Over-expression of neuropathy target esterase activity in bovine chromaffin cell cultures by adenovirus-mediated gene transfer.
Chromaffin cells in culture show high neuropathy target esterase (NTE) activity. It is well known that inhibition and specific modification of NTE by some organophosphorus (OPs) compounds induces a neurodegenerative neuropathy. It has been suggested that NTE is responsible for phosphatidylcholine homeostasis, although its role in neuropathy induction remains unclear. The cDNA of human NTE (4.4kbp) was inserted into an adenoviral vector. Bovine chromaffin cells cultured at 50,000 cells/well were incubated with the vector for 2h and after removing the volume of infection, cells were maintained in the incubator. After 24h, NTE activity was 6.8+/-0.5mU/10(6) cells in untreated cells and 14.8+/-1.5mU/10(6) cells, 19.3+/-2.9mU/10(6) cells, 24.8+/-0.9mU/10(6) cells and 30.9+/-1.0mU/10(6) cells in cells incubated with 2, 4, 8 and 16microl of vector, respectively. After 60min of inhibition with mipafox increased concentrations, the calculated I(50) (60min) values were 5.5, 6.2 and 6.6microM for cells infected with 0, 2 and 10microl of vector preparation. We confirm that the adenoviral vector containing the human NTE gene is active in bovine chromaffin cells in culture and that the NTE activity expressed by the vector shows the same inhibition pattern by the neuropathic OP mipafox as the NTE activity of bovine chromaffin cells and cells remained viable after the high NTE activity expression.
Quesada
Encarna
E
Unidad de Toxicología y Seguridad Química, Instituto de Bioingeniería, Universidad Miguel Hernández. Avda. de la Universidad, s/n. E-03202 Elche (Alicante), Spain. equesada@umh.es
Castell
José Vicente
JV
Vilanova
Eugenio
E
Carrera
Victoria
V
D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D002265:Carboxylic Ester Hydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000201:enzymology*; D005822:Genetic Vectors; D006801:Humans; D007531:Isoflurophate / Q000031:analogs & derivatives / Q000633:toxicity; D010261:Paraoxon / Q000633:toxicity; D014162:Transfection; D014631:Valerates / Q000378:metabolism
D014631:Valerates; D007531:Isoflurophate; C049867:phenyl valerate; C005238:mipafox; D002265:Carboxylic Ester Hydrolases; C021859:neurotoxic esterase; D010261:Paraoxon