Chemical inhibition of immune signaling rescues defects during heart development in Down syndrome via regulating canonical Wnt pathway

Congenital heart defects (CHDs) are very frequent in children with Down syndrome (DS), the genetic condition caused by trisomy of chromosome 21 (T21). However, the mechanisms by which T21 causes susceptibility to CHDs are poorly understood. Here, using a combination of human induced pluripotent stem cell (iPSC)-based model and Dp(16)1Yey/+ (Dp16) a mouse model of DS, we identified downregulation of canonical Wnt signaling that is caused by increased dosage of interferon (IFN) receptors encoded on chromosome 21 (HSA21) as a causative factor of CHDs in DS. We differentiated human iPSCs derived from individuals with DS as well as CHDs (DS/CHD iPSCs), and controls (control iPSCs) into cardiac cells. We observed that T21 upregulates IFN signaling, downregulates the canonical WNT pathway, and impairs cardiac differentiation. Furthermore, genetic and pharmacological normalization of IFN pathways restored canonical WNT signaling and rescued defects during cardiac differentiation of DS/CHD iPSCs. Strikingly, treatment with an inhibitor of Janus kinase, which is activated by IFN receptor engagement normalized the canonical Wnt pathway and ameliorated CHDs in Dp16 embryos. Our findings provide new mechanisms underlying CHDs in DS, ultimately aiding the development of novel therapeutic strategies. Comparative gene expression profiling analysis of RNA-seq data for: 1) human cells derived from healty controls, patients with Down syndrome as well as congenital heart defects, with or without Wnt signaling activator (CHIR99021) and/or JAK inhibitor (JAKi, Tofacitinib) treatments by using the indicated dosage, harvested at different day points during cardiac differentiation as indicated; 2) E15.5 mouse hearts from WT, Dp16 with or without 10 mg/kg bodyweight/day of JAK inhibitor (JAKi, Tofacitinib) treatments from E6.5 to E14.5, low p-STAT1 indicated the p-STAT1 levels are comparable to WT without JAKi treatment, high p-STAT1 indicated the p-STAT1 levels are significantly higher than WT without JAKi treatment.

唐氏综合征（Down syndrome, DS）患儿罹患先天性心脏缺陷（Congenital heart defects, CHDs）的概率极高，该病由21号染色体三体（trisomy of chromosome 21, T21）引发。然而目前学界对T21导致心脏缺陷易感性的具体机制仍知之甚少。本研究结合基于人类诱导多能干细胞（induced pluripotent stem cell, iPSC）的模型与唐氏综合征小鼠模型Dp(16)1Yey/+（Dp16），发现21号染色体（HSA21）上编码的干扰素（interferon, IFN）受体剂量升高会引发经典Wnt信号通路下调，这正是唐氏综合征患者并发先天性心脏缺陷的致病因素。我们分别将源自唐氏综合征合并先天性心脏缺陷患者的诱导多能干细胞（DS/CHD iPSCs）与健康对照诱导多能干细胞（control iPSCs）定向分化为心肌细胞。实验观察到，T21会上调干扰素信号通路、抑制经典Wnt信号通路，并损害心肌分化过程。进一步研究发现，通过遗传或药理学手段干预干扰素通路，可恢复经典Wnt信号通路的活性，并挽救DS/CHD iPSCs在心肌分化过程中的缺陷。值得注意的是，使用经干扰素受体激活的Janus激酶抑制剂处理，能够修复Dp16胚胎的经典Wnt通路异常，并改善其先天性心脏缺陷表型。本研究的转录组测序（RNA-seq）数据比较基因表达谱分析涵盖以下两组样本：1）心肌分化过程中不同时间点收集的人类细胞，包括健康对照、唐氏综合征患者以及合并先天性心脏缺陷的患者来源细胞，且部分细胞经不同剂量的Wnt信号通路激活剂CHIR99021和/或JAK抑制剂托法替布（Tofacitinib, JAKi）处理；2）于E6.5至E14.5期间以10 mg/kg体重/日的剂量接受JAK抑制剂托法替布处理或未处理的野生型（WT）与Dp16小鼠的E15.5心脏组织，其中低p-STAT1水平组的p-STAT1水平与未使用JAK抑制剂的野生型小鼠相当，高p-STAT1水平组的p-STAT1水平则显著高于未使用JAK抑制剂的野生型小鼠。本研究结果为唐氏综合征并发先天性心脏缺陷的潜在机制提供了新的见解，最终有望助力新型治疗策略的开发。