Generation of a novel antibody probe to the apical sodium-dependent bile acid transporter that inhibits ileal bile acid absorption.
Intestinal bile acid absorption is mediated by a sodium-dependent transporter located in the brush border apical membrane of ileocytes. The transmembrane topology and the role of individual amino acid residues in the bile acid transport process have been investigated by means of various experimental approaches, leading to multiple hypotheses. We raised a monoclonal antibody against a segment of the transporter comprising vicinal cysteine residues, in order to evaluate its functional role. A 14 amino acid peptide, corresponding to amino acids 104-117 of the transporter, was synthesized, and a monoclonal anti-peptide antibody was raised. In vitro uptake-inhibition studies in the presence of the monoclonal anti-peptide antibody were performed using ileal brush border membrane vesicles. Rabbit ileum was perfused in vivo with 5 mM taurocholic acid in the presence of the monoclonal antibody, and bile acid absorption inhibition was evaluated. The anti-peptide monoclonal antibody significantly reduced the in vitro uptake and in vivo absorption of taurocholic acid. The present data demonstrate the functional relevance of the 104-117 peptide segment and report the generation of a novel antibody against the apical sodium-dependent bile acid transporter (ASBT) that may be used as a therapeutic agent in hypercholesterolemia and in cholestatic pruritus.
Department of Internal Medicine and Gastroenterology, University of Bologna, Bologna, Italy.
M
M
Montagnani
A
A
Marangoni
A
A
Roda
F
F
Azzaroli
G
G
Mazzella
E
E
Roda
M
M
Tsivian
F
F
Neri
M
M
Jovani
M
M
Giandinoto
A
A
Caponi
R
R
Aldini
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology*; D001647:Bile Acids and Salts / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D007082:Ileum / Q000187:drug effects* / Q000378:metabolism*; D029363:Organic Anion Transporters, Sodium-Dependent / Q000737:chemistry / Q000276:immunology* / Q000502:physiology*; D011817:Rabbits; D027981:Symporters / Q000737:chemistry / Q000276:immunology* / Q000502:physiology*
D000911:Antibodies, Monoclonal; D001647:Bile Acids and Salts; D029363:Organic Anion Transporters, Sodium-Dependent; D027981:Symporters; C071908:sodium-bile acid cotransporter