Age-related macular degeneration (AMD) is a major cause of blindness in the developed world. Oxidative stress and inflammation are implicated in AMD, but precise mechanisms remain poorly defined. Carboxyethylpyrrole (CEP) is an AMD-associated lipid peroxidation product. We previously demonstrated that mice immunized with CEP-modified albumin developed AMD-like degenerative changes in the outer retina. Here, we examined the kinetics of lesion development in immunized mice and the presence of macrophages within the interphotoreceptor matrix (IPM), between the retinal pigment epithelium and photoreceptor outer segments. We observed a significant and time-dependent increase in the number of macrophages in immunized mice relative to young age-matched controls prior to overt pathology. These changes were more pronounced in BALB/c mice than in C57BL/6 mice. Importantly, IPM-infiltrating macrophages were polarized toward the M1 phenotype but only in immunized mice. Moreover, when Ccr2-deficient mice were immunized, macrophages were not present in the IPM and no retinal lesions were observed, suggesting a deleterious role for these cells in our model. This work provides mechanistic evidence linking immune responses against oxidative damage with the presence of proinflammatory macrophages at sites of future AMD and experimentally demonstrates that manipulating immunity may be a target for modulating the development of AMD. 1. Introduction Age-related macular degeneration (AMD) is the most common cause of legal blindness in the elderly population of developed countries with over 300,000 newly diagnosed patients per year in Europe and North America [1, 2]. It is widely believed that AMD starts with the insidious, slowly progressing “dry” form (dry AMD) and can later develop into the more severe “wet” AMD, which advances very rapidly and is characterized by abnormal development of blood vessels, a process called choroidal neovascularization (CNV) [3, 4] that affects the macular region of the retina and leads to loss of central vision. In turn, dry AMD without CNV can proceed to focal loss of the retinal pigment epithelium (RPE), termed geographic atrophy (GA), which is accompanied by loss of vision over these slowly expanding areas of RPE atrophy [5]. RPE loss or dysfunction renders the surrounding tissue vulnerable to damage by reactive oxygen species and loss of photoreceptor density ensues [6]. It has been recognized that accumulation of debris below the RPE in the macula, also known as drusen, is a risk factor for AMD. Identification of complement factor
References
[1]
C. A. Augood, J. R. Vingerling, P. T. V. M. De Jong et al., “Prevalence of age-related maculopathy in older Europeans: the European Eye Study (EUREYE),” Archives of Ophthalmology, vol. 124, no. 4, pp. 529–535, 2006.
[2]
J. C. Javitt, Z. Zhou, M. G. Maguire, S. L. Fine, and R. J. Willke, “Incidence of exudative age-related macular degeneration among elderly Americans,” Ophthalmology, vol. 110, no. 8, pp. 1534–1539, 2003.
[3]
S. B. Bressler, M. G. Maguire, N. M. Bressler, and S. L. Fine, “Relationship of drusen and abnormalities of the retinal pigment epithelium to the prognosis of neovascular macular degeneration,” Archives of Ophthalmology, vol. 108, no. 10, pp. 1442–1447, 1990.
[4]
S. H. Sarks, D. Van Driel, L. Maxwell, and M. Killingsworth, “Softening of drusen and subretinal neovascularization,” Transactions of the Ophthalmological Societies of the United Kingdom, vol. 100, no. 3, pp. 414–422, 1980.
[5]
F. G. Holz, C. Bellman, S. Staudt, F. Schütt, and H. E. V？lcker, “Fundus autofluorescence and development of geographic atrophy in age-related macular degeneration,” Investigative Ophthalmology and Visual Science, vol. 42, no. 5, pp. 1051–1056, 2001.
[6]
Y. Imamura, S. Noda, K. Hashizume et al., “Drusen, choroidal neovascularization, and retinal pigment epithelium dysfunction in SOD1-deficient mice: a model of age-related macular degeneration,” Proceedings of the National Academy of Sciences of the United States of America, vol. 103, no. 30, pp. 11282–11287, 2006.
[7]
J. W. Crabb, M. Miyagi, X. Gu et al., “Drusen proteome analysis: an approach to the etiology of age-related macular degeneration,” Proceedings of the National Academy of Sciences of the United States of America, vol. 99, no. 23, pp. 14682–14687, 2002.
[8]
A. O. Edwards, R. Ritter, K. J. Abel, A. Manning, C. Panhuysen, and L. A. Farrer, “Complement factor H polymorphism and age-related macular degeneration,” Science, vol. 308, no. 5720, pp. 421–424, 2005.
[9]
B. Gold, J. E. Merriam, J. Zernant et al., “Variation in factor B (BF) and complement component 2 (C2) genes is associated with age-related macular degeneration,” Nature Genetics, vol. 38, no. 4, pp. 458–462, 2006.
[10]
G. S. Hageman, D. H. Anderson, L. V. Johnson et al., “A common haplotype in the complement regulatory gene factor H (HF1/CFH) predisposes individuals to age-related macular degeneration,” Proceedings of the National Academy of Sciences of the United States of America, vol. 102, no. 20, pp. 7227–7232, 2005.
[11]
J. L. Haines, M. A. Hauser, S. Schmidt et al., “Complement factor H variant increases the risk of age-related macular degeneration,” Science, vol. 308, no. 5720, pp. 419–421, 2005.
[12]
R. J. Klein, C. Zeiss, E. Y. Chew et al., “Complement factor H polymorphism in age-related macular degeneration,” Science, vol. 308, no. 5720, pp. 385–389, 2005.
[13]
J. R. W. Yates, T. Sepp, B. K. Matharu et al., “Complement C3 variant and the risk of age-related macular degeneration,” New England Journal of Medicine, vol. 357, no. 6, pp. 553–561, 2007.
[14]
D. H. Anderson, R. F. Mullins, G. S. Hageman, and L. V. Johnson, “A role for local inflammation in the formation of drusen in the aging eye,” American Journal of Ophthalmology, vol. 134, no. 3, pp. 411–431, 2002.
[15]
R. B. Nussenblatt and F. Ferris III, “Age-related macular degeneration and the immune response: implications for therapy,” American Journal of Ophthalmology, vol. 144, no. 4, pp. 618–626, 2007.
[16]
D. Weismann, K. Hartvigsen, N. Lauer et al., “Complement factor H binds malondialdehyde epitopes and protects from oxidative stress,” Nature, vol. 478, no. 7367, pp. 76–81, 2011.
[17]
D. S. Friedman, B. J. O'Colmain, S. C. Tomany et al., “Prevalence of age-related macular degeneration in the United States,” Archives of Ophthalmology, vol. 122, no. 4, pp. 564–572, 2004.
[18]
P. J. Rosenfeld, D. M. Brown, J. S. Heier et al., “Ranibizumab for neovascular age-related macular degeneration,” New England Journal of Medicine, vol. 355, no. 14, pp. 1419–1431, 2006.
[19]
X. Gu, S. G. Meer, M. Miyagi et al., “Carboxyethylpyrrole protein adducts and autoantibodies, biomarkers for age-related macular degeneration,” Journal of Biological Chemistry, vol. 278, no. 43, pp. 42027–42035, 2003.
[20]
J. Gu, G. J. T. Pauer, X. Yue et al., “Proteomic and genomic biomarkers for age-related macular degeneration,” Advances in Experimental Medicine and Biology, vol. 664, pp. 411–417, 2010.
[21]
R. E. Anderson, “Lipids of ocular tissues. IV. A comparison of the phospholipids from the retina of six mammalian species,” Experimental Eye Research, vol. 10, no. 2, pp. 339–344, 1970.
[22]
J. G. Hollyfield, V. L. Bonilha, M. E. Rayborn et al., “Oxidative damage-induced inflammation initiates age-related macular degeneration,” Nature Medicine, vol. 14, no. 2, pp. 194–198, 2008.
[23]
J. G. Hollyfield, V. L. Perez, and R. G. Salomon, “A hapten generated from an oxidation fragment of docosahexaenoic acid is sufficient to initiate age-related macular degeneration,” Molecular Neurobiology, vol. 41, no. 2-3, pp. 290–298, 2010.
[24]
R. B. Nussenblatt, B. Iu, and Z. Li, “Age-related macular degeneration: an immunologically driven disease,” Current Opinion in Investigational Drugs, vol. 10, no. 5, pp. 434–442, 2009.
[25]
P. S. Mettu, A. R. Wielgus, S. S. Ong, and S. W. Cousins, “Retinal pigment epithelium response to oxidant injury in the pathogenesis of early age-related macular degeneration,” Molecular Aspects of Medicine, vol. 33, no. 4, pp. 376–398, 2012.
[26]
S. K. Biswas and A. Mantovani, “Macrophage plasticity and interaction with lymphocyte subsets: cancer as a paradigm,” Nature Immunology, vol. 11, no. 10, pp. 889–896, 2010.
[27]
A. Sica and A. Mantovani, “Macrophage plasticity and polarization: in vivo veritas,” Journal of Clinical Investigation, vol. 122, no. 3, pp. 787–795, 2012.
[28]
Y. Wang, M. Burnier, B. Detrick, and J. J. Hooks, “Genetic predisposition to coronavirus-induced retinal disease,” Investigative Ophthalmology and Visual Science, vol. 37, no. 1, pp. 250–254, 1996.
[29]
J. Ambati, A. Anand, S. Fernandez et al., “An animal model of age-related macular degeneration in senescent Ccl-2- or Ccr-2-deficient mice,” Nature Medicine, vol. 9, no. 11, pp. 1390–1397, 2003.
[30]
S. Y. Schmidt and R. D. Peisch, “Melanin concentration in normal human retinal pigment epithelium. Regional variation and age-related reduction,” Investigative Ophthalmology and Visual Science, vol. 27, no. 7, pp. 1063–1067, 1986.
[31]
T. F. Ng and J. W. Streilein, “Light-induced migration of retinal microglia into the subretinal space,” Investigative Ophthalmology and Visual Science, vol. 42, no. 13, pp. 3301–3310, 2001.
[32]
U. F. O. Luhmann, S. Robbie, P. M. G. Munro et al., “The drusenlike phenotype in aging Ccl2-knockout mice is caused by an accelerated accumulation of swollen autofluorescent subretinal macrophages,” Investigative Ophthalmology and Visual Science, vol. 50, no. 12, pp. 5934–5943, 2009.
[33]
J. Kelly, A. A. Khan, J. Yin, T. A. Ferguson, and R. S. Apte, “Senescence regulates macrophage activation and angiogenic fate at sites of tissue injury in mice,” Journal of Clinical Investigation, vol. 117, no. 11, pp. 3421–3426, 2007.
[34]
C. A. Lange, S. Robbie, P. M. G. Munro et al., “Differential modulation of retinal degeneration by Ccl2 and Cx3cr1 chemokine signalling,” PLoS ONE, vol. 7, no. 4, Article ID e35551, 2012.
[35]
M. J. Mattapallil, E. F. Wawrousek, C. C. Chan et al., “The Rd8 mutation of the Crb1 gene is present in vendor lines of C57BL/6N mice and embryonic stem cells, and confounds ocular induced mutant phenotypes,” Investigative Ophthalmology & Visual Science, vol. 53, no. 6, pp. 2921–2927, 2012.
[36]
W. Ma, L. Zhao, A. M. Fontainhas, R. N. Fariss, and W. T. Wong, “Microglia in the mouse retina alter the structure and function of retinal pigmented epithelial cells: a potential cellular interaction relevant to AMD,” PLoS ONE, vol. 4, no. 11, Article ID e7945, 2009.
[37]
S. L. Doyle, M. Campbell, E. Ozaki et al., “NLRP3 has a protective role in age-related macular degeneration through the induction of IL-18 by drusen components,” Nature Medicine, vol. 18, no. 5, pp. 791–798, 2012.
[38]
A. London, E. Itskovich, I. Benhar et al., “Neuroprotection and progenitor cell renewal in the injured adult murine retina requires healing monocyte-derived macrophages,” Journal of Experimental Medicine, vol. 208, no. 1, pp. 23–39, 2011.
[39]
X. Cao, D. Shen, M. M. Patel et al., “Macrophage polarization in the maculae of age-related macular degeneration: a pilot study,” Pathology International, vol. 61, no. 9, pp. 528–535, 2011.
[40]
L. Lu, X. Gu, L. Hong et al., “Synthesis and structural characterization of carboxyethylpyrrole-modified proteins: mediators of age-related macular degeneration,” Bioorganic and Medicinal Chemistry, vol. 17, no. 21, pp. 7548–7561, 2009.
