Benjamin E. Mead

Engineering Stem Cell Organoids.

Organoid systems leverage the self-organizing properties of stem cells to create diverse multi-cellular tissue proxies. Most organoid models only represent single or partial components of a tissue, and it is often difficult to control the cell type, organization, and cell-cell/cell-matrix interactions within these systems. Herein, we discuss basic approaches to generate stem cell-based organoids, their advantages and limitations, and how bioengineering strategies can be used to steer the cell composition and their 3D organization within organoids to further enhance their utility in research and therapies.

Generating iPSCs: translating cell reprogramming science into scalable and robust biomanufacturing strategies.

Induced pluripotent stem cells (iPSCs) have the potential to transform drug discovery and healthcare in the 21(st) century. However, successful commercialization will require standardized manufacturing platforms. Here we highlight the need to define standardized practices for iPSC generation and processing and discuss current challenges to the robust manufacture of iPSC products.

Culturing human intestinal stem cells for regenerative applications in the treatment of inflammatory bowel disease

Both the incidence and prevalence of inflammatory bowel disease (IBD) is increasing globally; in the industrialized world up to 0.5% of the population are affected and around 4.2 million individuals suffer from IBD in Europe and North America combined. Successful engraftment in experimental colitis models suggests that intestinal stem cell transplantation could constitute a novel treatment strategy to re‐establish mucosal barrier function in patients with severe disease. Intestinal stem cells can be grown in vitro in organoid structures, though only a fraction of the cells contained are stem cells with regenerative capabilities. Hence, techniques to enrich stem cell populations are being pursued through the development of multiple two‐dimensional and three‐dimensional culture protocols, as well as co‐culture techniques and multiple growth medium compositions. Moreover, research in support matrices allowing for efficient clinical application is in progress. In vitro culture is accomplished by modulating the signaling pathways fundamental for the stem cell niche with a suitable culture matrix to provide additional contact‐dependent stimuli and structural support. The aim of this review was to discuss medium compositions and support matrices for optimal intestinal stem cell culture, as well as potential modifications to advance clinical use in IBD.

Harnessing single-cell genomics to improve the physiological fidelity of organoid-derived cell types

BackgroundSingle-cell genomic methods now provide unprecedented resolution for characterizing the component cell types and states of tissues such as the epithelial subsets of the gastrointestinal tract. Nevertheless, functional studies of these subsets at scale require faithful in vitro models of identified in vivo biology. While intestinal organoids have been invaluable in providing mechanistic insights in vitro, the extent to which organoid-derived cell types recapitulate their in vivo counterparts remains formally untested, with no systematic approach for improving model fidelity.ResultsHere, we present a generally applicable framework that utilizes massively parallel single-cell RNA-seq to compare cell types and states found in vivo to those of in vitro models such as organoids. Furthermore, we leverage identified discrepancies to improve model fidelity. Using the Paneth cell (PC), which supports the stem cell niche and produces the largest diversity of antimicrobials in the small intestine, as an exemplar, we uncover fundamental gene expression differences in lineage-defining genes between in vivo PCs and those of the current in vitro organoid model. With this information, we nominate a molecular intervention to rationally improve the physiological fidelity of our in vitro PCs. We then perform transcriptomic, cytometric, morphologic and proteomic characterization, and demonstrate functional (antimicrobial activity, niche support) improvements in PC physiology.ConclusionsOur systematic approach provides a simple workflow for identifying the limitations of in vitro models and enhancing their physiological fidelity. Using adult stem cell-derived PCs within intestinal organoids as a model system, we successfully benchmark organoid representation, relative to that in vivo, of a specialized cell type and use this comparison to generate a functionally improved in vitro PC population. We predict that the generation of rationally improved cellular models will facilitate mechanistic exploration of specific disease-associated genes in their respective cell types.

Bioprocess Decision Support Tool For Scalable Manufacture of Extracellular Vesicles: NG et al.

Newly recognized as natural nanocarriers that deliver biological information between cells, extracellular vesicles (EVs), including exosomes and microvesicles, provide unprecedented therapeutic opportunities. Large‐scale and cost‐effective manufacturing is imperative for EV products to meet commercial and clinical demands; successful translation requires careful decisions that minimize financial and technological risks. Here, we develop a decision support tool (DST) that computes the most cost‐effective technologies for manufacturing EVs at different scales, by examining the costs of goods associated with using published protocols. The DST identifies costs of labor and consumables during EV harvest as key cost drivers, substantiating a need for larger‐scale, higher‐throughput, and automated technologies for harvesting EVs. Importantly, we highlight a lack of appropriate technologies for meeting clinical demands, and propose a potentially cost‐effective solution. This DST can facilitate decision‐making very early on in development and be used to predict, and better manage, the risk of process changes when commercializing EV products.