纤维母细胞生长因子21（FGF21）类似物已被用作潜在的物质使用障碍治疗药物。先前的研究表明，FGF21的给予可能会影响酒精摄入和奖赏行为。我们最近的报告显示，血浆FGF21水平与酒精使用障碍（AUD）患者的酒精摄入呈正相关。FGF21的半衰期较短（0.5-2小时）且能够穿过血脑屏障。因此，我们着手寻找天然形式的FGF21和长效FGF21分子（PF-05231023）在诱导多能干细胞（iPSC）衍生的前脑神经元中的分子机制。我们在生理相关浓度下对FGF21和PF-05231023处理的iPSC衍生前脑神经元进行了RNA-seq分析。我们从先前的AUD临床试验（n = 442）中获得了FGF21和GABA的血浆水平。我们在iPSC衍生前脑神经元和前脑器官块中进行了FGF21的ELISA实验。我们使用共免疫沉淀技术确定了蛋白质相互作用。最后，我们应用ChIP实验来确认FGF21在iPSC衍生的前脑神经元中与REST、EZH2和H3K27me3的结合。我们发现，对于天然形式的FGF21和PF-05231023处理，相应地有4701和1956个差异表达基因（FDR＜0.05）。值得注意的是，在天然形式的FGF21和PF-05231023处理中，有974个差异表达基因重叠。REST是差异表达基因中最重要的上游调控因子。GABA能突触通路是我们发现的最显著的通路，基于这些重叠基因。我们还观察到AUD患者的FGF21和GABA浓度之间存在显著正相关。与此相符，FGF21和PF-05231023明显诱导iPSC衍生神经元的GABA水平增加。最后，功能基因组学研究显示，在GABA降解相关基因的启动子区域，FGF21的作用导致了REST、EZH2和H3K27me3的药物依赖性占位，从而导致了转录抑制。我们的研究结果强调了与FGF21作用相关的GABA降解基因表观遗传调控中的显著作用。（ClinicalTrials.gov识别号：NCT00662571）。版权所有©2023年作者。由Elsevier GmbH出版。保留所有权利。

Fibroblast growth factor 21 (FGF21) analogs have been tested as potential therapeutics for substance use disorders. Prior research suggests that FGF21 administration might affect alcohol consumption and reward behaviors. Our recent report showed that plasma FGF21 levels were positively correlated with alcohol use in patients with alcohol use disorder (AUD). FGF21 has a short half-life (0.5-2 hours) and crosses the blood-brain barrier. Therefore, we set out to identify molecular mechanisms for both the naïve form of FGF21 and a long-acting FGF21 molecule (PF-05231023) in induced pluripotent stem cell (iPSC)-derived forebrain neurons.We performed RNA-seq in iPSC-derived forebrain neurons treated with naïve FGF21 or PF-05231023 at physiologically relevant concentrations. We obtained plasma levels of FGF21 and GABA from our previous AUD clinical trial (n=442). We performed ELISA for FGF21 in both iPSC-derived forebrain neurons and forebrain organoids. We determined protein interactions using co-immunoprecipitation. Finally, we applied ChIP assays to confirm the occupancy of REST, EZH2 and H3K27me3 by FGF21 using iPSC-derived forebrain neurons with and without drug exposure.We identified 4701 and 1956 differentially expressed genes in response to naïve FGF21 or PF-05231023, respectively (FDR<0.05). Notably, 974 differentially expressed genes overlapped between treatment with naïve FGF21 and PF-05231023. REST was the most important upstream regulator of differentially expressed genes. The GABAergic synapse pathway was the most significant pathway identified using the overlapping genes. We also observed a significant positive correlation between plasma FGF21 and GABA concentrations in AUD patients. In parallel, FGF21 and PF-05231023 significantly induced GABA levels in iPSC-derived neurons. Finally, functional genomics studies showed a drug-dependent occupancy of REST, EZH2, and H3K27me3 in the promoter regions of genes involved in GABA catabolism which resulted in transcriptional repression.Our results highlight a significant role in the epigenetic regulation of genes involved in GABA catabolism related to FGF21 action. (The ClinicalTrials.gov Identifier: NCT00662571).Copyright © 2023 The Author(s). Published by Elsevier GmbH.. All rights reserved.