AUTHOR=Lasch Manuel , Kumaraswami Konda , Nasiscionyte Simona , Kircher Susanna , van den Heuvel Dominic , Meister Sarah , Ishikawa-Ankerhold Hellen , Deindl Elisabeth 

TITLE=RNase A Treatment Interferes With Leukocyte Recruitment, Neutrophil Extracellular Trap Formation, and Angiogenesis in Ischemic Muscle Tissue

JOURNAL=Frontiers in Physiology

VOLUME=11

YEAR=2020

URL=https://www.frontiersin.org/journals/physiology/articles/10.3389/fphys.2020.576736

DOI=10.3389/fphys.2020.576736

ISSN=1664-042X

ABSTRACT=<p><bold>Background:</bold> RNase A (the bovine equivalent to human RNase 1) and RNase 5 (angiogenin) are two closely related ribonucleases. RNase 5 is described as a powerful angiogenic factor. Whether RNase A shares the same angiogenic characteristic, or interferes with vessel growth as demonstrated for arteriogenesis, has never been investigated and is the topic of this present study.</p><p><bold>Methods and Results:</bold> To investigate whether RNase A shows a pro‐ or anti-angiogenic effect, we employed a murine hindlimb model, in which femoral artery ligation (FAL) results in arteriogenesis in the upper leg, and, due to provoked ischemia, in angiogenesis in the lower leg. C57BL/6J male mice underwent unilateral FAL, whereas the contralateral leg was sham operated. Two and seven days after the surgery and intravenous injection of RNase A (50 μg/kg dissolved in saline) or saline (control), the gastrocnemius muscles of mice were isolated from the lower legs for (immuno-) histological analyses. Hematoxylin and Eosin staining evidenced that RNase A treatment resulted in a higher degree of ischemic tissue damage. This was, however, associated with reduced angiogenesis, as evidenced by a reduced capillary/muscle fiber ratio. Moreover, RNase A treatment was associated with a significant reduction in leukocyte infiltration as shown by CD45<sup>+</sup> (pan-leukocyte marker), Ly6G<sup>+</sup> or MPO<sup>+</sup> (neutrophils), MPO<sup>+</sup>/CitH<sub>3</sub><sup>+</sup> [neutrophil extracellular traps (NETs)], and CD68<sup>+</sup> (macrophages) staining. CD68/MRC1 double staining revealed that RNase A treated mice showed a reduced percentage of M1-like polarized (CD68<sup>+</sup>/MRC1<sup>−</sup>) macrophages whereas the percentage of M2-like polarized (CD68<sup>+</sup>/MRC1<sup>+</sup>) macrophages was increased.</p><p><bold>Conclusion:</bold> In contrast to RNase 5, RNase A interferes with angiogenesis, which is linked to reduced leukocyte infiltration and NET formation.</p>