Megakaryocytes are a rare population of cells that develop in the bone marrow and function to produce platelets that circulate throughout the body and form clots to stop or prevent bleeding. A major challenge in studying megakaryocyte development, and the diseases that arise from their dysfunction, is the identification, classification, and enrichment of megakaryocyte progenitor cells that are produced during hematopoiesis. Here, we present a high throughput strategy for identifying and isolating megakaryocytes and their progenitor cells from a heterogeneous population of bone marrow samples. Specifically, we couple thrombopoietin (TPO) induction, image flow cytometry, and principal component analysis (PCA) to identify and enrich for megakaryocyte progenitor cells that are capable of self-renewal and directly differentiating into mature megakaryocytes. This enrichment strategy distinguishes megakaryocyte progenitors from other lineage-committed cells in a high throughput manner. Furthermore, by using image flow cytometry with PCA, we have identified a combination of markers and characteristics that can be used to isolate megakaryocyte progenitor cells using standard flow cytometry methods. Altogether, these techniques enable the high throughput enrichment and isolation of cells in the megakaryocyte lineage and have the potential to enable rapid disease identification and diagnoses ahead of severe disease progression.
Synthetic biology is a multidisciplinary field that focuses on the rational design and construction of novel genetic tools for the purpose of engineering cells to behave in controllable and predictable ways. The promise of this modern technology relies on our understanding of basic genetics and gene expression to engineer cells with unique functions. This is accomplished by designing biological parts and assembling them into higher-order gene circuits that control cell operations through tight regulation of gene expression, effectively reprogramming and rewiring the cells. In this article, we review the basic concepts of gene expression, discuss the framework of how synthetic biologists reprogram cells and outline how cells can be engineered to function as new vehicles for delivering therapeutic proteins.
6Megakaryocytes (MKs) are rare cells that develop in the bone marrow and give rise to platelets. A 7 major challenge in studying megakaryocyte development, and the diseases that arise from these cells, 8 is the identification and classification of MK progenitor cells that are produced during hematopoiesis.
9Here, we utilize combination image flow cytometry and principal component analysis (PCA) to
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