RS Mehta Jain Department of Biochemistry and Cell Biology, KBIRVO block, Vision Research Foundation, Sankara Nethralaya, Chennai, Tamil Nadu, India.
Tamil Nadu Dr MGR Medical University, Chennai, Tamil Nadu, India.
Br J Ophthalmol. 2019 Jul;103(7):985-992. doi: 10.1136/bjophthalmol-2018-312088. Epub 2018 Sep 24.
Pseudoexfoliation syndrome (PXF) is an idiopathic, elastogenesis-associated systemic disease characterised by amyloid-like material aggregates in the eye. Elevated plasma and aqueous humour (aqH) homocysteine (Hcy) is reportedly associated with PXF. This study is aimed to probe Hcy-mediated alterations in elastin expression.
Lens level of Hcy (total Hcy (tHcy)), mRNA expression of , and in lens capsule, protein expression of elastin in aqH were estimated by enzyme immunoassay, quantitative PCR and western blot, respectively in PXF, PXF with glaucoma (PXF-G) cases, in comparison with cataract-alone disease controls. Human lens epithelial cells (hLECs) were exposed to Hcy and homocysteine thiolactone (HCTL) to evaluate elastin expression in vitro. Furthermore, elastin recombinant protein was incubated with Hcy and HCTL to assess secondary and tertiary structural modifications based on circular dichroism spectroscopy, spectrophotometric and SEM studies.
The lens tHcy was significantly high in PXF (p=0.02) and PXF-G (p=0.009). expression was elevated in PXF and PXF-G (p=0.0007). Elastin level in aqH was elevated in PXF (p=0.01) and PXF-G (p=0.002). Hcy (200 µM) and HCTL (1 µM) promoted elastin expression at mRNA level by 36-fold (p=0.02) and 10-fold (p=0.05), respectively, and at protein level by nearly two-fold in cultured hLECs. Secondary structure changes in elastin protein caused by Hcy were evident from 34.11% drop in α-helix and 6.17% gain in β-sheet. Fluorescence, spectral assays and SEM analyses showed aggregation and amyloid formation of elastin with homocysteinylation.
The study reveals that lens accumulation of Hcy associated with hyperhomocysteinaemia is characteristic of PXF that augments elastin expression. Hcy causes structural changes promoting elastin aggregation, thereby contributing to defective elastin in PXF and PXF-G.
假性剥脱综合征（PXF）是一种特发性、弹性蛋白相关的系统性疾病，其特征是眼睛中存在类淀粉样物质聚集。据报道，血浆和房水中升高的同型半胱氨酸（Hcy）与 PXF 有关。本研究旨在探讨 Hcy 介导的弹性蛋白表达改变。
通过酶免疫测定法、定量 PCR 和 Western blot 分别评估 PXF、PXF 伴青光眼（PXF-G）病例和白内障单独疾病对照的晶状体中 Hcy（总 Hcy（tHcy））、晶状体囊 mRNA 表达水平、 、 以及房水中的弹性蛋白蛋白表达水平。将人晶状体上皮细胞（hLECs）暴露于 Hcy 和同型半胱氨酸硫内酯（HCTL）中，以评估体外的弹性蛋白表达。此外，将弹性蛋白重组蛋白与 Hcy 和 HCTL 孵育，基于圆二色性光谱、分光光度法和 SEM 研究评估二级和三级结构修饰。
PXF 中晶状体 tHcy 显著升高（p=0.02），PXF-G 中 tHcy 也显著升高（p=0.009）。PXF 和 PXF-G 中 表达升高（p=0.0007）。PXF 中房水中的弹性蛋白水平升高（p=0.01），PXF-G 中也升高（p=0.002）。Hcy（200µM）和 HCTL（1µM）在 mRNA 水平上分别使弹性蛋白表达增加 36 倍（p=0.02）和 10 倍（p=0.05），在培养的 hLECs 中蛋白水平增加近两倍。Hcy 引起的弹性蛋白蛋白二级结构变化，α-螺旋下降 34.11%，β-折叠增加 6.17%。荧光、光谱分析和 SEM 分析显示同型半胱氨酸化引起弹性蛋白的聚集和淀粉样形成。
本研究表明，与高同型半胱氨酸血症相关的晶状体 Hcy 积累是 PXF 的特征，可增加弹性蛋白表达。Hcy 导致结构变化，促进弹性蛋白聚集，从而导致 PXF 和 PXF-G 中弹性蛋白缺陷。
