TY - JOUR
T1 - Generation of a novel antibody probe to the apical sodium-dependent bile acid transporter that inhibits ileal bile acid absorption
AU - Montagnani, M.
AU - Marangoni, A.
AU - Roda, A.
AU - Azzaroli, F.
AU - Mazzella, G.
AU - Roda, E.
AU - Tsivian, M.
AU - Neri, F.
AU - Jovani, M.
AU - Giandinoto, M.
AU - Caponi, A.
AU - Aldini, Rita
PY - 2009/6/1
Y1 - 2009/6/1
N2 - 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.
AB - 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.
KW - Apical sodium-dependent bile acid transporter (ASBT)
KW - Brush border membrane vescicles (BBMV)
KW - Cholic acid (CA)
KW - Liver sodium taurocholate transport protein (NTCP)
KW - Taurocholic acid (TCA)
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U2 - 10.1021/mp800117b
DO - 10.1021/mp800117b
M3 - Article
C2 - 19366255
AN - SCOPUS:67249132776
VL - 6
SP - 1012
EP - 1018
JO - Molecular Pharmaceutics
JF - Molecular Pharmaceutics
SN - 1543-8384
IS - 3
ER -