Effect of LDL-Ox on genes related to inflammatory process in macrophages

ESTEVE RAFOLS MONTSERRAT; LEDDA ANGELO; GRASSA MARÍA DEL MAR; TOLEDO JUAN; GARDA HORACIO; LAURA MANSILLA; TARRAGA WILSON ALBERTO; GONZALEZ MARINA CECILIA

La Plata

Congreso; I Congreso Internacional de la Facultad de Ciencias Médicas; 2013

Facultad de Ciencias Médicas de La Plata

Isolated LDL fraction from human plasmas was peroxidized in vitro with Cu++ (5 µM) for 4 h (low, L), 8 h (medium, M) and 24 h (high, H) and dialyzed overnight to wash-out the copper ions. Three types of oxidized LDL were obtained. Percentage of dead cells -evaluated by trypan dye exclusion- compared to control flasks incubated with native LDL fraction was increased in the L, M and H assays. We selected LDL-Ox (M) to evaluate the expression of genes involved in the inflammatory process (TNFα, iNOS, IL6, FAT/CD3) and 11ß hydroxysteroid dehydrogenase type 2 (11ßHSD2). RAW 264.7 cells were treated with LDL-Ox (M) for 4, 8, 12 and 24 h to a final concentration of 100 µM. The mARN level of different genes were measured by quantitative real-time PCR. Results showed an increase of TNFα, IL6 and iNOS gene expression which was more marked between 4 and 12 h. FAT/CD36 expression increased also in a range of 10-fold at 12 and 24 h versus controls indicating uptakes of fatty acids and formation of foam cells. The effect of LDL-Ox in the culture medium promotes a significant increase of 11ßHSD2 expression between 8 and 12 h. The increase of 11ßHSD2 expression indicates the oxidation of corticosterone to dehydrocorticosterone (glucocorticoid inactive form). This fact would prevent the differentiation towards an antiinflammatory macrophage profile promoted by glucocorticoids.