Background/Purpose: ANCA-associated vasculitis (AAV) are relapsing diseases with high morbidity and mortality. The vascular damage present in these patients requires continuous repair with the participation of the endothelial progenitor cells (EPC). The circulating EPC have a fundamental role in angiogenesis, repair and vascular homeostasis. These cells isolated from peripheral blood generate in vitro endothelial colony-forming cells (ECFC) that have high proliferative capacity and stable phenotype throughout the culture. It is believed that there is a deficiency of the EPC’s repair capacity in AAV. There are no studies on the functional capacity of EPC and ECFC in AAV. Understanding the functional disturbances of EPC in AAV patients will result in a better knowledge of its pathogenesis and possible future therapeutic targets. The aim of the study was to evaluate the functional capacity of ECFC in patients with AAV in vitro through angiogenesis and migration assays.

Methods: Patients with a previous diagnosis of AAV without immunosuppression for at least 18 months and healthy controls were selected. First, we isolated ECFC from peripheral blood and characterized them by flow cytometry (FACS). After expansion, we evaluated angiogenesis on Matrigel ™ plates and ECFCs migration capacity. For angiogenesis, ECFC were evaluated and photographed 15 and 24 hours after the preparation. The ECFC migration was performed by the scratch (wound-healing assay) methodology and photographed hourly in an phase-contrast microscope using the Aviocam 506zen 2Pro, for 24 hours after the scratch. The images obtained in both experiments were analyzed using the Image J® software. All the tests consisted of the evaluation of ECFC from patients, healthy controls and human umbilical vein endothelial cells (HUVEC), performed in triplicate.

Results: Peripheral blood was collected from 12 patients diagnosed with AAV. The collection of 7 of these patients was performed at the diagnosis, 5 with granulomatosis with polyangiitis (GPA) and 2 with eosinophilic granulomatosis with polyangiitis (EGPA). The other 5 patients were on follow-up and without immunosuppression for at least 18 months and had a maximum prednisone dose of 10mg daily, three of which were diagnosed with GPA, one with EGPA and one with microscopic polyangiitis (MPA). Success in the isolation and growth in ECFC culture was obtained in 8/12 (66%) of the cases. ECFC from GPA patients had lower rates of migration compared to healthy controls and HUVEC (p=0.002 and p=0.0079, respectively). ECFC from patients with AAV had a higher angiogenesis capacity as shown by a significantly higher number of structures (meshes, nodes and junctions) when compared with ECFCs from healthy controls (p=0.0023, p<0.00001 and p=0.0371, for each structure) and HUVEC (p=0.00001, p=0.0001 and p=0.056).

Conclusion: These findings suggest distinct functional profiles in ECFC from patients with AAV, which could influence disease pathogenesis.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/evaluation-of-the-functional-activity-of-endothelial-progenitor-cells-in-patients-with-anca-associated-vasculitis/