title: CFH Y402H polymorphism is associated with elevated vitreal GM-CSF and choroidal macrophages in the postmortem human eye.
abstract: OBJECTIVE
  Age-related macular degeneration (AMD) is the leading cause of irreversible blindness in people 50 years of age or older in developed countries. The homozygous CC genotype in the complement factor H (CFH) Y402H single nucleotide polymorphism (SNP; rs1061170) is widely recognized as a risk factor for the development of AMD. In this study, we examined vitreal levels of granulocyte macrophage colony-stimulating factor (GM-CSF), a hematopoietic cytokine, and macrophages in the choroid of postmortem human eyes genotyped for the CFH Y402H SNP.
  
  
  METHODS
  Twenty-two pairs of postmortem, non-diseased, human donor eyes were obtained. The vitreous and retinal tissues of the left eyes were collected for GM-CSF level measurement and CFH Y402H genotyping, respectively. The right eyes were paraffin-embedded and sectioned for immunohistochemistry using a macrophage and microglia marker, CD68. Cell cultures of RPE cells were stimulated with complement C3a, C5a, 4-hydroxynonenal (HNE), or tumor necrosis factor alpha (TNF-α), and GM-CSF expression was measured with a suspension assay or quantitative PCR.
  
  
  RESULTS
  Eyes genotyped with the CC or the CT risk variant of the CFH Y402H SNP showed significantly increased levels of GM-CSF in the vitreous compared to eyes with the protective TT variant (mean ± standard error of mean, 607.54±85.83 pg/ml or 656.32±15.20 pg/ml versus 286.69±81.96 pg/ml, p<0.05). The choroid of eye tissues genotyped with the CC variant showed higher levels of CD68 immunoreactivity than the tissues genotyped with the TT variant (p<0.05). The GM-CSF levels detected in the supernatant of RPE cells in culture treated with HNE or TNF-α were significantly higher compared to the non-treated control (145.88±5.06 pg/ml and 149.32±3.76 pg/ml versus 123.27±4.05 pg/ml, p<0.05). Furthermore, the gene expression of GM-CSF detected in the lysate of RPE cells stimulated with complement C3a or C5a showed significantly increased fold changes compared to the non-treated control (C3a: 2.38±0.31 fold, p<0.05; C5a: 2.84±0.54 fold, p<0.01).
  
  
  CONCLUSIONS
  Our data showed a relationship between the CFH Y402H polymorphism and GM-CSF levels in the vitreous and accumulation of choroidal macrophages in the postmortem eye. These data suggest that the at-risk variant of the CFH gene may contribute to the dysregulation of proinflammatory cytokines locally in the eye.
authors: Department of Ophthalmology and Visual Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Jay Ching Chieh
authors: JC
authors: Wang
authors: Department of Ophthalmology and Visual Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Sijia
authors: S
authors: Cao
authors: Department of Ophthalmology and Visual Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Aikun
authors: A
authors: Wang
authors: Department of Ophthalmology and Visual Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Eleanor
authors: E
authors: To
authors: Department of Ophthalmology and Visual Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Geoffrey
authors: G
authors: Law
authors: Department of Ophthalmology and Visual Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Jiangyuan
authors: J
authors: Gao
authors: Department of Ophthalmology and Visual Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Dean
authors: D
authors: Zhang
authors: Department of Ophthalmology and Visual Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Jing Z
authors: JZ
authors: Cui
authors: Department of Ophthalmology and Visual Sciences, University of British Columbia, Vancouver, BC, Canada.
authors: Joanne A
authors: JA
authors: Matsubara
mesh_terms: D000447:Aldehydes / Q000494:pharmacology; D019943:Amino Acid Substitution; D001344:Autopsy; D002478:Cells, Cultured; D002829:Choroid / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*; D015926:Complement C3a / Q000494:pharmacology; D015936:Complement C5a / Q000494:pharmacology; D017242:Complement Factor H / Q000235:genetics* / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D005838:Genotype; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008264:Macrophages / Q000166:cytology* / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D020641:Polymorphism, Single Nucleotide; D055213:Retinal Pigment Epithelium / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D014822:Vitreous Body / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*
chemical_list: D000447:Aldehydes; D014409:Tumor Necrosis Factor-alpha; D015926:Complement C3a; D015936:Complement C5a; D017242:Complement Factor H; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; C027576:4-hydroxy-2-nonenal