Department of Ophthalmology, Mureş County Clinical Hospital, Târgu Mureş, Romania;
Rom J Morphol Embryol. 2022 Jan-Mar;63(1):121-127. doi: 10.47162/RJME.63.1.12.
The aim of the study was to better understand the interplay between genetic factors and the aging process in the human retina through mapping complement factor H (CFH) and related proteins. Two human eyes, from 92- and 64-year-old donors, were genotyped for the expression of CFH-related 1 (CFHR1) and CFH-related 3 (CFHR3) genes. Deoxyribonucleic acid (DNA) was extracted and analyzed for concentration and purity with a spectrophotometer, at 260 nm. The results showed a DNA concentration of 469.17 ng∕μL in the aged retina and of 399.20 ng∕μL in the younger one. Through polymerase chain reaction (PCR) genotyping, the DNA CFHR1 and CFHR3 were visible as bands of 175 bp and 181 bp. Immunohistochemistry by immunofluorescence method was used with a panel of specific antibodies for CFH, CFHR1, CFHR3 and GFAP, a marker for Müller cells. All the samples were examined, and images captured using confocal microscopy. In the younger retina, CFH was localized in the inner plexiform layer and below the outer nuclear layer, while in the aged retina, it was found in the photoreceptors. CFH was also detected in the choriocapillaris and within the end-feet of the Müller cells. Our controls showed autofluorescence of the retinal pigment epithelium shedding light on a false positive CFH immunostaining of this layer. GFAP immunoreactivity highlighted an increased gliosis within the aged retina. CFHR3 signal was found in the microglia, while CFHR1 was detected in the choriocapillaris. In summary, underpinning the expression of these components can show the potential involvement of these modulators in implementing new treatment strategies.
本研究旨在通过绘制补体因子 H（CFH）及其相关蛋白，更好地了解遗传因素与人类视网膜衰老过程之间的相互作用。对来自 92 岁和 64 岁供体的两只人眼进行 CFH 相关 1（CFHR1）和 CFH 相关 3（CFHR3）基因的表达基因分型。提取脱氧核糖核酸（DNA）并用分光光度计在 260nm 处分析浓度和纯度。结果显示，老年视网膜的 DNA 浓度为 469.17ng/μL，年轻视网膜的 DNA 浓度为 399.20ng/μL。通过聚合酶链反应（PCR）基因分型，可在 175bp 和 181bp 处看到 DNA CFHR1 和 CFHR3 条带。通过免疫荧光法进行免疫组织化学，使用 CFH、CFHR1、CFHR3 和 GFAP（Müller 细胞的标志物）的特异性抗体进行面板检测。所有样本均进行了检查，并使用共聚焦显微镜捕获图像。在年轻的视网膜中，CFH 定位于内丛状层和外核层下方，而在老年视网膜中，CFH 位于光感受器中。CFH 也在脉络膜毛细血管和 Müller 细胞的终足内被检测到。我们的对照显示视网膜色素上皮的自发荧光，揭示了该层 CFH 免疫染色的假阳性。GFAP 免疫反应性突出显示老年视网膜内胶质增生增加。CFHR3 信号在小胶质细胞中发现，而 CFHR1 在脉络膜毛细血管中被检测到。总之，这些成分的表达可以表明这些调节剂在实施新的治疗策略中的潜在作用。
