Abstract 20466: De-Sumoylation and Phosphorylation of Magi1 Induce Microcirculation Dysfunction via Endothelial P90rsk Activation in Endotoxemia
Background: In sepsis, intermittent blood flow (a form of disturbed blood flow) occurs along with microcirculatory dysfunction, leading to multiple tissue/organ damage. Membrane associated guanylate kinase-1 (MAGI1) is a p90RSK-binding protein, but the functional significance of this association remains unknown.
Methods and Results: When endothelial cells (ECs) were exposed to disturbed flow or thrombin, p90RSK was activated and MAGI1-S741 was phosphorylated. Normally, MAGI1 is SUMOylated at K931 and localized in the membrane/cytosol. However, after disturbed flow or thrombin stimulation, MAGI1 not only was phosphorylated at S741 but also was de-SUMOylated. These post-translational modifications occurred independently, and were significantly inhibited by dominant negative p90RSK adenovirus transduction (Ad-DN-p90RSK). Thrombin-induced NF-κB activation, E-selectin expression, and EC permeability measured by trans-endothelial electric resistance were significantly inhibited in ECs transduced by Ad-DN-p90RSK and Ad-MAGI1-S741A mutant. Expression of a p90RSK-MAGI1 binding inhibitory fragment (MAGI1aa362-543) significantly inhibited thrombin-induced NF-κB activation. In contrast to the MAGI1-S741A phosphorylation site mutant, MAGI1-K931R SUMOylation defect mutant enhanced thrombin-induced NF-κB activation and EC permeability, while MAGI1-SUMO3 chimera expression inhibited it. Together, these results suggest that MAGI1 de-SUMOylation plays a key role in up-regulating both EC inflammation and permeability. We then studied effects of FMK-MEA, a specific p90RSK inhibitor, on capillary perfusion in the cerebral cortex of mice exposed to endotoxin using in vivo 2-photon microscopy. p90RSK inhibition reduced the incidence of intermittent blood flow (disturbed flow) and the number of rolling leukocytes, and restored the velocity of capillary blood flow in endotoxemic mice. Systemic blood pressure, blood oxygenation, heart rate, and respiratory rate were not affected by FMK-MEA.
Conclusions: MAGI1-S741 phosphorylation and MAGI1-K931 de-SUMOylation via p90RSK activation are critical in regulating endothelial inflammation and permeability and may play a major role in micro-circulatory dysfunction in sepsis.
Author Disclosures: N. Le: None. O. Polesskaya: None. C.J. Giancursio: None. M. Kim: None. J. Taunton: None. S. Dewhurst: None. A. Pietropaoli: None. K. Fujiwara: None. J. Abe: None.
This research has received full or partial funding support from the American Heart Association
- © 2014 by American Heart Association, Inc.