Differential roles of FOXC2 in the trabecular meshwork and Schlemm's canal in glaucomatous pathology

Impaired development and maintenance of Schlemm's canal (SC) are associated with perturbed aqueous humor outflow and intraocular pressure. The angiopoietin (ANGPT)/TIE2 signaling pathway regulates SC development and maintenance, whereas the molecular mechanisms of crosstalk between SC and the neural crest (NC)-derived neighboring tissue, the trabecular meshwork (TM), are poorly understood. Here, we show NC-specific forkhead box (Fox)c2 deletion in mice results in impaired SC morphogenesis, loss of SC identity, and elevated intraocular pressure. Visible-light optical coherence tomography analysis further demonstrated functional impairment of the SC in response to changes in intraocular pressure in NC-Foxc2 -/- mice, suggesting altered TM biomechanics. Single-cell RNA-sequencing analysis identified that this phenotype is predominately characterized by transcriptional changes associated with extracellular matrix organization and stiffness in TM cell clusters, including increased matrix metalloproteinase expression, which can cleave the TIE2 ectodomain to produce soluble TIE2. Moreover, endothelial-specific Foxc2 deletion impaired SC morphogenesis because of reduced TIE2 expression, which was rescued by deleting the TIE2 phosphatase VE-PTP. Thus, Foxc2 is critical in maintaining SC identity and morphogenesis via TM-SC crosstalk. The 10X Genomics Chromium platform was used to obtain single cell transcriptomes from the anterior eye segment of 4 week old wild-type and neural crest-specific Foxc2 knockout mice施莱姆管（Schlemm's canal, SC）的发育与维持受损，与房水外流异常及眼压升高密切相关。血管生成素（angiopoietin, ANGPT）/TIE2信号通路可调控SC的发育与维持，但目前对于SC与神经嵴（neural crest, NC）来源的邻近组织——小梁网（trabecular meshwork, TM）之间的分子互作机制仍不甚明晰。本研究发现，小鼠体内神经嵴特异性叉头框C2（forkhead box C2, Foxc2）敲除会导致SC形态发生受损、SC身份丧失以及眼压升高。可见光光学相干断层成像分析进一步证实，神经嵴特异性Foxc2敲除小鼠的SC在应对眼压波动时存在功能损伤，提示TM的生物力学特性发生改变。单细胞RNA测序分析显示，该表型主要表现为TM细胞簇中与细胞外基质组织及基质刚度相关的转录组变化，包括基质金属蛋白酶表达上调，而该酶可切割TIE2胞外域以产生可溶性TIE2。此外，内皮特异性Foxc2敲除会因TIE2表达下调而损害SC形态发生，这一缺陷可通过敲除TIE2磷酸酶VE-PTP得以挽救。综上，Foxc2通过TM与SC之间的分子互作，对维持SC身份及形态发生至关重要。本研究采用10X Genomics Chromium平台，对4周龄野生型（wild-type）及神经嵴特异性Foxc2敲除小鼠的眼前节组织进行了单细胞转录组测序。