title: Regulation of C3 Activation by the Alternative Complement Pathway in the Mouse Retina.
abstract: The purpose of this study was to examine the retinas of mice carrying hemizygous and null double deletions of Cfb-/- and Cfh-/-, and to compare these with the single knockouts of Cfb, Cfh and Cfd. Retinas were isolated from wild type (WT), Cfb-/-/Cfh-/-, Cfb-/-/Cfh+/-, Cfh-/-/Cfb+/-, Cfb-/-, Cfh-/- Cfd-/-, and Cfd+/- mice. Complement proteins were evaluated by western blotting, ELISA and immunocytochemistry, and retinal morphology was assessed using toluidine blue stained semi-thin sections. WT mice showed staining for C3 and its breakdown products in the retinal vasculature and the basal surface of the retinal pigment epithelium (RPE). Cfb-/- mice exhibited a similar C3 staining pattern to WT in the retinal vessels but a decrease in C3 and its breakdown products at the basal surface of the RPE. Deletion of both Cfb and Cfh restored C3 to levels similar to those observed in WT mice, however this reversal of phenotype was not observed in Cfh-/-/Cfb+/- or Cfb-/-/Cfh+/- mice. Loss of CFD caused an increase in C3 and a decrease in C3 breakdown products along the basal surface of the RPE. Overall the retinal morphology and retinal vasculature did not appear different across the various genotypes. We observed that C3 accumulates at the basal RPE in Cfb-/-, Cfb-/-/Cfh-/-, Cfb-/-/Cfh+/-, Cfd-/- and WT mice, but is absent in Cfh-/- and Cfh-/-/Cfb+/- mice, consistent with its consumption in the serum of mice lacking CFH when CFB is present. C3 breakdown products along the surface of the RPE were either decreased or absent when CFB, CFH or CFD was deleted or partially deleted.
authors: Department of Cell Biology, UCL Institute of Ophthalmology, 11-43 Bath Street, London EC1V 9EL, United Kingdom.
authors: Jennifer A E
authors: JA
authors: Williams
authors: Department of Cell Biology, UCL Institute of Ophthalmology, 11-43 Bath Street, London EC1V 9EL, United Kingdom.
authors: Dimitris
authors: D
authors: Stampoulis
authors: Department of Cell Biology, UCL Institute of Ophthalmology, 11-43 Bath Street, London EC1V 9EL, United Kingdom.
authors: Chloe E
authors: CE
authors: Gunter
authors: Department of Cell Biology, UCL Institute of Ophthalmology, 11-43 Bath Street, London EC1V 9EL, United Kingdom.
authors: John
authors: J
authors: Greenwood
authors: Ophthiris Discovery Performance Unit and Department of Laboratory Animal Science, GlaxoSmithKline, Medicines Research Centre, Gunnelswood Road, Stevenage, Herts SG1 2NY, United Kingdom.
authors: Peter
authors: P
authors: Adamson
authors: Department of Cell Biology, UCL Institute of Ophthalmology, 11-43 Bath Street, London EC1V 9EL, United Kingdom.
authors: Stephen E
authors: http://orcid.org/0000-0002-7304-8879
authors: SE
authors: Moss
mesh_terms: D000818:Animals; D015153:Blotting, Western; D050577:Complement C3-C5 Convertases / Q000378:metabolism*; D003182:Complement C5 / Q000235:genetics / Q000378:metabolism; D011415:Complement Factor B / Q000235:genetics / Q000378:metabolism; D011416:Complement Factor D / Q000235:genetics / Q000378:metabolism; D017242:Complement Factor H / Q000235:genetics / Q000378:metabolism; D003170:Complement Pathway, Alternative / Q000235:genetics / Q000502:physiology*; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D051379:Mice; D008810:Mice, Inbred C57BL; D008817:Mice, Mutant Strains; D012160:Retina / Q000378:metabolism*
chemical_list: D003182:Complement C5; D017242:Complement Factor H; D050577:Complement C3-C5 Convertases; D011416:Complement Factor D; D011415:Complement Factor B