Abstract
High interstitial level of ATP and its lysate adenosine in the cancer microenvironment are considered a halo mark of cancer. Adenosine acts as a strong immune suppressor. However, the source of ATP release is unclear. We clarified the release of ATP via volume-regulated anion channels (VRACs) in breast cell lines using an ATP luminescence imaging system. We detected a slowly rising diffuse pattern of ATP release that was only observed in undifferentiated cells, not in differentiated primary cultured cells. This was confirmed by suppression with DCPIB, a blocker of VRACs, and shRNA for LRRC8A, an indispensable subunit of VRACs. We herein demonstrated that the inflammatory mediator sphingosine-1-phosphate (S1P), which exists abundantly in the cancer microenvironment, induced a diffuse pattern of ATP release isovolumetrically. The response was dose-dependent and suppressed by the knock-down of LRRC8A. It was also suppressed by blockers of S1P receptor 1 and 2 (W146 and JTE013, respectively). RTqPCR demonstrated the prominent presence of S1PR1 and S1PR2 mRNAs. We discussed the roles of S1P-induced ATP release in the cancer microenvironment.
Highlights
ATP is quickly hydrolyzed by ecto-ATPase in the extracellular space and is maintained at a concentration near zero in the interstitial fluids of unstressed tissues
Life 2021, 11, 851 changed to a transient-sharp pattern with cholera toxin treatment, and the reverse change was induced by TGFβ treatment; DCPIB (4-[(2-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihyby TGFβ treatment; DCPIB (4-[(2-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1hdro-1-oxo-1h-inden-5-yl)oxy]butanoic acid), an inhibitor of volume-regulated anion chaninden-5-yl)oxy]butanoic acid), an inhibitor of volume-regulated anion channels (VRACs), nels (VRACs), only suppressed the diffuse pattern; knockdown of LRRC8A, the essential only suppressed the diffuse pattern; knockdown of LRRC8A, the essential molecular molecular entity of VRACs, suppressed the diffuse pattern (Figure 1). These results entity of VRACs, suppressed the diffuse pattern (Figure 1). These results suggest that suggest that abundantly expressed VRACs contribute to the release of ATP in undifferenabundantly expressed VRACs contribute to the release of ATP in undifferentiated cells tiated cellscancer including including cells.cancer cells
(4) Knockdown of LRRC8A, the essential molecular entity of VRACs, suppressed thepattern. These results suggest that abundantly expressed VRACs are a conduit of ATP release in undifferentiated cells, including pattern
Summary
ATP is quickly hydrolyzed by ecto-ATPase in the extracellular space and is maintained at a concentration near zero in the interstitial fluids of unstressed tissues. This provides a condition under which ATP works as a ubiquitous extracellular signaling molecule through its instantaneous release. One important function is the production of adenosine by hydrolysis with ecto-ATPase, CD39 and CD73 and the maintenance of a high level of adenosine in the cancer microenvironment. Despite increasing evidence to support the importance of purinergic pathways in the cancer microenvironment, the source of ATP, which is itself a major source of adenosine, remains unclear
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