Аннотация
In this study, we compared the pharmacological and biochemical characteristics
of A(2B) adenosine receptors in recombinant (hA(2B)HEK293 cells)
and native cells (neutrophils, lymphocytes) by using a new potent
8-pyrazole xanthine derivative, (3)HN-benzo1,3dioxol-5-yl-2-5-(1,3-dipropyl-2,6-dioxo-2,3,6,7-tetrahy
dro-1H-purin-8-yl)-1-methyl-1H-pyrazol-3-yl-oxy-acetamide ((3)HMRE
2029-F20), that has high affinity and selectivity for hA(2B) versus
hA(1),hA(2A), and hA(3) subtypes. (3)HMRE 2029-F20 bound specifically
to the hA(2B) receptor stably transfected in human embryonic kidney
(HEK) 293 cells with K(D) of 2.8 +/- 0.2 nM and B(max) of 450 +/-
42 fmol/mg of protein. Saturation experiments of (3)HMRE 2029-F20
binding in human neutrophils and lymphocytes detected a single high-affinity
binding site with K(D) values of 2.4 +/- 0.5 and 2.7 +/- 0.7 nM,
respectively, and B(max) values of 79 +/- 10 and 54 +/- 8 fmol/mg
of protein, respectively, in agreement with real-time reverse transcription
polymerase chain reaction studies showing the presence of A(2B) mRNA.
The rank order of potency of typical adenosine ligands with recombinant
hA(2B) receptors was consistent with that typically found for interactions
with the A(2B) subtype and was also similar in peripheral blood cells.
5'-N-Ethyl-carboxamidoadenosine stimulated cAMP accumulation in both
hA(2B)HEK293 and native cells, whereas phospholipase C activation
was observed in recombinant receptors and endogenous subtypes expressed
in neutrophils but not in lymphocytes. MRE 2029-F20 was revealed
to be a potent antagonist in counteracting the agonist effect in
both signal transduction pathways. In conclusion, (3)HMRE 2029-F20
is a selective and high-affinity radioligand for the hA(2B) adenosine
subtype and may be used to quantify A(2B) endogenous receptors. In
this work, we demonstrated their presence and functional coupling
in neutrophils and lymphocytes that play a role in inflammatory processes
in which A(2B) receptors may be involved.
Линки и ресурсы
тэги