PMID- 33239022 OWN - NLM STAT- MEDLINE DCOM- 20210916 LR - 20240330 IS - 1742-2094 (Electronic) IS - 1742-2094 (Linking) VI - 17 IP - 1 DP - 2020 Nov 25 TI - Macrophage to myofibroblast transition contributes to subretinal fibrosis secondary to neovascular age-related macular degeneration. PG - 355 LID - 10.1186/s12974-020-02033-7 [doi] LID - 355 AB - BACKGROUND: Macular fibrosis causes irreparable vision loss in neovascular age-related macular degeneration (nAMD) even with anti-vascular endothelial growth factor (VEGF) therapy. Inflammation is known to play an important role in macular fibrosis although the underlying mechanism remains poorly defined. The aim of this study was to understand how infiltrating macrophages and complement proteins may contribute to macular fibrosis. METHODS: Subretinal fibrosis was induced in C57BL/6J mice using the two-stage laser protocol developed by our group. The eyes were collected at 10, 20, 30 and 40 days after the second laser and processed for immunohistochemistry for infiltrating macrophages (F4/80 and Iba-1), complement components (C3a and C3aR) and fibrovascular lesions (collagen-1, Isolectin B4 and alpha-SMA). Human retinal sections with macular fibrosis were also used in the study. Bone marrow-derived macrophages (BMDMs) from C57BL/6J mice were treated with recombinant C3a, C5a or TGF-beta for 48 and 96 h. qPCR, Western blot and immunohistochemistry were used to examine the expression of myofibroblast markers. The involvement of C3a-C3aR pathway in macrophage to myofibroblast transition (MMT) and subretinal fibrosis was further investigated using a C3aR antagonist (C3aRA) and a C3a blocking antibody in vitro and in vivo. RESULTS: Approximately 20~30% of F4/80(+) (or Iba-1(+)) infiltrating macrophages co-expressed alpha-SMA in subretinal fibrotic lesions both in human nAMD eyes and in the mouse model. TGF-beta and C3a, but not C5a treatment, significantly upregulated expression of alpha-SMA, fibronectin and collagen-1 in BMDMs. C3a-induced upregulation of alpha-SMA, fibronectin and collagen-1 in BMDMs was prevented by C3aRA treatment. In the two-stage laser model of induced subretinal fibrosis, treatment with C3a blocking antibody but not C3aRA significantly reduced vascular leakage and Isolectin B4(+) lesions. The treatment did not significantly alter collagen-1(+) fibrotic lesions. CONCLUSIONS: MMT plays a role in macular fibrosis secondary to nAMD. MMT can be induced by TGF-beta and C3a but not C5a. Further research is required to fully understand the role of MMT in macular fibrosis. Macrophage to myofibroblast transition (MMT) contributes to subretinal fibrosis. Subretinal fibrosis lesions contain various cell types, including macrophages and myofibroblasts, and are fibrovascular. Myofibroblasts are key cells driving pathogenic fibrosis, and they do so by producing excessive amount of extracellular matrix proteins. We have found that infiltrating macrophages can transdifferentiate into myofibroblasts, a phenomenon termed macrophage to myofibroblast transition (MMT) in macular fibrosis. In addition to TGF-beta1, C3a generated during complement activation in CNV can also induce MMT contributing to macular fibrosis. RPE = retinal pigment epithelium. BM = Bruch's membrane. MMT = macrophage to myofibroblast transition. TGFB = transforming growth factor beta. a-SMA = alpha smooth muscle actin. C3a = complement C3a. FAU - Little, Karis AU - Little K AD - The Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry & Biomedical Sciences, Queen's University Belfast, 97 Lisburn Road, Belfast, BT9 7BL, UK. FAU - Llorian-Salvador, Maria AU - Llorian-Salvador M AD - The Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry & Biomedical Sciences, Queen's University Belfast, 97 Lisburn Road, Belfast, BT9 7BL, UK. FAU - Tang, Miao AU - Tang M AD - The Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry & Biomedical Sciences, Queen's University Belfast, 97 Lisburn Road, Belfast, BT9 7BL, UK. FAU - Du, Xuan AU - Du X AD - The Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry & Biomedical Sciences, Queen's University Belfast, 97 Lisburn Road, Belfast, BT9 7BL, UK. FAU - Marry, Stephen AU - Marry S AD - The Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry & Biomedical Sciences, Queen's University Belfast, 97 Lisburn Road, Belfast, BT9 7BL, UK. FAU - Chen, Mei AU - Chen M AD - The Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry & Biomedical Sciences, Queen's University Belfast, 97 Lisburn Road, Belfast, BT9 7BL, UK. FAU - Xu, Heping AU - Xu H AUID- ORCID: 0000-0003-4000-931X AD - The Wellcome-Wolfson Institute for Experimental Medicine, School of Medicine, Dentistry & Biomedical Sciences, Queen's University Belfast, 97 Lisburn Road, Belfast, BT9 7BL, UK. heping.xu@qub.ac.uk. LA - eng GR - 722717/H2020 Marie Sklodowska-Curie Actions/ GR - 5057/5058/Fight for Sight UK/ PT - Journal Article DEP - 20201125 PL - England TA - J Neuroinflammation JT - Journal of neuroinflammation JID - 101222974 RN - 80295-42-7 (Complement C3a) SB - IM MH - Animals MH - Cells, Cultured MH - Complement C3a/toxicity MH - Female MH - Fibrosis MH - Humans MH - Macrophages/drug effects/immunology/*pathology MH - Macular Degeneration/chemically induced/immunology/*pathology MH - Male MH - Mice MH - Mice, Inbred C57BL MH - Myofibroblasts/drug effects/immunology/*pathology MH - Neovascularization, Pathologic/chemically induced/immunology/*pathology MH - Retina/drug effects/immunology/*pathology PMC - PMC7690191 OTO - NOTNLM OT - Age-related macular degeneration OT - Complement OT - Inflammation OT - Macrophage to myofibroblast transition OT - Retinal fibrosis COIS- The authors declare that they have no competing interests. EDAT- 2020/11/27 06:00 MHDA- 2021/09/18 06:00 PMCR- 2020/11/25 CRDT- 2020/11/26 05:30 PHST- 2020/07/14 00:00 [received] PHST- 2020/11/11 00:00 [accepted] PHST- 2020/11/26 05:30 [entrez] PHST- 2020/11/27 06:00 [pubmed] PHST- 2021/09/18 06:00 [medline] PHST- 2020/11/25 00:00 [pmc-release] AID - 10.1186/s12974-020-02033-7 [pii] AID - 2033 [pii] AID - 10.1186/s12974-020-02033-7 [doi] PST - epublish SO - J Neuroinflammation. 2020 Nov 25;17(1):355. doi: 10.1186/s12974-020-02033-7.