Nucleotides released by apoptotic cells act as a find-me signal to promote phagocytic clearance.
Phagocytic removal of apoptotic cells occurs efficiently in vivo such that even in tissues with significant apoptosis, very few apoptotic cells are detectable. This is thought to be due to the release of 'find-me' signals by apoptotic cells that recruit motile phagocytes such as monocytes, macrophages and dendritic cells, leading to the prompt clearance of the dying cells. However, the identity and in vivo relevance of such find-me signals are not well understood. Here, through several lines of evidence, we identify extracellular nucleotides as a critical apoptotic cell find-me signal. We demonstrate the caspase-dependent release of ATP and UTP (in equimolar quantities) during the early stages of apoptosis by primary thymocytes and cell lines. Purified nucleotides at these concentrations were sufficient to induce monocyte recruitment comparable to that of apoptotic cell supernatants. Enzymatic removal of ATP and UTP (by apyrase or the expression of ectopic CD39) abrogated the ability of apoptotic cell supernatants to recruit monocytes in vitro and in vivo. We then identified the ATP/UTP receptor P2Y(2) as a critical sensor of nucleotides released by apoptotic cells using RNA interference-mediated depletion studies in monocytes, and macrophages from P2Y(2)-null mice. The relevance of nucleotides in apoptotic cell clearance in vivo was revealed by two approaches. First, in a murine air-pouch model, apoptotic cell supernatants induced a threefold greater recruitment of monocytes and macrophages than supernatants from healthy cells did; this recruitment was abolished by depletion of nucleotides and was significantly decreased in P2Y(2)(-/-) (also known as P2ry2(-/-)) mice. Second, clearance of apoptotic thymocytes was significantly impaired by either depletion of nucleotides or interference with P2Y receptor function (by pharmacological inhibition or in P2Y(2)(-/-) mice). These results identify nucleotides as a critical find-me cue released by apoptotic cells to promote P2Y(2)-dependent recruitment of phagocytes, and provide evidence for a clear relationship between a find-me signal and efficient corpse clearance in vivo.
Elliott
Michael R
MR
Beirne B. Carter Center for Immunology Research, University of Virginia, Charlottesville, Virginia 22908, USA.
Chekeni
Faraaz B
FB
Trampont
Paul C
PC
Lazarowski
Eduardo R
ER
Kadl
Alexandra
A
Walk
Scott F
SF
Park
Daeho
D
Woodson
Robin I
RI
Ostankovich
Marina
M
Sharma
Poonam
P
Lysiak
Jeffrey J
JJ
Harden
T Kendall
TK
Leitinger
Norbert
N
Ravichandran
Kodi S
KS
D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002460:Cell Line; D002478:Cells, Cultured; D002630:Chemotactic Factors / Q000378:metabolism / Q000494:pharmacology; D002633:Chemotaxis / Q000187:drug effects; D017077:Culture Media, Conditioned / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D019169:Jurkat Cells; D008262:Macrophage Activation / Q000187:drug effects; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009000:Monocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D010586:Phagocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D010587:Phagocytosis / Q000187:drug effects / Q000502:physiology*; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D058473:Receptors, Purinergic P2Y2; D015398:Signal Transduction / Q000187:drug effects; D013950:Thymus Gland / Q000166:cytology*; D014544:Uridine Triphosphate / Q000378:metabolism* / Q000494:pharmacology
D002630:Chemotactic Factors; D017077:Culture Media, Conditioned; C546385:P2RY2 protein, human; C546386:P2ry2 protein, mouse; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2; D058473:Receptors, Purinergic P2Y2; D000255:Adenosine Triphosphate; D014544:Uridine Triphosphate