Bone Marrow Niche dysfunction in sickle cell disease

镰状细胞病的骨髓生态位功能障碍

• 批准号: 10250697
• 负责人: Nadia Carlesso
• 金额: $ 2.36万
• 依托单位: BECKMAN RESEARCH INSTITUTE/CITY OF HOPE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-04 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10250697
• 关键词: Abnormal Hemoglobins; Acute Pain; Address; Affect; Age; American; Anemia; Biology; Biopsy; Blood flow; Bone Marrow; Bone Marrow Diseases; Bone Marrow Transplantation; Bone marrow biopsy; CD34 gene; Cells; Cellular Stress; Cessation of life; Chronic; Clinical; Coagulation Process; Data; Endothelial Cells; Engraftment; Enrollment; Erythrocytes; Exhibits; Frequencies; Functional disorder; Genes; Hematological Disease; Heterogeneity; Histologic; Human; IL8 gene; Immune; Infection; Inflammation; Inflammatory; Inherited; Interleukin-6; Intervention; Lead; Mesenchymal; Molecular; Molecular Analysis; Molecular Profiling; Morphology; NF-kappa B; Organ; Organ failure; Outcome; Oxidative Stress; Pain; Pain management; Pathway interactions; Patients; Phenotype; Population; Prevention; Process; Reperfusion Injury; Risk; Sampling; Severity of illness; Sickle Cell Anemia; Specimen; Stroke; TLR4 gene; Therapeutic; Time; Tissues; Transplantation; acute chest syndrome; base; chronic pain; curative treatments; cytokine; functional status; gene therapy; improved; insight; novel; novel therapeutic intervention; parent grant; premature; sickle vasoocclusion; single-cell RNA sequencing; transcriptome sequencing; transcriptomics

SUMMARY/ABSTRACT Background. Sickle cell disease (SCD) is the most common form of inherited blood disorder affecting approximately 100,000 Americans. Patients with SCD suffer from repeated red cell sickling and vaso-occlusion episodes, which, since early age, lead to acute and chronic pain, stroke, anemia, infections, organ failure and premature death. Vaso-occlusion episodes cause ischemia-reperfusion injury, activation of endothelial cells, immune cells and the coagulation cascade. These processes lead to a state of chronic inflammation in SCD, associated with increased pro-inflammatory cytokines and induction of TLR4/NF-kB activation and oxidative stress in all organs, including the bone marrow (BM). To date, little is known about how inflammation impacts the BM microenvironment and HSC functions in SCD. Notably, few and limited studies have been conducted on human samples. To address this gap, we will examine the impact of SCD in the molecular profile and functional status of bone marrow-resident cells. Preliminary results. We observed that SCD patients enrolled for transplant could be divided in two groups based on the frequency of CD34+ cells. Group 1 displayed reduced frequency of CD34+ population whereas group 2 exhibited higher or similar frequency when compared to controls. Single cell RNA-sequencing (scRNA-seq) of CD34+ enriched cells from one patient assigned in group 1 showed changes in the number of distinct cell populations and their transcriptomic profiles, with enrichment in pathway related to cellular stress. We also found that SCD BM-derived mesenchymal cells exhibit an inflammatory profile, compared to controls, characterized by increased expression of the proinflammatory related genes miR-155, miR-146a, IL-8, and IL-6 by qPCR. Furthermore, we observed that BM tissue biopsies of patients at pre-transplant show evidence of dysfunctional stroma. We hypothesize that a pro-inflammatory state induced by SCD alters the composition of the BM microenvironment and damages bone-marrow resident HSPC cells. Aims and Strategy. This study will be conducted on BM specimens of SCD patients (minimum n=10) prior HSCT and at different times after HSCT. To better understand the heterogeneity of SCD BM samples and correlate it with disease severity and clinical outcome after transplant, we propose an in-depth molecular analysis,which will be integrated with data from the parent grant. We will examine: i) the transcriptomic landscape of the HSCs in the BM of SCD patients by single cell RNA-sequencing (scRNA-seq); ii) determine the molecular pathways contributing to a pro-inflammatory phenotype in BM-derived mesenchymal cells by bulk RNA-seq, and iii) characterize the morphological and histological features in the BM biopsies of SCD patients. Relevance. This study represents a unique opportunity to better understand the biology of SCD in human specimens. It will also provide critical insights into the function of SCD CD34+ cells and the BM niche to explore new therapeutic approaches for improving current HSCT and gene therapy strategies.

摘要/摘要 背景。镰状细胞病（SCD）是影响遗传性血液疾病的最常见形式 大约有100,000名美国人。 SCD患者患有红细胞反复的疾病和血管咬合 发作，从小就导致急性和慢性疼痛，中风，贫血，感染，器官衰竭和 过早死亡。血管封闭发作会导致缺血 - 再灌注损伤，内皮细胞的激活， 免疫细胞和凝血级联。这些过程导致SCD中的慢性炎症状态， 与促炎性细胞因子增加以及TLR4/NF-KB激活的诱导和氧化有关 所有器官的压力，包括骨髓（BM）。迄今为止，关于炎症的影响知之甚少 BM微环境和HSC在SCD中起作用。值得注意的是，很少有关于 人类样品。为了解决这一差距，我们将检查SCD在分子剖面和功能中的影响 骨髓居民细胞的状态。初步结果。我们观察到SCD患者参加了 基于CD34+细胞的频率，可以将移植分为两组。第1组显示减少 与CD34+人口的频率相比，第2组的频率更高或类似 控件。 CD34+富集细胞的单细胞RNA - 序列（SCRNA-SEQ）来自组中的一名患者 1显示了不同细胞群体数量及其转录谱的变化，并具有丰富 与细胞应激有关的途径。我们还发现，SCD BM衍生的间充质细胞表现出 与对照组相比，炎症性特征的特征是促炎相关的表达增加 基因miR-155，miR-146a，IL-8和IL-6的QPCR。此外，我们观察到患者的BM组织活检 在移植前，显示了功能失调的基质的证据。我们假设促炎性状态引起 通过SCD改变了BM微环境的组成，并损坏了骨row的居民HSPC细胞。 目的和策略。这项研究将在先前对SCD患者的BM标本（至少n = 10）进行 HSCT和HSCT之后的不同时间。更好地了解SCD BM样品的异质性和 将其与疾病的严重程度和移植后的临床结果相关，我们提出了一个深入的分子 分析将与父母赠款的数据集成。我们将检查：i）转录组 通过单细胞RNA-Serecting（SCRNA-Seq），SCD患者BM中HSC的景观； ii）确定 通过大量促进BM衍生间充质细胞中促炎表型的分子途径 RNA-Seq和III）表征了SC​​D患者BM活检中的形态学和组织学特征。 关联。这项研究代表了更好地了解人类SCD生物学的独特机会 标本。它还将提供有关SCD CD34+细胞和BM域名功能的关键见解以探索 改善当前HSCT和基因治疗策略的新治疗方法。