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Non‐canonical Cytokine Stimulation of Recurrent Lymphangiogenesis and Investigation of Macrophages in Recurrent Inflammation in Mouse Cornea
Author(s) -
Engels Sarah,
Indermuhle Thad,
Harms Rebecca,
Fink Darci
Publication year - 2021
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2021.35.s1.03202
Subject(s) - lymphangiogenesis , cytokine , inflammation , stimulation , cornea , medicine , immunology , ophthalmology , metastasis , cancer
Macrophage recruitment and lymphangiogenesis are key markers of inflammation.Vascular endothelial growth factor (VEGF‐C), thought to be derived from macrophages, is the primary signaling protein that drives lymphangiogenesis in initial corneal inflammation induced by sutures.However, VEGF‐C is not required for lymphangiogenesis that occurs after cornea has healed and is reinjured causing recurrent inflammation.The purpose of this study was to test the ability of noncanonical (non‐VEGF‐C) cytokines to induce lymphangiogenesis during recurrent corneal inflammation and to identify the polarization of macrophage populations during initial corneal inflammation, wound healing, and recurrent inflammation.Based on the literature, we hypothesized that IGF‐1/2, SHH, and EGF were possible candidate drivers of recurrent corneal lymphangiogenesis.We expected M1 pro‐inflammatory macrophages to be prevalent during initial and recurrent inflammation and that these cells might be the source of the unknown non‐canonical lymphangiogenic factor driving recurrent lymphangiogenesis.Candidate factors were loaded onto slow‐release micropellets and implanted into wound‐recovered corneas that had previously been injured by suturing.Lymphangiogenesis in response to these candidates, VEGF‐C or re‐sutured positive controls, or PBS negative control pellet was quantified by counting lymphatic endothelial cell nuclei per corneal area. Macrophage polarization was determined by immunofluorescence staining for markers of M1 and M2 macrophages, such as IL‐10, CD80, CD206, Arg1, and iNOS.IGF‐1/2, SHH, and EGF did not induce recurrent lymphangiogenesis above the level of the negative control.Surprisingly, VEGF‐C induced recurrent lymphangiogenesis, suggesting that it is sufficient but not necessary for this process.This finding also demonstrates that this assay has sufficient dynamic range to identify drivers of recurrent lymphangiogenesis.Future cytokine pellet experiments will test other candidate mediators of recurrent lymphangiogenesis, while RNAseq and protein array studies are designed to identify new candidate factors and macrophage and lymphatic endothelial cell gene expression during the various phases of this inflammation model. A better understanding of the regulation of lymphangiogenesis during recurrent inflammation and how macrophage subpopulations may shape this response could facilitate future development of treatments for inflammatory disease.