P2Y2 and P2Y6 receptor activation elicits intracellular calcium responses in human adipose-derived mesenchymal stromal cells

Abstract

Adipose tissue contains self-renewing multipotent cells termed mesenchymal stromal cells. In situ, these cells serve to expand adipose tissue by adipogenesis, but their multipotency has gained interest for use in tissue regeneration. Little is known regarding the repertoire of receptors expressed by adipose-derived mesenchymal stromal cells (AD-MSCs). The purpose of this study was to undertake a comprehensive analysis of purinergic receptor expression. Mesenchymal stromal cells were isolated from human subcutaneous adipose tissue and confirmed by flow cytometry. The expression profile of purinergic receptors was determined by quantitative real-time PCR and immunocytochemistry. The molecular basis for adenine and uracil nucleotide-evoked intracellular calcium responses was determined using Fura-2 measurements. All the known subtypes of P2X and P2Y receptors, excluding P2X2, P2X3 and P2Y 12 receptors, were detected at the mRNA and protein level. ATP, ADP and UTP elicited concentration-dependent calcium responses in mesenchymal cells (N = 7-9 donors), with a potency ranking ADP (EC 50 1.3 ± 1.0 μM) > ATP (EC 50 2.2 ± 1.1 μM) = UTP (3.2 ± 2.8 μM). Cells were unresponsive to UDP (< 30 μM) and UDPglucose (< 30 μM). ATP responses were attenuated by selective P2Y 2 receptor antagonism (AR-C118925XX; IC 50 1.1 ± 0.8 μM, 73.0 ± 8.5% max inhibition; N = 7 donors), and UTP responses were abolished. ADP responses were attenuated by the selective P2Y 6 receptor antagonist, MRS2587 (IC 50 437 ± 133nM, 81.0 ± 8.4% max inhibition; N = 6 donors). These data demonstrate that adenine and uracil nucleotides elicit intracellular calcium responses in human AD-MSCs with a predominant role for P2Y 2 and P2Y 6 receptor activation. This study furthers understanding about how human adipose-derived mesenchymal stromal cells can respond to external signalling cues.

Adult mesenchymal stem cells: is there a role for purine receptors in their osteogenic differentiation?

“…In contrast, P2X6R mRNA and protein levels decrease in the same ASCs committed toward osteogenesis, so that such a variable expression may be critical for early MSCs commitment to differentiate [198]. In more recent studies, in which human ASCs were again investigated, transcripts for multiple purinoreceptors were found, confirming the presence of P2R previously detected [199], besides most adenosine receptors [197]. In these cells, the effects of specific agonists and antagonists suggested that both P2Y1R and P2Y13R were mainly responsive to ADP, while P2Y4R and P2Y11R served as primary UTP and ATP receptors, respectively.…”

P2Y2 Receptor Promotes High-Fat Diet-Induced Obesity

“…Recruitment to this lineage gives rise to preadipocytes, which when induced, undergo mitotic clonal expansion and differentiate into adipocytes. It has been reported that human adipose tissue derived MSC express several purinergic receptors (P2X and P2Y series), but with a predominant role for P2Y 2 and P2Y 6 receptors in eliciting intracellular calcium response (54). There is also evidence that purinergic receptors play a role in stem cell differentiation (55).…”

Adenosine triphosphate enhances osteoblast differentiation of rat dental pulp stem cells via the PLC–IP3 pathway and intracellular Ca2+ signaling

“…It is well established that activation of P2Y receptors increases PLC activity and IP 3 formation for Ca 2+ signaling (Ciciarello et al, 2013). This is supported by findings in adipose-derived mesenchymal stem cells where ATP induces intracellular Ca 2+ signals via P2Y2 receptors (Ali, Turner, & Fountain, 2018). The family of P2X receptors are ATP-gated ion channels that conduct Na + , K + and Ca 2+ ions (Roberts et al, 2006).…”