Serotonin metabolism in rat mesangial cells: involvement of a serotonin transporter and monoamine oxidase A.
BACKGROUND
Serotonin is one of the factors regulating mesangial cell proliferation, and convergent evidence supports its involvement in the development of glomerulonephritis. In this study, we identified a serotonin transporter and the amine-degrading enzyme monoamine oxidases (MAOs) in mesangial cells, and we studied their involvement in serotonin degradation.


METHODS
MAOs were characterized in membrane preparations and intact mesangial cells by enzyme assay using [14C]5-hydroxytryptamine and [14C]beta-phenylethylamine as specific substrates for MAO-A and MAO-B, respectively, and by Western blot analysis. The expression of a serotonin transporter was determined by [14C]5-hydroxytryptamine uptake experiments and Western blot. Mesangial cell proliferation was measured by BrdU incorporation.


RESULTS
Quantitation of the MAO isoforms by enzyme assay and Western blot analysis showed that MAO-A was largely predominant in mesangial cells, accounting for approximately 90% of the total enzyme population. The MAO substrate [14C]serotonin was transported into mesangial cells by a saturable uptake system (Vmax 310 +/- 36 pmol/30 min/mg protein; Km 5.9 +/- 1.4 microM) displaying the pharmacological properties of a serotonin transporter. The expression of a serotonin transporter was confirmed by Western blot analysis. MAO activity measured in intact cells showed that after accumulation into mesangial cells, [14C]serotonin was metabolized by MAO-A. Finally, serotonin-mediated mesangial cell proliferation was significantly increased after irreversible MAO inhibition.


CONCLUSIONS
Our results suggest that serotonin concentration and function in glomeruli may be regulated in part by its transport into mesangial cells and degradation by MAO-A.
Pizzinat
N
N
Institut National de la Santé et de la Recherche Médicale U388, Institut Louis Bugnard, Centre Hospitalier Universitaire Rangueil, Toulouse, France.
Girolami
J P
JP
Parini
A
A
Pecher
C
C
Ordener
C
C
D000595:Amino Acid Sequence; D000818:Animals; D000963:Antimetabolites / Q000378:metabolism / Q000494:pharmacology; D001692:Biological Transport / Q000502:physiology; D015153:Blotting, Western; D001973:Bromodeoxyuridine / Q000378:metabolism / Q000494:pharmacology; D002250:Carbon Radioisotopes; D002352:Carrier Proteins / Q000378:metabolism*; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000502:physiology; D005920:Glomerular Mesangium / Q000737:chemistry* / Q000166:cytology / Q000201:enzymology*; D007672:Kidney Cortex / Q000737:chemistry / Q000166:cytology / Q000201:enzymology; D008562:Membrane Glycoproteins / Q000378:metabolism*; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D008995:Monoamine Oxidase / Q000032:analysis / Q000378:metabolism*; D008996:Monoamine Oxidase Inhibitors / Q000494:pharmacology; D009419:Nerve Tissue Proteins; D010293:Pargyline / Q000494:pharmacology; D051381:Rats; D012701:Serotonin / Q000378:metabolism* / Q000493:pharmacokinetics; D050486:Serotonin Plasma Membrane Transport Proteins
D000963:Antimetabolites; D002250:Carbon Radioisotopes; D002352:Carrier Proteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D008996:Monoamine Oxidase Inhibitors; D009419:Nerve Tissue Proteins; D050486:Serotonin Plasma Membrane Transport Proteins; C493232:Slc6a4 protein, rat; D012701:Serotonin; D010293:Pargyline; D008995:Monoamine Oxidase; D001973:Bromodeoxyuridine