Biology subjects

Deans, T. L.

Publications and source records attributed to Deans, T. L..

4 recordsLinked to original sources

bioRxiv2023

Reprogramming megakaryocytes to engineer platelets as delivery vehicles

Platelets are anucleate cells naturally filled with secretory granules that store large amounts of protein to be released in response to certain physiological conditions. Cell engineering can endow platelets with the ability to deliver non-native proteins by modifying them as they develop during the cell fate process. This study presents a strategy to efficiently generate mouse platelets from pluripotent stem cells and demonstrates their potential as bioengineered protein delivery platforms. By modifying megakaryocytes, the progenitor cells of platelets, we successfully engineered platelets capable of packaging and delivering non-native proteins. These engineered platelets can offer flexible delivery platforms to release non-native proteins in a controlled manner upon activation when packaged into -granules or deliver active enzymes to genetically alter recipient cells. Our findings highlight platelets as a promising tool for protein delivery in cell therapy applications.

synthetic biology↗

bioRxiv2023

In vitro generation of megakaryocytes from engineered mouse embryonic stem cells

The in vitro differentiation of pluripotent stem cells into desired lineages enables mechanistic studies of cell transitions into more mature states that can provide insights into the design principles governing cell fate control. We are interested in reprogramming pluripotent stem cells with synthetic gene circuits to drive mouse embryonic stem cells (mESCs) down the hematopoietic lineage for the production of megakaryocytes, the progenitor cells for platelets. Here, we describe the methodology for growing and differentiating mESCs, in addition to inserting a transgene to observe its expression throughout differentiation. This entails four key methods: (1) growing and preparing mouse embryonic fibroblasts for supporting mESC growth and expansion, (2) growing and preparing OP9 feeder cells to support the differentiation of mESCs, (3) the differentiation of mESCs into megakaryocytes, and (4) utilizing an integrase mediated docking site to insert transgenes for their stable integration and expression throughout differentiation. Altogether, this approach demonstrates a streamline differentiation protocol that emphasizes the reprogramming potential of mESCs that can be used for future mechanistic and therapeutic studies of controlling cell fate outcomes.

synthetic biology↗

bioRxiv2020

Rolling Signal-based Ripley's K: A new algorithm to identify spatial patterns in histological specimens

The spatial distribution of cells within a tissue underlies organ function. However, these spatial distributions are often difficult to identify, making it challenging to evaluate how cells establish these patterns during development or how diseases may disrupt these patterns and impair function. To address this, we developed an image analysis tool based on a novel algorithm that identifies spatial patterns within tissues. This analytical tool was used to study the bone marrow, a specialized microenvironment in which spatial patterning of regulatory cells may influence the differentiation and survival of hematopoietic stem cells. Using this algorithm, we discovered clusters of regulatory cells within the bone marrow that suggest an organization of micro-niches, which may form the basis of the hematopoietic stem cell microenvironment. This work provides a new tool for the detection and analysis of tissue morphology that enables identification of spatial patterns within tissues that can lead to a deeper understanding of tissue function, provide clues for early onset of disease, and be used as a tool for studying the impact of pharmaceutics on tissue development and regeneration. In BriefThis work introduces a new statistic to analyze the patterning of cells and physiological features in histological images. This statistic was used on a published set of immunofluorescent images of murine bone to identify novel spatial structures within the bone marrow that may provide new inisghts to the organization of the hematopoietic stem cell microenvironment. HighlightsO_LIRSRK, a statistical tool for analyzing the spatial distribution of features in histological images, is introduced. C_LIO_LIRSRK incorporates the quantification of signal distribution to identify unique spatial patterns. C_LIO_LISpatial patterns in hematopoietic stem cell microenvironments are identified. C_LI Graphical Abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

systems biology↗

bioRxiv2019

Combination image flow cytometry for single-cell analysis reveals novel methods for isolating subsets of megakaryocyte progenitor populations

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 principle components 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.

cell biology↗