title: Apyrase activity and adenosine diphosphate induced platelet aggregation inhibition by the salivary gland proteins of Culicoides variipennis, the North American vector of bluetongue viruses.
abstract: Salivary gland homogenates of Culicoides variipennis, the primary vector of bluetongue (BLU) viruses in North America, were analyzed for apyrase activity. Apyrase (ATP diphosphohydrolase, EC 3.6.1.5) is an anti-hemostatic and anti-inflammatory salivary enzyme of most hematophagous arthropods. The enzyme activity was measured by the release of orthophosphate using ATP, ADP, and AMP as substrates with Ca2+ as the divalent cation. ATPase (11.5 +/- 1 mU/pair of glands), ADPase (7.3 +/- 0.7 mU/pair of glands), and insignificant (P < 0.05) AMPase (0.07 mU/pair of glands) activities were detected in female salivary glands. Male salivary glands contained lower amounts of ATPase and ADPase activity (P < 0.05). The ATPase and ADPase activities were greatest at pH 8.5, and were similarly activated by Mg2+. Molecular sieving HPLC of salivary gland homogenates generated a single peak which coincided with ATPase and ADPase, but no AMPase, activity; the protein has an estimated molecular mass of 35,000 Da. ATPase and ADPase activity, and total protein concentration, were reduced (P < 0.05) in the salivary glands of females after taking a blood meal from a sheep. Salivary gland homogenates also inhibited ADP-induced platelet aggregation in vitro. It is concluded that the salivary ATPase and ADPase activities of C. variipennis reside in one enzyme, and that this enzyme is likely an apyrase. The apyrase activity is thought to be responsible for the inhibition of ADP-induced platelet aggregation, as indicated by the apparent discharge of apyrase from salivary glands into the host during blood feeding. This suggests that apyrase is one of the salivary proteins present in C. variipennis acting as antigens in the development of Culicoides hypersensitivity in ruminants and horses. Apyrase may inhibit an inflammatory response at the feeding site through the subsequent degradation of its end-product, AMP, to adenosine, a potent anti-inflammatory substance, by the ecto-5' nucleotidase activity of neutrophils.
authors.affiliation: Arthropod-borne Animal Diseases Research Laboratory, Agricultural Research Service, USDA, Laramie, WY 82071-3965, USA.
authors.forename: A A
authors.initials: AA
authors.lastname: Pérez de León
authors.forename: W J
authors.initials: WJ
authors.lastname: Tabachnick
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D001081:Apyrase / Q000378:metabolism*; D001819:Bluetongue / Q000635:transmission; D001820:Bluetongue virus; D002519:Ceratopogonidae / Q000821:virology; D005260:Female; D007303:Insect Vectors; D007700:Kinetics; D009656:North America; D010974:Platelet Aggregation / Q000187:drug effects / Q000502:physiology*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D012469:Salivary Glands / Q000201:enzymology*; D012471:Salivary Proteins and Peptides / Q000302:isolation & purification / Q000494:pharmacology*
chemical_list: D010975:Platelet Aggregation Inhibitors; D012471:Salivary Proteins and Peptides; D000244:Adenosine Diphosphate; D000251:Adenosine Triphosphatases; D001081:Apyrase