Regulated Extracellular Nucleotide Metabolism and Function at the Mucosa - Extracellular ATP and Adenosine as Regulators of Endothelial Cell Function - page 123

ATP over ADP. NTPDase2 has been identified for acting almost exclusively to
hydrolyze ATP [51, 62]. Together, these NTPDases act in a concerted manner to
regulate the production of extracellular adenosine monophosphate (AMP).
The final step in generation of extracellular adenosine is conferred by the gly-
cosyl phosphatidylinositol (GPI)-anchored membrane protein, ecto-5 nucleotidase
(CD73). CD73 is the predominant source for accumulation of extracellular adeno-
sine from released adenine nucleotides [75]. CD73 metabolizes AMP to adenosine,
which is then either free to act as a ligand to one of four adenosine receptors (A1,
A2A, A2B, A3), or transported into the cell by dipyridamole-sensetive channels
and degraded by the purine salvage pathway. As with the NTPDases, CD73 is a
widely distributed cell surface enzyme, whose expression has been shown across a
variety of tissues. It has been demonstrated that the AMP hydrolyzing activity of
CD73 varies widely from tissue to tissue, with the rank order of tissue activity as
follows: colon > kidney brain > liver > lung > heart >> muscle [69]. As high

levels of activity are reported at a number of mucosal sites, it is important to under-
stand the physiological role played at this terminal step of the nucleotide hydrolysis
pathway.

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8.3 Physiological and Pathophysiologic Influences of Nucleotide
Metabolites

Mucosal tissues such as the kidney, lung and intestine are lined with an epithelial
layer that serves as both a barrier to the external environment and a conduit for nutri-
ents, ions and gas exchange. During mucosal inflammation, the major sources of
extracellular nucleotides are considered to be platelet- and leukocyte-derived [44].
Of particular interest is the regulated release of adenine nucleotides by polymor-
phonuclear cells (PMN, neutrophils) as they cross the endothelium and epithelium
to a site of infection and/or inflammation. A number of years ago, it was appreciated
that when interacting with endothelia and epithelia, PMN actively release adenine
nucleotides. Original studies by Madara et al. examining biological properties of
soluble mediators derived from activated inflammatory cells (e.g. neutrophils and
eosinophils) identified a small, protease-resistant fraction termed neutrophil-derived
secretagogue (NDS), which when incubated on epithelia, activated electrogenic
chloride secretion and fluid transport (responsible for the hydration of mucosal
surfaces). Subsequent biophysical analysis of NDS identified this molecule to be
AMP [42]. With no known AMP receptor, studies turned toward defining poten-
tial metabolic pathways for adenosine generation. Biochemical and pharmacologic
studies demonstrated the polarized expression of CD73 on the apical surface of
cultured and primary intestinal epithelial cells [63]. Further biochemical and mor-
phological studies revealed that CD73 exists in both a GPI-linked surface fraction as
well as in a sub-apical caveolin-rich domain within the epithelium. Such expression
patterns have subsequently been shown in a variety of mucosal epithelial cell types.
Subsequent studies have addressed the role of adenine nucleotides in the regula-
tion of tissue barrier function. Indeed, as PMN cross both epithelial and endothelial