Blood vessel assembly from multipotent hemangioma-derived stem cells

项目摘要 摘要婴儿血管瘤是一种常见的血管肿瘤，具有快速血管形成的独特生命周期。 在婴儿期的前6-9个月，随后几个月血管缓慢自发退化 好几年了。对于大多数儿童来说，IH并不构成严重威胁，治疗是不必要的；然而，在大约 在10%的病例中，IH可显著增大，威胁器官，并导致永久性毁容。在过去的几年里 普萘洛尔是一种著名的非选择性β肾上腺素能受体拮抗剂，10年来已成为一线治疗药物 危害IH的治疗方法，然而它如何以及为什么在减少IH血管过度生长方面效果如此好 仍然是个谜。有必要改进心得安治疗，因为高达18%的IHS失败 作为回应，当药物停止时，高达25%的婴儿恢复生长，37%的普萘洛尔治疗的婴儿需要 在5-6岁时进行手术，以减少残留的纤维脂肪造成的畸形。改进的方法 心得安，有必要阐明它的作用机制，以防止血管过度生长，然后 为推进IH的医学治疗，以及潜在的其他新生血管疾病提供了一条前进的道路。 在之前的资助周期中，我们从人类IH手术标本中鉴定出一种血管瘤干细胞(HemSC)。 可分化为内皮细胞、周细胞和脂肪细胞，并在体内形成血管瘤样血管 7天后植入免疫缺陷小鼠体内。我们实验室和其他实验室的后续研究证实 HbSCs作为IH的启动细胞。我们最近的研究结果表明，一种小分子转录抑制物 因子SOX18和心得安均能有效地阻断向内皮细胞的分化。此外， 心得安的R(+)对映体缺乏β肾上腺素能受体拮抗活性，同样有效。 这一新发现确定了一种β肾上腺素能受体不依赖、SOX18依赖的机制 心得安可减少IH患者的血管过度生长。为了深入研究，我们提出了三个具体目标。目标1 将使用我们的体内模型直接而严格地测试在IH血管形成中对SOX18的要求 该细胞在裸鼠体内形成类IH血管。目标2将调查二聚化状况 在IH中的SOX18(次级目标2a)，心得安和R(+)对映体如何破坏SOX18的二聚化和亚目标 细胞定位(亚目标2b)，以及这如何改变转录以防止血液干细胞血管形成 (次级目标2c)。并行进行的AIM 3将使用以下工具分析我们现有的下一代测序数据 新的生物信息学工具用于识别潜在的染色体易位或小拷贝数变异 可产生具有新活动的融合转录本(次级目标3a)，并将深入报道RNA-Seq Ih组织和新分离的Ih细胞作为鉴定融合转录本的替代方法(次级目标3b)；一次 确定了与SOX18和IH血管形成的联系将在体外和体内模型中进行测试。

Project Abstract Infantile hemangioma (IH) is a common vascular tumor with a unique lifecycle of rapid blood vessel formation over the first 6-9 months of infancy, followed by a slow spontaneous involution of blood vessels over several years. For most children, IH does not pose a serious threat and therapy is unnecessary; however, in about 10% of cases, IH can enlarge dramatically, threaten organs and cause permanent disfigurement. Over the last 10 years, propranolol, a well-known non-selective β-adrenergic receptor antagonist, has emerged as first-line therapy for endangering IH, yet how and why it works so well in reducing the vascular overgrowth in IH has remained a mystery. There is a significant need to improve propranolol therapy because up to 18% of IHs fail to respond, up to 25% resume growth when the drug is stopped, and 37% of propranolol-treated infants require surgery at 5-6 years of age to minimize deformity caused by remaining fibrofatty residua. To improve on propranolol, it is essential to elucidate it’s mechanism of action against vascular overgrowth, which will then provide a path forward to advance IH medical therapy, and potentially other neovascular diseases as well. In previous funding cycles, we identified a hemangioma stem cell (HemSC) from human IH surgical specimens that can differentiate into endothelial cells, pericytes and adipocytes and form hemangioma-like vessels within 7 days when implanted into immune-deficient mice. Subsequent studies from our lab and others validate HemSCs as the IH-initiating cell. Our recent results show that a small molecule inhibitor of the transcription factor SOX18 and propranolol both effectively block HemSC-to-endothelial differentiation. Furthermore, the R(+) enantiomer of propranolol, which lacks β-adrenergic receptor antagonistic activity, is equally effective. This novel discovery identifies a β-adrenergic receptor-independent, SOX18-dependent mechanism by which propranolol reduces vascular overgrowth in IH. To investigate deeply, we propose three specific aims. Aim 1 will directly and rigorously test the requirement for SOX18 in IH vessel formation using our in vivo model in which IH-derived HemSC form IH-like blood vessels in nude mice. Aim 2 will investigate dimerization status of SOX18 in IH (sub-aim 2a), how propranolol and the R(+) enantiomer disrupt SOX18 dimerization and sub- cellular localization (sub-aim 2b), and how this alters transcription to prevent HemSC-blood vessel formation (sub-aim 2c). Aim 3, conducted in parallel, will analyze our existing next generation sequencing data using new bioinformatic tools to identify potential chromosomal translocations or small copy number variants that could produce fusion transcripts with new activities (sub-aim 3a) and will perform deep coverage RNA-Seq on IH tissue and freshly isolated IH cells as an alternative method to identify fusion transcripts (sub-aim 3b); once identified, the connection to SOX18 and IH vessel formation will tested in in vitro and in vivo models.