Department of Biomedical Engineering, The City College of the City University of New York, New York, NY 10031, USA.
Department of Psychiatry, Columbia University, New York, NY 10032, USA.
Cells. 2023 Jan 27;12(3):422. doi: 10.3390/cells12030422.
We have previously established that the integrity of the induced blood-brain barrier (iBBB) formed by brain microvascular endothelial cells derived from the iPSC of 22q11.2 DS (22q11.2 Deletion Syndrome, also called DiGeorge Syndrome) patients is compromised. We tested the possibility that the haploinsufficiency of CRKL, a gene within the 22q11.2 DS deletion region, contributes to the deficit. The CRKL is a major substrate of the Abl tyrosine kinase, and the Abl/CRKL signaling pathway is critical for endothelial barrier functions. Imatinib, an FDA-approved drug, inhibits Abl kinase and has been used to treat various disorders involving vascular leakages. To test if imatinib can restore the compromised iBBB, we treated the patient's iBBB with imatinib. After treatment, both trans-endothelial electrical resistance and solute permeability returned to comparable levels of the control iBBB. Correspondingly, changes in tight junctions and endothelial glycocalyx of the iBBB were also restored. Western blotting showed that imatinib increased the level of active forms of the CRKL protein. A transcriptome study revealed that imatinib up-regulated genes in the signaling pathways responsible for the protein modification process and down-regulated those for cell cycling. The KEGG pathway analysis further suggested that imatinib improved the gene expression of the CRKL signaling pathway and tight junctions, which agrees with our expectations and the observations at protein levels. Our results indicate that the 22q11.2DS iBBB is at least partially caused by the haploinsufficiency of CRKL, which can be rescued by imatinib via its effects on the Abl/CRKL signaling pathway. Our findings uncover a novel disease mechanism associated with 22q11.2DS.
我们之前已经证实,由 22q11.2 DS(22q11.2 缺失综合征,也称为 DiGeorge 综合征)患者的诱导血脑屏障(iBBB)形成的脑微血管内皮细胞的完整性受到损害。我们测试了 22q11.2 DS 缺失区域内的 CRKL 基因的单倍不足是否导致了这种缺陷的可能性。CRKL 是 Abl 酪氨酸激酶的主要底物,而 Abl/CRKL 信号通路对于内皮屏障功能至关重要。伊马替尼是一种获得 FDA 批准的药物,可抑制 Abl 激酶,已用于治疗各种涉及血管渗漏的疾病。为了测试伊马替尼是否可以恢复受损的 iBBB,我们用伊马替尼处理患者的 iBBB。治疗后,跨内皮电阻和溶质通透性均恢复到对照 iBBB 的可比水平。相应地,iBBB 的紧密连接和内皮糖萼的变化也得到了恢复。Western blot 显示伊马替尼增加了 CRKL 蛋白的活性形式的水平。转录组研究表明,伊马替尼上调了负责蛋白质修饰过程的信号通路中的基因,并下调了那些与细胞周期有关的基因。KEGG 通路分析进一步表明,伊马替尼改善了 CRKL 信号通路和紧密连接的基因表达,这与我们的预期和蛋白质水平的观察结果一致。我们的结果表明,22q11.2 DS iBBB 至少部分是由 CRKL 的单倍不足引起的,伊马替尼可以通过其对 Abl/CRKL 信号通路的影响来挽救这种情况。我们的发现揭示了与 22q11.2 DS 相关的一种新的疾病机制。