Transcriptome Analysis of the Mesenteric Adipose and Liver Reveals Novel Mechanisms by Which Prenatal Alcohol Exposure Contributes to Worse Stroke Outcome

Prenatal alcohol exposure (PAE) is linked to elevated risk for systemic adult-onset diseases like hypertension, impaired glucose and immune regulation, and in animal models, to impaired recovery from acute onset diseases like cerebrovascular ischemic stroke. Recent evidence suggests that the gastrointestinal system rapidly becomes dysbiotic following cerebrovascular stroke, resulting in systemic inflammation. We hypothesized that a history of PAE would modify the systemic effects of stroke, and transduce exposure-dependent transcriptomic changes in downstream sentinel tissues of the enteric portal circulation that have previously been linked to biobehavioral outcomes in rodent PAE models. Pregnant Sprague Dawley rats were exposed to repeated episodes of vaporized ethanol or room air from gestational day 8 to 19. At 5 months, progeny from each treatment condition were subjected to unilateral endothelin-1 induced occlusion of the middle cerebral artery and outcomes evaluated after 2 days while other progeny stayed stroke-naïve. Stroke induced disabilities were assessed by behavioral assays (adhesive removal, Vibrissae evoked fore-limb placement, circling) and infarct size. The mesenteric adipose tissue and liver transcriptomes were assessed by sequencing, from age-matched stroke-exposed and stroke-naïve offspring. In stroke-naïve animals, pathway analysis identified rRNA processing as downregulated and citric acid cycle as upregulated in mesenteric adipose. Weighted gene correlation network analysis (WGCNA) identified, in the liver of stoke-naïve animals, a moderate but significant correlation between PAE status and necroptosis, a proinflammatory form of programmed cell death (Pearson’s r=0.554, p<0.05). Two days after a stroke, PAE rats exhibited worse neurological scores compared to controls (p<0.05). WGCNA after stroke identified an adipose gene network associated with B cell differentiation and NF-kappa B signaling as moderately correlated with post-stroke neurological function (Pearson's r=0.52, p=0.05). Post-stroke WGCNA also identified a liver proinflammatory gene network strongly correlated with post-stroke neurological function (Pearson's r=-0.63, p<0.01). PAE persistently alters the transcriptome of afferent tissue targets of enteric circulation in adult rat offspring. Moreover, PAE-linked enteric inflammation is correlated to worse outcomes following cerebrovascular ischemic stroke in adulthood. Enteric disturbances may mediate adverse brain health outcomes due to PAE in adulthood.产前酒精暴露（Prenatal alcohol exposure, PAE）与成年后全身性成年发病疾病的发病风险升高密切相关，此类疾病包括高血压、血糖调节与免疫功能受损；在动物模型中，其还与急性发作性疾病（如缺血性脑卒中）的恢复能力受损存在关联。近期研究证据表明，缺血性脑卒中发作后，胃肠道系统会快速出现菌群失调，进而引发全身性炎症反应。本研究假设，产前酒精暴露史会改变脑卒中带来的全身性影响，并会在肠门循环的下游哨兵组织中诱导暴露依赖性的转录组变化——这类组织此前在啮齿类产前酒精暴露模型中，已被证实与生物行为学结局相关。 研究人员让妊娠第8天至第19天的斯普拉格·道利大鼠（Sprague Dawley rats）反复暴露于汽化乙醇或洁净空气环境中。当子代大鼠长至5月龄时，对两组处理的子代分别实施单侧内皮素-1（endothelin-1）诱导的大脑中动脉（middle cerebral artery）闭塞手术，其中一组于术后2天评估结局；另一组则不进行脑卒中造模，作为未造模对照组。脑卒中诱导的功能缺损通过行为学实验（黏贴移除实验、触须诱发前肢放置实验、转圈实验）与梗死体积进行评估。研究人员对年龄匹配的造模与未造模子代大鼠的肠系膜脂肪组织与肝脏转录组进行测序分析。 在未造模的大鼠中，通路分析显示肠系膜脂肪组织内的核糖体RNA加工通路呈下调状态，而三羧酸循环通路则呈上调状态。加权基因共表达网络分析（Weighted Gene Correlation Network Analysis, WGCNA）显示，在未造模大鼠的肝脏中，产前酒精暴露状态与坏死性凋亡（necroptosis，一种促炎型程序性细胞死亡）存在中度且显著的相关性（皮尔逊相关系数r=0.554, P<0.05）。脑卒中造模后2天，产前酒精暴露组大鼠的神经功能评分显著低于对照组（P<0.05）。脑卒中造模后的加权基因共表达网络分析显示，一个与B细胞分化及核因子κB（NF-κB）信号通路相关的脂肪组织基因网络，与脑卒中后神经功能存在中度相关性（r=0.52, P=0.05）。脑卒中造模后的加权基因共表达网络分析还显示，肝脏内的促炎基因网络与脑卒中后神经功能存在强相关性（r=-0.63, P<0.01）。 产前酒精暴露会持续改变成年大鼠子代肠门循环传入组织靶点的转录组特征。此外，成年期缺血性脑卒中后，与产前酒精暴露相关的肠道炎症与不良预后存在相关性。肠道功能紊乱可能介导了产前酒精暴露导致的成年期脑健康不良结局。