Biomechanical Stimulation of Hemogenic Endothelial Cells to Develop Clinical-Grade Hematopoietic Stem Cells

生物力学刺激造血内皮细胞发育临床级造血干细胞

• 批准号: 9077910
• 负责人: DHVANIT INDRAVADAN SHAH
• 金额: $ 42.89万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-05-01 至 2021-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9077910
• 关键词: Angiography; Aorta; Biomechanics; Blood; Blood Circulation; Blood Vessels; Blood flow; Bone Marrow; Bone Marrow Diseases; Cardiac Output; Cations; Cells; Child; Clinical; Data; Development; Disease; Doppler Ultrasound; Echocardiography; Embryo; Endothelial Cells; Endothelium; Engraftment; Family; Fetal Development; Fetal Liver; Foundations; Gene Expression; Gene Expression Profiling; Gonadal structure; Grant; Heart; Hematological Disease; Hematopoiesis; Hematopoietic; Hematopoietic Neoplasms; Hematopoietic Stem Cell Transplantation; Hematopoietic stem cells; Human; Image; Immune System Diseases; Immunologics; In Situ; Incubated; Inherited; Ion Channel; Mediating; Membrane; Mesonephric structure; Methods; Modeling; Molecular; Mus; Nylons; Organ; Organ Culture Techniques; Parents; Patients; Process; Property; Pulse Pressure; Race; Role; Signal Transduction; Source; Stem Cell Development; Stem cell transplant; Stretching; Suspension substance; Suspensions; Therapeutic Uses; Time; Tissues; Transgenic Organisms; Ultrasonography; Umbilical cord structure; Vanilloid; Yin; Zebrafish; base; biracial; cellular engineering; digital; embryo culture; epithelial Na+ channel; flexibility; human tissue; member; mouse model; novel; public health relevance; receptor; shear stress; Angiography; Aorta; Biomechanics; Blood; Blood Circulation; Blood Vessels; Blood flow; Bone Marrow; Bone Marrow Diseases; Cardiac Output; Cations; Cells; Child; Clinical; Data; Development; Disease; Doppler Ultrasound; Echocardiography; Embryo; Endothelial Cells; Endothelium; Engraftment; Family; Fetal Development; Fetal Liver; Foundations; Gene Expression; Gene Expression Profiling; Gonadal structure; Grant; Heart; Hematological Disease; Hematopoiesis; Hematopoietic; Hematopoietic Neoplasms; Hematopoietic Stem Cell Transplantation; Hematopoietic stem cells; Human; Image; Immune System Diseases; Immunologics; In Situ; Incubated; Inherited; Ion Channel; Mediating; Membrane; Mesonephric structure; Methods; Modeling; Molecular; Mus; Nylons; Organ; Organ Culture Techniques; Parents; Patients; Process; Property; Pulse Pressure; Race; Role; Signal Transduction; Source; Stem Cell Development; Stem cell transplant; Stretching; Suspension substance; Suspensions; Therapeutic Uses; Time; Tissues; Transgenic Organisms; Ultrasonography; Umbilical cord structure; Vanilloid; Yin; Zebrafish; base; biracial; cellular engineering; digital; embryo culture; epithelial Na+ channel; flexibility; human tissue; member; mouse model; novel; public health relevance; receptor; shear stress

 DESCRIPTION (provided by applicant): The therapeutic use of hematopoietic stem cell (HSC) transplantation for inherited or acquired blood diseases is often restricted due to the difficulty with finding HLA-matched donors and immunologic intolerance between donor and recipient. Therefore, there is a critical need to identify new methods to generate high throughput, clinical-grade functional HSCs from human tissues. The objective of this application is to analyze novel cell-extrinsic mechanisms that stimulate the developmental transition from hemogenic endothelial cells to HSCs. We recently established that functional HSCs emerge in the cdh5-deleted zebrafish embryos, despite an impaired blood circulation. These findings led us to investigate shear-stress independent biomechanical mechanisms stimulating endothelial to HSC transition. Using echocardiography, micro-angiography, 3D Doppler, and confocal imaging, we demonstrate that heartbeat mediated pulse-pressure, and thus circumferential stretch stimulates HSC formation. Pharmacological inhibition of stretch-activated ion channels further reduced HSC formation. We, therefore, hypothesize that pulse-pressure mediated circumferential stretch stimulates the transition of endothelial cells to HSCs by stimulating stretch-activated ion channels. We will use advances in organ-in-a-dish and cell-engineering to recapitulate in situ circumferential stretch conditions on murine hemogenic endothelial cells. Since the silent-heart (tnnt2, sih)-silenced embryos lack heartbeat and blood circulation, we will also investigate if pharmacological stimulation of stretch-activated ion channels could rescue hematopoietic deficiency in the sih-silenced embryos. In addition, we will use transgenic zebrafish, ex vivo mouse embryo and explant culture methods, as well as confocal imaging to analyze how stimulation of stretch activated ion channels, Trpv4 and/or Piezo1, influences the emergence of functional HSCs. Our aims will establish a new conserved cell-extrinsic role and mechanisms of circumferential stretch during HSC formation. Identification of new factors stimulating hemogenic properties of endothelium would expedite the development of endothelial cells as a source of HSCs for the treatment of human blood diseases.

 描述（通过应用提供）：由于难以在供体和受体之间找到HLA匹配的供体和免疫学性供体，造血干细胞（HSC）移植对遗传或获得的血液疾病的理论使用通常受到限制。因此，迫切需要确定从人体组织产生高通量的临床级功能HSC的新方法。该应用的目的是分析新型的细胞超支机制，这些机制刺激了从血浆内皮细胞到HSC的发育过渡。我们最近确定，尽管血液循环受损，但在CDH5骨骨鱼胚胎的胚胎中出现了功能性HSC。这些发现使我们研究了刺激内皮到HSC过渡的剪切压力独立的生物力学机制。使用超声心动图，微血管造影，3D多普勒和共聚焦成像，我们证明了心跳介导的脉冲压力，因此圆周拉伸刺激了HSC的形成。拉伸激活离子通道的药理抑制进一步降低了HSC的形成。因此，我们假设脉冲压力介导的圆周拉伸通过刺激拉伸激活的离子通道刺激内皮细胞向HSC的过渡。我们将利用在静脉内和细胞工程的器官中的进步来概括鼠血质内皮细胞的原位圆周伸展条件。由于沉默的心脏（TNNT2，SIH）缺乏心血管缺乏心跳和血液循环，因此我们还将调查是否可以刺激拉伸激活的离子通道的药物刺激，可以挽救SIH脱毛的胚胎中的造血性造血缺乏。此外，我们将使用转基因斑马鱼，离体小鼠胚胎和外植体培养方法，以及共焦成像进行分析，分析拉伸激活的离子通道，TRPV4和/或PIEZO1如何影响功能HSC的出现。我们的目标将在HSC形成期间建立新的保守细胞肢体作用和圆周拉伸的机制。鉴定刺激森林血质特性的新因素将加快森林细胞的发展，作为治疗人类血液疾病的HSC的来源。