Regulation of endothelial CD73 by adenosine: paracrine pathway for enhanced endothelial barrier function.
During episodes of inflammation, multiple cell types release adenine nucleotides in the form of ATP, ADP, 5'-AMP, and adenosine. In particular, following activation, polymorphonuclear leukocytes release larger quantities of 5'-AMP. Extracellular 5'-AMP is metabolized to adenosine by surface-expressed 5'-ectonucleotidase (CD73). Adenosine liberated by this process activates surface adenosine A(2B) receptors, results in endothelial junctional reorganization, and promotes barrier function. We hypothesized that adenosine signaling to endothelia provides a paracrine loop for regulated expression of CD73 and enhanced endothelial barrier function. Using an in vitro microvascular endothelial model, we investigated the influence of 5'-AMP; adenosine; and adenosine analogues on CD73 transcription, surface expression, and function. Initial experiments revealed that adenosine and adenosine analogues induce CD73 mRNA (RT-PCR), surface expression (immunoprecipitation of surface biotinylated CD73), and function (HPLC analysis of etheno-AMP conversion to ethenoadenosine) in a time- and concentration-dependent fashion. Subsequent studies revealed that similar exposure conditions increase surface protein through transcriptional induction of CD73. Analysis of DNA-binding activity by EMSA identified a functional role for CD73 cAMP response element and, moreover, indicated that multiple cAMP agonists induce transcriptional activation of functional CD73. Induced CD73 functioned to enhance 5'-AMP-mediated promotion of endothelial barrier (measured as a paracellular flux of 70-kDa FITC-labeled tracer). These results provide an example of transcriptional induction of enzyme (CD73) by enzymatic product (adenosine) and define a paracrine pathway for the regulated expression of vascular endothelial CD73 and barrier function.
Narravula
S
S
Center for Experimental Therapeutics and Reperfusion Injury, Brigham and Women's Hospital, and Division of Hematology and Oncology, Beth Israel-Deaconess Medical Center and Harvard Medical School, Boston, MA 02115, USA.
Lennon
P F
PF
Mueller
B U
BU
Colgan
S P
SP
D015720:5'-Nucleotidase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D000241:Adenosine / Q000031:analogs & derivatives / Q000378:metabolism / Q000502:physiology*; D000328:Adult; D002199:Capillary Permeability / Q000502:physiology*; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000819:agonists / Q000378:metabolism / Q000502:physiology; D020405:Dermis / Q000098:blood supply; D004730:Endothelium, Vascular / Q000201:enzymology* / Q000378:metabolism / Q000502:physiology; D004789:Enzyme Activation; D006801:Humans; D019899:Paracrine Communication / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism; D018047:Receptors, Purinergic P1 / Q000378:metabolism; D015533:Transcriptional Activation
D012333:RNA, Messenger; D018047:Receptors, Purinergic P1; D000242:Cyclic AMP; D015720:5'-Nucleotidase; D000241:Adenosine