Self-regulation of inflammatory cell trafficking in mice by the leukocyte surface apyrase CD39

Abstract

Leukocyte and platelet accumulation at sites of cerebral ischemia exacerbate cerebral damage. The ectoenzyme CD39 on the plasmalemma of endothelial cells metabolizes ADP to suppress platelet accumulation in the ischemic brain. However, the role of leukocyte surface CD39 in regulating monocyte and neutrophil trafficking in this setting is not known. Here we have demonstrated in mice what we believe to be a novel mechanism by which CD39 on monocytes and neutrophils regulates their own sequestration into ischemic cerebral tissue, by catabolizing nucleotides released by injured cells, thereby inhibiting their chemotaxis, adhesion, and transmigration. Bone marrow reconstitution and provision of an apyrase, an enzyme that hydrolyzes nucleoside tri-and diphosphates, each normalized ischemic leukosequestration and cerebral infarction in CD39-deficient mice. Leukocytes purified from Cd39 -/-mice had a markedly diminished capacity to phosphohydrolyze adenine nucleotides and regulate platelet reactivity, suggesting that leukocyte ectoapyrases modulate the ambient vascular nucleotide milieu. Dissipation of ATP by CD39 reduced P2X 7 receptor stimulation and thereby suppressed baseline leukocyte α M β 2 -integrin expression. As α M β 2 -integrin blockade reversed the postischemic, inflammatory phenotype of Cd39 -/-mice, these data suggest that phosphohydrolytic activity on the leukocyte surface suppresses cell-cell interactions that would otherwise promote thrombosis or inflammation. These studies indicate that CD39 on both endothelial cells and leukocytes reduces inflammatory cell trafficking and platelet reactivity, with a consequent reduction in tissue injury following cerebral ischemic challenge.

Liver damage and systemic inflammatory responses are exacerbated by the genetic deletion of CD39 in total hepatic ischemia

“…Hyman et al have recently shown that CD39 on both endothelial cells and leukocytes reduces inflammatory cell trafficking and platelet reactivity following cerebral ischemia [31]. CD39 is also protective in cerebrovascular ischemia [32] and soluble CD39 infusion can restore postischemic cerebral perfusion [32].…”

“…31 P NMR All experiments were performed at a 31 P Frequency of 145.7 MHz (360 MHz for 1 H) on a, 9-cm vertical bore, Bruker DRX (Burker Biospin Inc. Bellerica, MA) spectrometer. Mice were anesthetized with a mixture of Ketamine/ Xylazine (100 mg and 10 mg/kg) and the liver was surgically exposed.…”

“…31 P spectra were displayed at the same vertical scale and calibrated so that chemical shifts of the other metabolites can be visualized with respect to the phosphocreatine peak Soluble NTPDase treatment Mice that were randomly assigned to NTPDase treatment groups received a single intravenous injection of soluble grade VII NTPDase (0.2 units/g bodyweight) of a 20 units/ml stock solution in saline (apyrase, Sigma, St. Louis, MO, USA) or adenosine (Sigma, St. Louis, MO, USA) 1 mmol/kg/min and amrinone (Sigma, St. Louis, MO, USA) 0.05 mmol/kg/min for 60 min prior to reperfusion. Controls were injected with an equivalent volume of saline.…”

“…31 P spectra from the left lateral mouse liver lobe were tested at three time points (top) pre-ischemia, (middle) 45 min of ischemia, and (bottom) after 30-60 min of reperfusion (Fig. 4).…”

“…5 Hepatic reperfusion injury: NMR -P31 spectra with ATP depletion in the wild-type mouse. 31 P spectra from the left lateral mouse liver lobe at the three different time frames (top) pre-ischemia, (middle) 45 min of ischemia, and (bottom) 30-60 min of reperfusion. The intensity of the three peaks due to the abg ATP is reduced during ischemia and recovery in the wild-type upon reperfusion.…”