Novel Role for the AnxA1-Fpr2/ALX Signaling Axis as a Key Regulator of Platelet Function to Promote Resolution of Inflammation
Authors: Senchenkova Elena Y. , Ansari Junaid , Becker Felix , Vital Shantel A. , Al-Yafeai Zaki , Sparkenbaugh Erica M. , Pawlinski Rafal , Stokes Karen Y. , Carroll Jennifer L. , Dragoi Ana-Maria , Qin Cheng Xue , Ritchie Rebecca H. , Sun Hai , Cuellar-Saenz Hugo H. , Rubinstein Mara R. , Han Yiping W. , Orr A. Wayne , Perretti Mauro , Granger D. Neil , Gavins Felicity N.E. 
Published: June 2019

Background:
Ischemia reperfusion injury (I/RI) is a common complication of cardiovascular diseases. Resolution of detrimental I/RI-generated prothrombotic and proinflammatory responses is essential to restore homeostasis. Platelets play a crucial part in the integration of thrombosis and inflammation. Their role as participants in the resolution of thromboinflammation is underappreciated; therefore we used pharmacological and genetic approaches, coupled with murine and clinical samples, to uncover key concepts underlying this role.

Methods:

Middle cerebral artery occlusion with reperfusion was performed in wild-type or annexin A1 (AnxA1) knockout (AnxA1
−/−
) mice. Fluorescence intravital microscopy was used to visualize cellular trafficking and to monitor light/dye–induced thrombosis. The mice were treated with vehicle, AnxA1 (3.3 mg/kg), WRW4 (1.8 mg/kg), or all 3, and the effect of AnxA1 was determined in vivo and in vitro.

Results:

Intravital microscopy revealed heightened platelet adherence and aggregate formation post I/RI, which were further exacerbated in AnxA1
−/−
mice. AnxA1 administration regulated platelet function directly (eg, via reducing thromboxane B
2
and modulating phosphatidylserine expression) to promote cerebral protection post-I/RI and act as an effective preventative strategy for stroke by reducing platelet activation, aggregate formation, and cerebral thrombosis, a prerequisite for ischemic stroke. To translate these findings into a clinical setting, we show that AnxA1 plasma levels are reduced in human and murine stroke and that AnxA1 is able to act on human platelets, suppressing classic thrombin-induced inside-out signaling events (eg, Akt activation, intracellular calcium release, and Ras-associated protein 1 [Rap1] expression) to decrease α
IIb
β
3
activation without altering its surface expression. AnxA1 also selectively modifies cell surface determinants (eg, phosphatidylserine) to promote platelet phagocytosis by neutrophils, thereby driving active resolution. (n=5–13 mice/group or 7–10 humans/group.)

Conclusions:

AnxA1 affords protection by altering the platelet phenotype in cerebral I/RI from propathogenic to regulatory and reducing the propensity for platelets to aggregate and cause thrombosis by affecting integrin (α
IIb
β
3
) activation, a previously unknown phenomenon. Thus, our data reveal a novel multifaceted role for AnxA1 to act both as a therapeutic and a prophylactic drug via its ability to promote endogenous proresolving, antithromboinflammatory circuits in cerebral I/RI. Collectively, these results further advance our knowledge and understanding in the field of platelet and resolution biology.