Association of STAT4 Gene Polymorphisms (rs10181656, rs7574865, rs7601754, rs10168266) and Serum STAT4 Levels in Age-Related Macular Degeneration

**Background:** Age-related macular degeneration (AMD) is a progressive retinal disease and a leading cause of central vision loss in older adults. Its pathogenesis involves drusen formation, local inflammation, and neovascularization. STAT4 is a transcription factor involved in pro-inflammatory signaling and has been linked to autoimmune and inflammatory diseases. The authors hypothesized that STAT4 gene polymorphisms and serum STAT4 levels might influence AMD development, particularly through effects on new vessel formation. This study aimed to investigate associations of four STAT4 single nucleotide polymorphisms (SNPs) — rs10181656, rs7574865, rs7601754, and rs10168266 — and serum STAT4 levels with AMD.

**Methods:** This case-control study included 500 participants: 150 with early AMD, 150 with exudative AMD, and 200 healthy controls. All subjects were examined ophthalmologically, including visual acuity, slit-lamp biomicroscopy, funduscopy, and optical coherence tomography. AMD was classified according to the Age-Related Eye Disease Study criteria. DNA was extracted from venous blood using the salting-out method. Genotyping of the four STAT4 SNPs was performed using TaqMan assays and real-time PCR. Serum STAT4 levels were measured by ELISA in a subset of 40 exudative AMD patients and 40 controls, and separately in early AMD vs controls. Statistical analyses included chi-square tests, Mann-Whitney U tests, and binary logistic regression under multiple genetic models (codominant, dominant, recessive, overdominant, additive). Bonferroni correction set significance at p<0.0125.

**Key Results:** No statistically significant differences were found in genotype or allele frequencies of any of the four STAT4 SNPs between early AMD and controls, or between exudative AMD and controls. Logistic regression analyses under all genetic models showed no significant associations with AMD, even after stratification by gender or age (≤65, >65–≤75, >75 years). However, serum STAT4 levels were significantly lower in exudative AMD patients (median IQR: 0.118 (0.042)) compared to controls (0.262 (0.385), p=0.005). No significant difference was observed between early AMD (mean 0.164 (0.068)) and controls (0.859 (2.122), p=0.226). Further analysis revealed that exudative AMD patients carrying at least one G allele of rs10181656 had lower serum STAT4 than controls (p=0.011), and those with at least one T allele of rs10168266 also had lower levels (p=0.039).

**Clinical Implications:** This study is the first to investigate these specific STAT4 polymorphisms in AMD. The lack of association between STAT4 SNPs and AMD suggests that these variants do not play a major role in AMD susceptibility in this Lithuanian population. However, the significantly lower serum STAT4 levels in exudative AMD patients, particularly those with certain genotypes, point to a potential role of STAT4 in the neovascular (exudative) stage of AMD. This could indicate that STAT4-mediated inflammatory pathways are differentially involved in advanced AMD. The findings highlight the need for larger studies to confirm these observations and to explore STAT4 as a biomarker or therapeutic target for exudative AMD. Limitations include the modest sample size, use of serum STAT4 as a proxy for retinal expression, and lack of adjustment for environmental factors.