-shRNA cells were much less sensitive to radiation. Moreover, both the results of the γH2AX foci staining assay and the comet assay further revealed that ID1 negatively regulates DNA repair processes by downregulating the expression of genes such as DNA ligase IV (LIG4) and ataxia-telangiectasia-mutated. Additionally, ID1 induces G2/M arrest in U87 cells. Taken together, these results suggest that ID1 may be a new prognostic marker for GBM and have important implications for the therapeutic strategies used to treat GBM patients.

245 DYRK1a Modulates the Self-renewal Capacity of Neural Stem Cells and Tumor Initiating Cells: Targetting the Achilles Heels of EGFR Addicted Glioblastomas

245 DYRK1a Modulates the Self-renewal Capacity of Neural Stem Cells and Tumor Initiating Cells: Targetting the Achilles Heels of EGFR Addicted Glioblastomas

Improvement of neurological recovery and stimulation of neural progenitor cell proliferation by intrathecal administration of Sonic hedgehog

Sonic hedgehog (Shh) is a glycoprotein molecule that has been shown to be associated with the proliferative capacity of endogenous neural precursor cells during embryonic development. It has also been shown to regulate the proliferative capacity of neural stem cells in the adult subventricular zone (SVZ), which are also upregulated in animal models of ischemic stroke. In the present study, the effects of exogenous administration of intrathecal Shh protein were examined in the setting of a rodent model of ischemic stroke, with particular attention given to endogenous neural stem cell proliferation and migration as well as inducible differences in behavioral recovery. A rodent model of ischemic stroke was created using the intraluminal suture method of reversible middle cerebral artery occlusion. Animals were treated with intrathecal administration of Shh protein at 24 hours after the onset of the stroke. Behavioral testing was performed, and the animals were killed for measurements of infarct volume 7 days after stroke. Immunohistochemical staining was performed and measurements of cellular proliferation were obtained, with a focus on the proportion and distribution of neural progenitor cells in the SVZ. These values were compared across experimental groups. Treatment with intrathecal Shh protein resulted in significant improvement in behavioral function compared with the control group, with a significant reduction of ischemic tissue in the cerebral hemisphere. An increase of nestin immunoreactive cells was observed along the SVZ. Intrathecal Shh agonist at doses that upregulate spinal cord GLI1 transcription increases the population of neural precursor cells after spinal cord injury in adult rats. Intrathecal administration of Shh protein appears to have a neuroprotective effect in animal models of ischemic stroke and is associated with improved behavioral recovery, which may be related to its effects on neurogenesis in the SVZ and could be associated with improved functional recovery.

Proliferation capacity of cord blood derived neural stem cell line on different micro-scale biofunctional domains.

Physical interactions of cells with the adhesive substrates of the microenvironment as well as the presence of the soluble growth factors are important for the proliferation capacity of neural stem cells. We have used biofunctionalized surface domains microcontact printed with either synthetic polyaminoacid poly-L-lysine or extracellular matrix (ECM) component such as fibronectin, to study the proliferation capacity of human umbilical cord blood-derived neural stem cells (HUCB-NSC). The proliferation measured by the expression of Ki-67 protein was accompanied by the investigation of the cell morphology under the transmission and scanning electron microscopy in different culture time, plating densities of cells and medium condition (serum-free or 2 percent of FBS). The poly-L-lysine domains of defined micro-scale area promoted the presence of round, loosely attached Ki-67-positive cells, while fibronectin domains of the same size allowed appearance of flattened, strongly attached cells with more differentiated phenotype. These results were in agreement with the non-specific, electrostatic type of interaction between cell and substrate on poly-L-lysine and integrin receptor-mediated specific adhesion on fibronectin. In this report we have described in vitro culture conditions, which allow for immobilization of the non-differentiated and highly proliferating population of neural stem/progenitor cells to the biofunctionalized surface. The microarrays with bioactive domains allocating non-differentiated and proliferating neural stem/progenitor cells may find application for drug and chemicals toxicology screening of diverse factors influencing neural development.