Association Study Between Human Retinal Ganglion Cell Regulatory Network Genes Single Nucleotide Polymorphism With Primary Open-angle Glaumoca

Primary open-angle glaucoma (POAG) is a condition of disorders that have optic nerve damage and slow progressive degeneration of retina ganglion cells, causing irreversible defects in the visual field usually associated with elevated intraocular pressure (IOP). It is projected that approximately 66 million individuals worldwide are estimated to have glaucoma, and POAG is the most widespread type. The molecular biological basis of the disease is not totally understood and the factors contributing to its progression are not fully characterized. Many studies have documented that POAG is thought to have multifactorial etiology, including family history, race, IOP and central corneal thickness (CCT). First-degree relatives of individuals with POAG have three- to nine folds greater risk than that of the population in general. In the past 20 years, an impressive amount of knowledge has been generated on molecular regulatory mechanisms that control retinal development in embryonic and neonatal life. This maybe can give us a new insight into POAG.Retinal ganglion cell (RGC) formation is a continuation of the early morphological and molecular processes of retina development. RGC specification and differentiation proceed as a stepwise process when the retina has developed to a certain stage. A specific gene regulatory program is required to accomplish each step. Thus, gene regulation during RGC formation is inherently hierarchical, with transcription factors regulating the early events positioned at the top of the hierarchy and those for the late events at the bottom. In the past decade, substantial inroads have been made into identifying many of the major transcriptional regulators within the hierarchy, including Pax6, Six3, Rx, Chx10, Notch and Notch pathway components, Ath5, and the Brn3 factors.In the present study, we choose Pax6, ATOH7, Brn3a, Brn3b, Isll and Six1 as the candidate genes. Sixteen SNPs were genotyped by using the SnaPshot method in a Caucasian cohort with 762 cases and 464 age and ethnicity matched controls. To confirm our association study result, we used three different models of glaucoma (optic nerve crush model, retinal transient ischemia model and N-methyl-d-aspartate (NMDA) induced retinal ganglion cell loss model) in mice and the expression of these genes was examined by real time polymerase chain reaction.For the two SNPs we tested near Pax6 gene, rs3026393 showed significant association with POAG in the Caucasian cohort (P=2.8x10-4 for an additive model, OR=1.69, 95%CI:1.27-2.26). Rs2239789 showed significant association with POAG in the Caucasian cohort (P=3.1x10-4 for an additive model, OR=1.69,95%CI:1.27-2.27). For the five SNPs we tested near ATOH7 gene, there were no significant difference between case and control. For the two SNPs we tested near Brn3a gene, rs9601092 showed significant association with POAG in the Caucasian cohort (P=0.043 for a recessive model, OR=0.78,95%CI:0.61-0.99). For the two SNPs we tested near ISL1 gene, rs2288468 showed significant association with POAG in the Caucasian cohort (P=0.023 for a recessive model, OR=0.76,95%CI:0.59-0.96). For the four SNPs we tested near Six1 gene, rs10483727 showed significant association with POAG in the Caucasian cohort (P=1.1x10-5 for an additive model, OR=1.37,95%CI:1.19-1.59).Early development gene, Six1, was induced significantly in crush model at as early as Day 3 and persisted till Day 7 while that significant induction of the gene was observed in NMDA and IOP models only in Day 5. On the other hand, Brn3a, a marker of ganglion cells, was significantly reduced in gene expression in all three ganglion cell death models, and the time-course reduction trend coincides with the induction trend of Six1 in respective model. The higher the induction of Six1 gene, the less Brn3a diminished after each injury. In the IOP model, other early eye-field genes like Pax6 and Isl-1, were induced like Six1. IOP is the only model that resulted in the reduction of Brn3b mRNA level.In summary, our results provide further demonstration that Pax6 gene, Brn3a gene and Six1 gene were associated with POAG and plays a potential role in POAG.