Center for Neural Repair and Rehabilitation Shriners Hospitals Pediatric Research Center, Lewis Katz School of Medicine, Temple University, Philadelphia, PA 19140, USA.
Department of Neurosurgery, Johns Hopkins Hospital, Baltimore, MD 21287, USA.
Int J Mol Sci. 2023 Sep 5;24(18):13714. doi: 10.3390/ijms241813714.
Fetal alcohol spectrum disorders (FASD) are leading causes of neurodevelopmental disability. The mechanisms by which alcohol (EtOH) disrupts fetal brain development are incompletely understood, as are the genetic factors that modify individual vulnerability. Because the phenotype abnormalities of FASD are so varied and widespread, we investigated whether fetal exposure to EtOH disrupts ribosome biogenesis and the processing of pre-ribosomal RNAs and ribosome assembly, by determining the effect of exposure to EtOH on the developmental expression of 18S rRNA and its cleaved forms, members of a novel class of short non-coding RNAs (srRNAs). In vitro neuronal cultures and fetal brains (11-22 weeks) were collected according to an IRB-approved protocol. Twenty EtOH-exposed brains from the first and second trimester were compared with ten unexposed controls matched for gestational age and fetal gender. Twenty fetal-brain-derived exosomes (FB-Es) were isolated from matching maternal blood. RNA was isolated using Qiagen RNA isolation kits. Fetal brain srRNA expression was quantified by ddPCR. srRNAs were expressed in the human brain and FB-Es during fetal development. EtOH exposure slightly decreased srRNA expression (1.1-fold; = 0.03). Addition of srRNAs to in vitro neuronal cultures inhibited EtOH-induced caspase-3 activation (1.6-fold, = 0.002) and increased cell survival (4.7%, = 0.034). The addition of exogenous srRNAs reversed the EtOH-mediated downregulation of srRNAs (2-fold, = 0.002). EtOH exposure suppressed expression of srRNAs in the developing brain, increased activity of caspase-3, and inhibited neuronal survival. Exogenous srRNAs reversed this effect, possibly by stabilizing endogenous srRNAs, or by increasing the association of cellular proteins with srRNAs, modifying gene transcription. Finally, the reduction in 18S rRNA levels correlated closely with the reduction in fetal eye diameter, an anatomical hallmark of FASD. The findings suggest a potential mechanism for EtOH-mediated neurotoxicity via alterations in 18S rRNA processing and the use of FB-Es for early diagnosis of FASD. Ribosome biogenesis may be a novel target to ameliorate FASD in utero or after birth. These findings are consistent with observations that gene-environment interactions contribute to FASD vulnerability.
胎儿酒精谱系障碍（FASD）是导致神经发育障碍的主要原因。酒精（EtOH）破坏胎儿大脑发育的机制尚不完全清楚，个体易感性的遗传因素也不清楚。由于 FASD 的表型异常如此多样和广泛，我们通过确定暴露于 EtOH 对 18S rRNA 及其切割形式的发育表达的影响，来研究胎儿暴露于 EtOH 是否会破坏核糖体生物发生以及前核糖体 RNA 的加工和核糖体组装。11-22 周）是根据经过 IRB 批准的方案收集的。将 20 个来自第一和第二个三个月的 EtOH 暴露的大脑与 10 个匹配的未暴露对照进行比较，这些对照的胎龄和胎儿性别相匹配。从匹配的母亲血液中分离出 20 个胎儿脑衍生的外泌体（FB-Es）。使用 Qiagen RNA 分离试剂盒分离 RNA。通过 ddPCR 定量胎儿脑 srRNA 的表达。srRNA 在人类大脑和 FB-Es 中在胎儿发育过程中表达。EtOH 暴露略微降低了 srRNA 的表达（1.1 倍； = 0.03）。向体外神经元培养物中添加 srRNAs 抑制了 EtOH 诱导的 caspase-3 激活（1.6 倍， = 0.002）并增加了细胞存活（4.7%， = 0.034）。添加外源性 srRNAs 逆转了 EtOH 介导的 srRNAs 下调（2 倍， = 0.002）。EtOH 暴露抑制了发育中大脑中 srRNAs 的表达，增加了 caspase-3 的活性，并抑制了神经元的存活。外源性 srRNAs 逆转了这种作用，可能是通过稳定内源性 srRNAs，或者通过增加细胞蛋白与 srRNAs 的结合，改变基因转录。最后，18S rRNA 水平的降低与胎儿眼睛直径的降低密切相关，这是 FASD 的解剖学标志。这些发现表明，通过改变 18S rRNA 加工和使用 FB-Es 进行 FASD 的早期诊断，可能存在 EtOH 介导的神经毒性的潜在机制。核糖体生物发生可能是改善宫内或出生后 FASD 的新靶点。这些发现与基因-环境相互作用导致 FASD 易感性的观察结果一致。
