Wanhainen, A. | Search bioRxiv

bioRxiv2026

A splenic reservoir of pro-angiogenic neutrophils supports blood flow recovery after peripheral tissue ischemia

Background: Neutrophils accumulate at sites of ischemic injury, where a distinct subset characterized by CD49d+CXCR4hiVEGFR1hi expression promotes angiogenesis and contributes to blood flow recovery. This study investigated the function, origin, and recruitment mechanisms of these pro-angiogenic neutrophils (PANs) in a murine model of hindlimb ischemia and explored whether a corresponding population responds to acute ischemia in humans. Methods: Using spectral flow cytometry and immunohistochemistry, PANs and classical neutrophils were characterized in ischemic gastrocnemius muscle following hindlimb ischemia in mice. Blood flow recovery was assessed by laser speckle flowmetry. To determine the origin and mobilization mechanisms of PANs, splenectomy, chemical sympathectomy with 6-hydroxydopamine (6-OHDA), and surgical splenic denervation were performed. Translational studies examined blood samples collected during elective abdominal aortic aneurysm repair to assess PAN mobilization following acute lower limb ischemia induced by aortic cross-clamping. Results: PANs rapidly accumulated in ischemic muscle through a CD49d-dependent mechanism, and blockade of CD49d impaired blood flow recovery. During homeostasis, PANs were maintained within a splenic reservoir and were rapidly released into circulation following ischemia. Splenectomy and disruption of splenic sympathetic signaling significantly reduced PAN recruitment to ischemic tissue and impaired functional recovery. Mobilization was driven by ischemia-induced sympathetic activation, which decreased CXCL12 levels in the splenic red pulp, disrupting CXCR4-mediated PAN retention. In humans, a phenotypically similar PAN population was identified in peripheral blood, which increased rapidly following lower limb ischemia during aortic cross-clamping. The speed of this response is consistent with release from an extramedullary reservoir, although its anatomical source remains unknown. Conclusion: These findings identify the spleen as a reservoir of pro-angiogenic neutrophils in mice and reveal a neuroimmune mechanism regulating their mobilization and recruitment after ischemic injury. The rapid appearance of PAN in human circulation after acute ischemia suggests that comparable extramedullary neutrophil reservoirs may also exist in humans.

immunology↗

bioRxiv2023

Glycoprotein VI is Critical for the Detection and Progression of Abdominal Aortic Aneurysms

A common feature in patients with abdominal aortic aneurysms (AAA) is the formation of a nonocclusive intraluminal thrombus (ILT) in regions of aortic dilation. Platelets are known to maintain hemostasis and propagate thrombosis through several redundant activation mechanisms, yet the role of platelet activation in the pathogenesis of AAA associated ILT is still poorly understood. Thus, we sought to investigate how platelet activation impacts the pathogenesis of AAA. Using RNA-sequencing, we identify that the platelet-associated transcripts are significantly enriched in the ILT compared to the adjacent aneurysm wall and healthy control aortas. We found that the platelet specific receptor glycoprotein VI (GPVI) is among the top enriched genes in AAA ILT and is increased on the platelet surface of AAA patients. Examination of a specific indicator of platelet activity, soluble GPVI (sGPVI), in two independent AAA patient cohorts is highly predictive of a AAA diagnosis and associates more strongly with aneurysm growth rate when compared to D-dimer in humans. Finally, intervention with the anti-GPVI antibody (J) in mice with established aneurysms blunted the progression of AAA in two independent mouse models. In conclusion, we show that levels of sGPVI in humans can predict a diagnosis of AAA and AAA growth rate, which may be critical in the identification of high-risk patients. We also identify GPVI as a novel platelet-specific AAA therapeutic target, with minimal risk of adverse bleeding complications, where none currently exist. KEY POINTSO_LISoluble glycoprotein VI, which is a platelet-derived blood biomarker, predicts a diagnosis of AAA, with high sensitivity and specificity in distinguishing patients with fast from slow-growing AAA. C_LIO_LIBlockade of glycoprotein VI in mice with established aneurysms reduces AAA progression and mortality, indicating therapeutic potential. C_LI

pathology↗