Abstract
Schwann cells (SCs) are fundamental for development, myelination and regeneration in the peripheral nervous system. Slow growth rate and difficulties in harvesting limit SC applications in regenerative medicine. Several molecules, including receptors for neurosteroids and neurotransmitters, have been suggested to be implicated in regulating physiology and regenerative potential of SCs. Adipose-derived stem cells (ASCs) can be differentiated into SC-like phenotype (dASC) sharing morphological and functional properties with SC, thus representing a valid SC alternative. We have previously shown that dASC express γ-aminobutyric-acid receptors, which modulate their proliferation and neurotrophic potential, although little is known about the role of other neurotransmitters in ASC. In this study, we investigated the expression of purinergic receptors in dASC. Using reverse transriptase (RT)-PCR, western blot analyses and immunocytochemistry, we have demonstrated that ASCs express P2X3, P2X4 and P2X7 purinoceptors. Differentiation of ASCs towards glial phenotype was accompanied by upregulation of P2X4 and P2X7 receptors. Using Ca2+-imaging techniques, we have shown that stimulation of purinoceptors with adenosine 5′-triphosphate (ATP) triggers intracellular Ca2+ signals, indicating functional activity of these receptors. Whole-cell voltage clamp recordings showed that ATP and BzATP induced ion currents that can be fully inhibited with specific P2X7 antagonists. Finally, using cytotoxicity assays we have shown that the increase of intracellular Ca2+ leads to dASC death, an effect that can be prevented using a specific P2X7 antagonist. Altogether, these results show, for the first time, the presence of functional P2X7 receptors in dASC and their link with critical physiological processes such as cell death and survival. The presence of these novel pharmacological targets in dASC might open new opportunities for the management of cell survival and neurotrophic potential in tissue engineering approaches using dASC for nerve repair.
Highlights
Nerve injury involves the response of Schwann cells (SCs), the glial cells of the peripheral nervous system.[6]
We have previously shown that differentiated ASCs (dASC) express c-aminobutyric-acid receptors, which modulate their proliferation and neurotrophic potential, little is known about the role of other neurotransmitters in Adipose-derived stem cells (ASCs)
Representative glial fibrillary acidic protein (GFAP) immunostainings of undifferentiated ASCs (uASC), dASC and neonatal SC (nSC) are shown in red in Figures 1d–f, respectively
Summary
Nerve injury involves the response of Schwann cells (SCs), the glial cells of the peripheral nervous system.[6]. Received 07.4.13; revised 24.5.13; accepted 19.6.13; Edited by D Bano rate and the neurotrophic potential of dASC could be the key requirement for their clinical employability in nerve repair Several molecules such as neurosteroids, growth hormones and neurotransmitters have been suggested as potential pharmacological modulators of SC physiology.[29] In particular, neurotransmitters such as g-aminobutyric acid (GABA) and adenosine 50-triphosphate (ATP) have been shown to affect SC functional responses and differentiation.[30,31,32,33,34] Recently, we have shown that dASC express functional GABAA and GABAB receptors that modulate SC proliferation and release of neurotrophic factors.[35,36,37] The expression of other neurotransmitter receptors in dASC has not been investigated, purinergic receptors influence the adipogenic and osteogenic differentiation of human ASC.[38]. Tissue damage is often associated with massive increase of ATP on the injury site, which induces neuronal cell death following spinal cord injuries, an effect that is prevented by P2X7-specific antagonists.[43]
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