Estudo do papel da proteína induzida por glicocorticóide Anexina ...

ABSTRACT
The inflammatory response is initially characterized by the release of pro-inflammatory
mediators and the migration of leukocytes to the inflammation site. On the other hand, this
process is finely controlled by the action of anti-inflammatory and/or pro-resolution mediators
that promote resolution of inflammation and helps the return to tissue homeostasis. Annexin
A1 (AnxA1) is a glucocorticoid (GC)-induced protein that mediates several GC functions.
AnxA1 acts as a potent endogenous modulator of inflammation by preventing the action of
enzymes that participate in the production of pro-inflammatory mediators, inhibiting the
leukocyte transmigration and inducing phagocytosis of apoptotic cells. Thus, this study
investigated the role of AnxA1 on the spontaneous and pharmacologically-induced resolution
of the inflammatory response. For this, we have used a murine model of pleurisy induced by
LPS (lipopolysaccharide) where BALB/c mice were injected with 250 ηg LPS by intrapleural
route. After that, the cells in the pleural lavage were collected at different post-injection
intervals and analyzed for total and differential leukocyte count, number of cells with
apoptotic morphology, signaling pathways involved in apoptosis and cell survival, and
AnxA1 protein expression. Our results have shown that the injection of LPS induced an
accumulation of neutrophils in the pleural cavity which was maximal between 8-24h,
decreased in 48h with complete resolution occurring at 72h. The natural resolution, showed
by the reduction of neutrophils numbers observed at the time of 48-72h, was accompanied by
an increase of mononuclear cells and was associated with an increase of apoptotic events
between 8-48h, as shown by morphological (apoptotic cells count) and biochemical (increase
of caspase-3 cleavage and Bax) criteria. The protein AnxA1 in intact active form (37 kDa)
was detected in animals injected with PBS, strongly decreased in the active phase of
neutrophilic inflammation (4-24h) and increased during the natural resolution, as seen in 48-
72h after LPS challenge. There was a predominance of AnxA1 protein cleavage (33 kDa
band) during the time-points of higher neutrophil recruitment. Treatment of animals with anti-
inflammatory drugs, which have been previously shown to induce resolution in the same
inflammatory model, to know: rolipram (PDE4 inhibitor), wortmannin (PI3K inhibitor) and
dexametasone (synthetic glucocorticoid) decreased the number of neutrophils into the pleural
cavity, with an increase of AnxA1 active form expression and prevention of its cleavage.
Also, the increase of AnxA1 induced by these drugs has been associated with activation of
pro-apoptotic proteins (increase of Bax and caspase-3 cleavage) and decreased