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Distinct kinetics for nucleotide hydrolysis in lymphocytes isolated from blood, spleen and cervical lymph nodes: Characterization of ectonucleotidase activity
Author(s) -
Doleski Pedro H.,
Cabral Fernanda L.,
Adefegha Stephen A.,
Jantsch Matheus H.,
Ebone Renan S.,
Leal Daniela B. R.,
Schetinger Maria R. C.
Publication year - 2021
Publication title -
cell biochemistry and function
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.933
H-Index - 61
eISSN - 1099-0844
pISSN - 0263-6484
DOI - 10.1002/cbf.3616
Subject(s) - atp hydrolysis , nucleotide , enzyme , chemistry , adenosine , extracellular , adenosine triphosphate , spleen , lymph node , hydrolysis , adenosine monophosphate , biochemistry , adenosine diphosphate , lymph , microbiology and biotechnology , biology , immunology , atpase , medicine , gene , platelet , pathology , platelet aggregation
Abstract Ectonucleotidases are a plasma membrane‐bound enzyme that hydrolyses extracellular adenosine triphosphate (eATP) and adenosine diphosphate (eADP) to adenosine monophosphate (AMP). It regulates normal function of lymphocytes, acts as an inflammatory marker and represents a molecular target for new therapeutics. Thus, this study sought to isolate lymphocytes from blood (BL), spleen (SL) and cervical lymph node (CLL), and characterize the eATP and eADP enzymatic hydrolysis in Wistar rats. The hydrolysis of the nucleotides occurred primarily at pH 8.0, 37°C in the presence of Ca 2+ or Mg 2+ . Chevillard‐plot showed the hydrolysis of eATP and eADP at the same active site. The inhibitors of some classical ATDPases did not cause any significant change on enzymatic activity. Inhibitors of E‐NTPDase (‐1, ‐2, ‐3 isoforms) and E‐NPP‐1 decrease the enzyme activity in all resident lymphocytes. Furthermore, kinetic parameters (Vmax and Km) revealed that SL had significantly ( P < .001) higher enzymatic activity when compared to BL and CLL. In conclusion, this study standardized kinetic values for eATP and eADP hydrolysis for resident lymphocytes isolated from BL, SL and CLL.