Department of Biochemistry and Molecular Genetics, University of Virginia School of Medicine.
Hematovascular Biology Center, Robert M. Berne Cardiovascular Research Center, University of Virginia School of Medicine.
J Vis Exp. 2021 May 26(171). doi: 10.3791/61875.
Clonal hematopoiesis is a prevalent age-associated condition that results from the accumulation of somatic mutations in hematopoietic stem and progenitor cells (HSPCs). Mutations in driver genes, that confer cellular fitness, can lead to the development of expanding HSPC clones that increasingly give rise to progeny leukocytes harboring the somatic mutation. Because clonal hematopoiesis has been associated with heart disease, stroke, and mortality, the development of experimental systems that model these processes is key to understanding the mechanisms that underly this new risk factor. Bone marrow transplantation procedures involving myeloablative conditioning in mice, such as total-body irradiation (TBI), are commonly employed to study the role of immune cells in cardiovascular diseases. However, simultaneous damage to the bone marrow niche and other sites of interest, such as the heart and brain, is unavoidable with these procedures. Thus, our lab has developed two alternative methods to minimize or avoid possible side effects caused by TBI: 1) bone marrow transplantation with irradiation shielding and 2) adoptive BMT to non-conditioned mice. In shielded organs, the local environment is preserved allowing for the analysis of clonal hematopoiesis while the function of resident immune cells is unperturbed. In contrast, the adoptive BMT to non-conditioned mice has the additional advantage that both the local environments of the organs and the hematopoietic niche are preserved. Here, we compare three different hematopoietic cell reconstitution approaches and discuss their strengths and limitations for studies of clonal hematopoiesis in cardiovascular disease.
克隆性造血是一种普遍存在的与年龄相关的疾病，它是由于造血干细胞和祖细胞（HSPCs）中的体细胞突变积累而导致的。驱动基因突变赋予细胞适应性，可能导致 HSPC 克隆的扩张，这些克隆越来越多地产生携带体细胞突变的祖细胞白细胞。由于克隆性造血与心脏病、中风和死亡率有关，因此开发模拟这些过程的实验系统对于理解这种新风险因素的机制至关重要。骨髓移植程序涉及小鼠的骨髓清除预处理，如全身照射（TBI），常用于研究免疫细胞在心血管疾病中的作用。然而，这些程序不可避免地会同时损害骨髓龛和其他感兴趣的部位，如心脏和大脑。因此，我们的实验室开发了两种替代方法来最小化或避免 TBI 可能引起的副作用：1）带有辐射屏蔽的骨髓移植，2）对非预处理的小鼠进行过继性 BMT。在屏蔽的器官中，局部环境得以保留，从而可以分析克隆性造血，同时不干扰常驻免疫细胞的功能。相比之下，对非预处理的小鼠进行过继性 BMT 还有一个额外的优势，即器官和造血龛的局部环境都得以保留。在这里，我们比较了三种不同的造血细胞重建方法，并讨论了它们在心血管疾病中研究克隆性造血的优缺点。
