Intestinal Hypoxia and reperfusion: role for Eph-ephrin system

Background and aim: Mesenteric ischemia-reperfusion (I/R) is a life-threatening clinical problem consisting in the temporary deprivation of blood flow to the gut. The ensuing local organ damage is followed by the development of a systemic inflammatory response, eventually leading to multiple organ failure. Several neuronal, paracrine and plasmatic messengers are involved in mesenteric I/R injury. However, up to now, no studies have been performed on Eph receptors, the largest family of tyrosine kinase receptors, and their cell-bound ephrin ligands, whose role in cell adhesion-based processes during inflammation and immunity is growingly emerging. Eph-ephrin interaction generates a bidirectional signaling affecting both the receptor bearing (forward signaling) and the ligand bearing cells (reverse signaling). Our aim was therefore to investigate the effects produced by unidirectional activation of forward signaling (administration of chimeric protein ephrinA1-Fc; 50, 100, 200microg/kg), of reverse signaling (EphA2-Fc; 180microg/kg), or inhibition of both signals (monomeric EphA2 at equimolar doses of ephrinA1-Fc) on the inflammatory responses caused by mesenteric I/R in mice. Methods: Eph-ephrin proteins were intravenously administered to Swiss mice 5 min prior to 45 min occlusion of the superior mesenteric artery (SMA) followed by 5h reperfusion: intestinal and lung myeloperoxidase (MPO) activity, index of neutrophil recruit- ment, gut malondialdehyde (MDA) levels, marker of lipoperoxidation, and edema, index of increased vascular permeability, were compared to those of control I/R mice, receiving only vehicle, and of sham operated (SO) mice, subjected to the same manipulation without SMA occlusion and receiving vehicle. All experiments were performed according to the guidelines for the Care and Use of Animals (DL26/2014). Results: I/R mice displayed increased gut (45.3±4.7 vs 0.8±0.5 U/g, P<0.001) and lung (289.9±21.0vs 144.6±45.1 U/g, P<0.05) MPO activity and higher MDA levels (595.0±87.3 vs 203.4±62.6 nmol/g dry tissue, P<0.05) and edema (4.4±0.1 vs 3.4±0.3 wet to dry weight ratio, P<0.01) compared to SO. EphrinA1- Fc markedly reduced leukocyte infiltration in the gut (24.9±2.9 U/g) at 100microg/kg and in the lung (128.0±19.1 U/g) at 200microg/kg, while monomeric EphA2 significantly lowered intestinal edema formation (3.9±0.1g/g, P<0.05 vs I/R) at the highest dose. Neither EphA2- Fc nor equimolar doses of IgG-Fc alone were effective in modifying I/R-induced inflammatory responses. Conclusions: These preliminary findings indicate that pharmacological modula- tion of Eph-ephrin system may be advantageous in limiting the inflammatory responses elicited by mesenteric I/R: forward signaling activation attenuates local and systemic leuko- cytes enrolment and reverse signaling silencing contrasts local changes in vascular perme- ability.