Uracil nucleotides stimulate human neural precursor cell proliferation and dopaminergic differentiation: involvement of MEK/ERK signalling

Abstract

Isolation and propagation of neural stem cells derived from human brain tissue uniquely enables the study of human neurogenesis in vitro. In addition, ex vivo-expanded human neural stem/precursor cells (NPCs) may offer novel therapeutic strategies. We investigated the effects of extracellular nucleotides on the proliferation and differentiation of human mesencephalic neural stem/precursor cells (hmNPCs). When combined with the mitogens epidermal growth factor and fibroblast growth factor 2, UTP (1 lM) boosted proliferation of hmNPCs as shown by increased expression of the proliferation marker proliferating cell nuclear antigen (330%). UTPinduced proliferation was abrogated by the preferential P2Y receptor blocker pyridoxal phosphate-6-azophenyl-2¢,4¢-disulphonic acid (PPADS). UTP also stimulated dopaminergic differentiation. Treatment with UTP (100 lM) increased the number of tyrosine hydroxylase (TH)-positive cells and TH protein by 267 and 319% respectively. UTP-stimulated dopaminergic differentiation of hmNPCs was blocked by the P2 receptor antagonists suramin (10 lM) and PPADS (100 lM). In addition, UDP (1 lM) enhanced TH protein expression by 194%. During differentiation, treatment with UTP stimulated the extracellular signal-regulated kinase (ERK) pathway. Both ERK1/2 phosphorylation and dopaminergic differentiation were inhibited by U0126, a selective ERK kinase inhibitor, as well as by suramin. When other P2 receptor agonists (ATP, ADP and adenosine 5¢-O-(2-thiophosphate) (ADPbS); all 100 lM) were applied, both proliferation and dopaminergic differentiation of NPCs were compromised. We conclude that uracil nucleotides exert specific P2 receptor-mediated effects on midbrain-derived human NPCs, and may be used to enhance both proliferation and dopaminergic differentiation. Keywords: extracellular signal-regulated kinase 1/2 pathway, neural stem/precursor cells, P2 receptors, tyrosine hydroxylase, UTP. Neural stem/precursor cells (NPCs) serve as a model of brain development, providing promise for currently incurable brain diseases and injuries. Based on their unique capacity to migrate throughout the brain and to target invading tumour cells, transplantation of NPCs offers a new

Activation of UTP-Sensitive P2Y2Receptor Induces the Expression of Cholinergic Genes in Cultured Cortical Neurons: A Signaling Cascade Triggered by Ca2+Mobilization and Extracellular Regulated Kinase Phosphorylation

“…In line with the previous reports, UTP and PSB1114 (P2Y 2 R-specific agonists) induced robust phosphorylation of ERK1 and ERK2 in a time-dependent manner. Our results suggest that MAP kinase signaling is activated by Ca 21 and PKC, and it appears that nucleotide-mediated P2YR activation initiates similar signaling cascades with similar cellular consequences in muscle and neurons (Tu et al, 2000;Choi et al, 2003;Milosevic et al, 2006;Siow et al, 2010). In our results, the responses of UTP in Ca 21 mobilization and Erk phosphorylation in Fig.…”

The PRKAA1/AMPKα1 pathway triggers autophagy during CSF1-induced human monocyte differentiation and is a potential target in CMML

“…There are few studies in the literature that show that P2Y6 receptor activation is involved in differentiation processes such as osteogenic, monocytic, or dopaminergic differentiation. 17,[33][34][35] Moreover, it has been previously reported that P2RY13 activation by ADP induces autophagy in hepatoma cell lines. 36 However to the best of our knowledge, this is the first description of purinergic receptor-mediated differentiation being dependent on autophagy induction.…”

Perspectives of purinergic signaling in stem cell differentiation and tissue regeneration

“…These data suggest the involvement of purinergic signaling in dopaminergic cell differentiation and possible applications for purinergic receptors in in vitro differenti-ation cultures for posterior PD cell therapy [74]. However, more studies are needed to clarify whether extracellular nucleotides may contribute to favorable endogenous niches for stem cell transplantation or even recruit endogenous NPCs for dopaminergic differentiation.…”

“…Exogenous molecules are known to guide neural differentiation and are responsible for the high grade of phenotype specification, including induction of axonal growth and establishment of synaptic contact [73]. Milosevic et al [74] detected P2Y4, P2Y6, and P2X4 receptor expression in cultured human NPCs from human fetal midbrain. UTP and UDP are known as agonists of the P2Y2/P2Y4 and P2Y6 receptors, respectively [75].…”