Extracellular guanosine and GTP promote expression of differentiation markers and induce S‐phase cell‐cycle arrest in human SH‐SY5Y neuroblastoma cells

Abstract

SH-SY5Y neuroblastoma cells, a model for studying neuronal differentiation, are able to differentiate into either cholinergic or dopaminergic/adrenergic phenotypes depending on media conditions. Using this system, we asked whether guanosine (Guo) or guanosine-5'-triphosphate (GTP) are able to drive differentiation towards one particular phenotype. Differentiation was determined by evaluating the frequency of cells bearing neurites and assessing neurite length after exposure to different concentrations of Guo or GTP for different durations. After 6 days, 0.3 mM Guo or GTP induced a significant increase in the number of cells bearing neurites and increased neurite length. Western blot analyses confirmed that purines induced differentiation; cells exposed to purines showed increases in the levels of GAP43, MAP2, and tyrosine hydroxylase. Proliferation assays and cytofluorimetric analyses indicated a significant anti-proliferative effect of purines, and a concentration-dependent accumulation of cells in S-phase, starting after 24 h of purine exposure and extending for up to 6 days. A transcriptional profile analysis using gene arrays showed that an up-regulation of cyclin E2/cdk2 evident after 24 h was responsible for S-phase entry, and a concurrent down-regulation of cell-cycle progression-promoting cyclin B1/B2 prevented S-phase exit. In addition, patch-clamp recordings revealed that 0.3 mM Guo or GTP, after 6 day incubation, significantly decreased Na(+) currents. In conclusion, we showed Guo- and GTP-induced cell-cycle arrest in neuroblastoma cells and suggest that this makes these cells more responsive to differentiation processes that favor the dopaminergic/adrenergic phenotype.

Purinergic signalling and cancer

“…The human SH-SY5Y neuroblastoma cell line expresses a functional P2X7 receptor that modulates voltage-dependent Ca 2+ channel function [549]. Extracellular guanosine and guanosine triphosphates promote the expression of differentiation markers and induce S-phase cell cycle arrest in SH-SY5Y cells [550]. Agonistinduced stimulation of both A 1 and A 2A receptor induced neurite outgrowth and differentiation of SH-SY5Y neuroblastoma cells in vitro [551].…”

Guanosine and its role in neuropathologies

“…In line with the proposed neuritogenic properties of guanosine, a study conducted by Guarnieri et al sought to investigate whether this nucleoside was capable of promoting a mature neuronal phenotype in human SH-SY5Y neuroblastoma cells [55]. Gene profiling revealed that guanosine is able to induce an S-phase cell cycle arrest and neurite outgrowth that are comparable to those elicited by well-known inducers of neuronal differentiation, such as retinoic acid and phorbol-12-myristate-13-acetate (PMA).…”

Different Mechanisms of Seed Kernel Extract from <i>Mangifera indica</i> on the Growth of Two Colon Cancer Cell Lines

“…This may be associated with the expression of cyclin E and suppression of cyclin B level. A recent report reveals that upregulation of cyclin E and concurrent downregulation of cyclin B prevent S phase exit [20]. This may be the main event that MSP-treated Colo 320DM cells are arrested at S phase rather than G2/M phase.…”

“…MSP arrests cell cycle of SW480 cells in G1 phase, which associates with downregulation of cyclin D1 and E and upregulation of p21. Not the same as SW480 cells, Colo 320DM cells are arrested by MSP treatment in S phase of cell cycle, which may be from the upregulation of cyclin E and downregulation of cyclin B [20]. Cyclin D1 and cyclin E are both key regulators to CDK4 and 6, which phosphorylate retinoblastoma protein and results in retinoblastoma protein to degeneration.…”