Flow cytometric analysis of the in situ hybridization of cyclooxygenase isoforms in mesangial cells treated with cyclosporine A.
BACKGROUND
Cyclosporine A increases oxidative stress in kidney and we hypothesized that cyclooxygenase (COX) may be involved in this effect.


METHODS
Mesangial cells of Cyclosporine A-treated (4, 7 or 10 days) rats were obtained to evaluate mRNA expression of COX-isoforms (COX-1, constitutive and COX-2, inducible) by "in situ" hybridization. Probes were labelled using "Gene Image Random Prime Labelling Protocol" and COX expression was measured by flow cytometry.


CONCLUSIONS
"In situ" hybridization by flow cytometry is an useful method to detect mRNA. We observed an increased COX-2 expression in a time-dependent manner in parallel with Reactive Oxygen Species synthesis. COX-1 expression increased only at 10 days.
Pérez de Hornedo
J
J
Unidad de Investigación, Hospital Universitario de Guadalajara, Spain.
Calvino Fernández
M
M
de Arriba de la Fuente
G
G
del Reino Fernández
P
P
Parra Cid
T
T
Benito Martinez
S
S
D000818:Animals; D051545:Cyclooxygenase 1 / Q000235:genetics; D051546:Cyclooxygenase 2 / Q000235:genetics; D016572:Cyclosporine / Q000494:pharmacology*; D005434:Flow Cytometry / Q000379:methods*; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D007166:Immunosuppressive Agents / Q000494:pharmacology; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D007527:Isoenzymes / Q000235:genetics; D008297:Male; D050527:Mesangial Cells / Q000187:drug effects* / Q000378:metabolism; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics*; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism
D007166:Immunosuppressive Agents; D007527:Isoenzymes; D017382:Reactive Oxygen Species; D016572:Cyclosporine; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases