P2Y11 Purinoceptor Mediates the ATP-Enhanced Chemotactic Response of Rat Neutrophils

Abstract

Abstract. ATP and hydrolysis products of ATP like adenosine regulate the chemotaxis of neutrophils by activating purinoceptors and adenosine receptors. The present study was designed to examine exogenous ATP, activation of purinoceptors, and activation of A 3 adenosine receptor as key steps in the signal cascades that control cell orientation and migration of rat neutrophils. One or more of those steps might be potential therapeutic targets for treatment of inflammatory diseases. The chemotaxis of rat neutrophils was stimulated with N-formyl-methionyl-leucyl-phenylalanine (fMLP) and measured with an EZ-TAXIScan apparatus. The effects of apyrase, exogenous ATP, suramin (P2X and P2Y blocker), PPADS (a P2X blocker), TNP-ATP (P2X 1 and P2X 3 antagonist), and Reactive Blue 2 (a P2Y blocker) on the chemotactic response were also investigated. Rat neutrophil chemotaxis was significantly suppressed by apyrase. fMLP induced rat neutrophil chemotaxis was potentiated by ATP, blocked by suramin, not affected by PPADS or TNP-ATP, and significantly inhibited by RB-2. Western blotting showed that A 3 , P2Y 2 , and P2Y 11 were expressed in rat neutrophils. The chemotactic response of rat neutrophils to fMLP stimulation is potentiated by ATP via P2Y 11 purinoceptors but not P2X purinoceptors or A 3 adenosine receptor, and that the response plays a critical role in host defense and pathogenicity.[Supplementary Movies: available only at http://dx

Role of Spinal P2Y6and P2Y11Receptors in Neuropathic Pain in Rats: Possible Involvement of Glial Cells

“…Currently there are no reports of a cloned P2Y 11 receptor in rats. However, this receptor has been recently reported in epithelial cells [36], primary glial cells [37], lacrimal glands [38], neutrophils [39], and brain [38] of rats. Thus, our data suggest that the P2Y 6 receptor is expressed and completely functional in the spinal cord.…”

The growth factor progranulin attenuates neuronal injury induced by cerebral ischemia-reperfusion through the suppression of neutrophil recruitment

“…The neutrophils were then pelleted by centrifugation (800 × g for 10 minutes) and washed twice with 5 mL of PBS. This fraction, which contained over 95% viable neutrophils, was used for the following experiment [22]. 125 Iodine-labeled recombinant human TNF-α was purchased from PerkinElmer, Inc. (Boston, MA, USA; specific activity 2.8 Bq/pg).…”

“…Screening for neutrophil chemotaxis was performed using an EZ-TAXIScan chemotaxis apparatus (Effector Cell Institute, Kawasaki, Japan), as has been previously described [22]. Briefly, neutrophils were stimulated by the addition TNF-α (100 ng/ mL; Recombinant Mouse TNF-α; R&D Systems, Inc.) and different amounts of r-PGRN (between 0 and 250 ng/mL; R&D Systems, Inc.) to one hole, while treated cells were placed in a contra-hole of the microchannel.…”

A critical look at the function of the P2Y11 receptor

“…Other antibodies have also been used to examine the existence of the murine P2Y 11 . The P2Y 11 antibody #AB9590 targeting the human P2Y 11 C-terminus detects a large, 75-kDa band on western blot, which is believed to be P2Y 11 on the basis of the results of experiments using rat tissue [41,42], and it has been claimed that another antibody with an unknown epitope confirms P2Y 11 receptor expression in rat neutrophils with a 40-kDa signal [43].…”

“…It is also worth noting that NF157, NF340, and NF546 have all been used to study the murine BP2Y 11 -like receptort hat shows many of the same properties as seen in humans [14,[41][42][43][82][83][84]. Assuming that no P2Y 11 receptor exists in murines, these compounds must have other mechanisms by which they interfere with cell signalling.…”