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Spilanthol from Acmella oleracea lowers Na + ‐K + ‐2Cl − ‐cotransporter activity and water channel aquaporin 2 membrane expression in mouse renal cells
Author(s) -
Gerbino Andrea,
Schena Giorgia,
Milella Luigi,
Miglionico Rocchina,
Armentano Mariafrancesca,
Barbosa Alan Franco,
Procino Giuseppe,
Svelto Maria,
Carmosino Monica
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.959.4
Subject(s) - chemistry , apical membrane , intracellular , aquaporin 2 , cotransporter , kidney , diuretic , endocrinology , medicine , biochemistry , sodium , membrane , biology , mechanical engineering , organic chemistry , water channel , engineering , inlet
Objective Acmella oleracea is well recognized in Brazilian traditional medicine as diuretic, although few scientific data have been published to support this effect. Aim of this study was to determine the molecular effect of Acmella oleracea extract and its main alkylamide spilanthol on two major processes involved in the urine concentrating mechanism: Na‐K‐2Cl symporter (NKCC2) activity in the thick ascending limb and water channel aquaporin 2 accumulation at the apical plasma membrane of collecting duct cells. Methods Phosphorylation of NKCC2 was evaluated as index of its activation by Western blotting. Rate of aquaporin 2 apical expression was analysed by confocal laser microscopy. Spilanthol‐induced intracellular signalling events were dissected by video‐imaging experiments. Results Exposure to spilanthol reduced the basal phosphorylation level of NKCC2 both in freshly isolated mouse kidney slices and in NKCC2‐expresing HEK293 cells. Spilanthol‐induced effect on NKCC2 activity was significantly larger than that exerted by the methanol plant extract. In addition, exposure to spilanthol strongly reduced both desmopressin and low Cl − ‐dependent increase in NKCC2 phosphorylation in mouse kidney slices and NKCC2‐expressing HEK293 cells, respectively. Similarly, exposure to spilanthol reduced both desmopressin‐ and forskolin‐stimulated aquaporin 2 accumulation at the apical plasma membrane of collecting duct in mouse kidney slice and MCD4 cells. Finally, at cellular level, spilanthol rapidly reduced or reversed basal and agonist‐increased cAMP levels through a mechanism involving increases in intracellular [Ca 2+ ]. Conclusions Spilanthol‐induced inhibition of cAMP production negatively modulates urine‐concentrating mechanisms thus holding great promise for its use as both diuretic and aquaretic. Support or Funding Information This work was supported by the Ricerca di interesse locale RIL to M. Carmosino, University of Basilicata and by Fondo per gli Investimenti della Ricerca di Base‐Rete Nazionale di Proteomica to M. Svelto (grant number RBRN07BMCT_009).