Identification and characterization of CD39/vascular ATP diphosphohydrolase

Area of Science:

• Biochemistry
• Molecular Biology
• Immunology

Background:

• Vascular ATP diphosphohydrolase (ATPDase) is an enzyme on the plasma membrane that breaks down extracellular ATP and ADP into AMP.
• Mammalian and avian ATPDases show homology with CD39, a marker for B-cell activation.

Purpose of the Study:

• To isolate and characterize human endothelial CD39.
• To determine if CD39 is identical to vascular ATPDase.
• To investigate the role of ATPDase in platelet aggregation and its response to oxidative stress.

Main Methods:

• CD39 cDNA isolation from human endothelial cells and expression in COS-7 cells.
• Immunological and functional assays to compare expressed CD39 with vascular ATPDase.
• Assessment of ATPDase activity inhibition of platelet aggregation.
• Analysis of ATPDase mRNA expression in various human tissues.
• Identification of conserved motifs within ATPDase sequences.

Main Results:

• Human endothelial CD39 demonstrated immunological identity and functional characteristics of vascular ATPDase.
• ATPDase effectively inhibited platelet aggregation induced by ADP, collagen, and thrombin.
• This inhibitory activity was lost in transfected COS-7 cells upon exposure to oxidative stress.
• ATPDase mRNA was detected in placenta, lung, skeletal muscle, kidney, and heart, but not in the brain.
• A conserved motif, DLGGASTQ, potentially crucial for nucleotide binding and activity, was identified.

Conclusions:

• Human CD39 is the vascular ATPDase, playing a role in regulating platelet aggregation.
• The loss of ATPDase activity under oxidative stress is a key finding.
• Overexpression of ATPDase may offer a strategy to prevent platelet activation in vascular inflammation.