The Niche

ISSCR Friday posters: cell-penetrators, differentiators, memory-storers, and more

NOTE: These write-ups of selected ISSCR posters are by Teisha Rowland, a volunteer Niche blogger and student at UC Santa Barbara.

They include the following:

Culturing MSCs in spheres may boost their differentiation capacity (Bray, Schilling, Burdon, Genever)

Small-molecule primer for ESC differentiation (Zhu, Wurdak, Wang, Schultz)

Alternative to iPSCs: sometimes fusion goes faster (Schneider, Zenke, et al)

A silky cell-penetrating protein for producing iPS cells (Park, Park, Park)


ISSCR poster: Culturing MSCs in spheres may boost their differentiation capacity

Although mesenchymal stem cells (MSCs) are thought to have restricted, defined multipotent fates, some recent evidence suggests that certain conditions could boost their differentiation potential. In particular, the standard two-dimensional culture conditions might actually limit their potential by failing to recapitulate certain aspects of in vivo conditions. Bray et al. found that culturing MSCs as spheroids changes their gene expression. The researchers generated spheroids by growing MSCs in wells of an untreated plastic culture plate under a range of environmental conditions and cell seeding densities. After 4 to 5 days, a 10-100 fold increase in the expression of several embryonic genes (Oct4, Nanog, Sox2, Rex1, and Lin28) was observed compared to cells grown under standard 2-D culture conditions. Furthermore, the spheroid MSCs appeared to more efficiently differentiate into multiple lineages; under conditions for inducing cardiomyocytes and neurons, the spheroid MSCs produced much higher levels of markers for these tissues than MSCs grown in 2-D culture. However, teratoma formation was not observed 12 weeks post injection into a nude mouse, a standard assay for pluriptotency. Such MSC spheroids, that may more closely resemble MSCs in vivo, could someday be of use in studying differentiation as well as in cell therapies.

Title: Induced Potency of Human Mesenchymal Stem Cells in a Defined 3D in Vitro Environment (#464)

Authors: Bray, Elen, Schilling, Tatjana, Burdon, Drew, Genever, Paul

The University of York and Smith and Nephew Research Centre, UK

ISSCR poster: Small-molecule primer for ESC differentiation

Differentiating embryonic stem cells (ESCs) into a specific target cell type is often not very efficient; it can require a significant amount of time even with the best known culture conditions. The authors of this poster initially screened many small molecules to look for one that increases levels of a protein known to be highly expressed during tissue differentiation in development, specifically Sox17. Zhu et al. discovered a small molecule, called stauprimide, that may increase efficiency of ESC differentiation. Compared to cultures lacking the small molecule, ESCs differentiated more quickly into the neural and mesoderm lineages in a dose-dependent manner when stauprimide was added to ESC culture under appropriate media conditions for differentiation to these lineages. Further work showed that stauprimide binds to and inhibits nuclear localization of Non-Metastatic Cells 2 (NME2), which when inhibited causes down-regulation of c-Myc expression. Overall, the authors suggest that stauprimide could be used as a tool to study ESCs and prime them for differentiation.

Title: Small Molecule Primes Mouse and Human Embryonic Stem Cells for Differentiation by Targeting NME2 (#748)

Authors: Zhu, Shoutian, Wurdak, Heiko, Wang, Jian, Schultz, Peter G.

Affiliation: The Scripps Research Institute, La Jolla, California

ISSCR poster: Alternative to iPSCs: sometimes fusion goes faster

With all the attention induced pluripotent stem cells (iPSCs) get, it’s easy to forget that there are other, possibly better routes to cells of interest. In fact, methods besides reprogramming might produce some desired cells more quickly and in greater numbers. To that end, this poster reported that fusing hematopoietic stem cells (HSCs) isolated from mouse bone marrow with embryonic stem cells (ESCs) results in unique tetraploid hybrids with the propensity to differentiate back into HSCs. The fused cells have ESC morphology and can differentiate into the three germ layers. The fused cells express genes characteristic of both parent cell types, as well as a unique set of genes. When these hybrids were allowed to undergo spontaneous and directed differentiation, they showed a preference for hematopoietic cell differentiation, indicating some memory of their previous state as HSCs. Contrary to these findings, previous studies have suggested that such embryonic fusions lose their somatic memory, while in the related iPSC field it is also unclear whether cells prefer to differentiate into their non-reprogrammed precursors; much work remains to be done before researchers know whether or not they should start with the cell type they want to end up with.

Title: Somatic Memory in Pluripotent Hybrids of Mouse Hematopoietic Stem Cells and ES Cells (#904)

Authors: Schneider, Rebekka K., Ensenat-Waser, Roberto, Breitbach, Martin, Do, Jeong Tae, Fugemann, Christoph, Kuzmenkin, Alexey, Ruau, David, Wang, Mengxi, Xu, Guoxing, Fleischmann, Bernd K., Hescheler, Jurgen, Scholer, Hans R., Zenke, Martin

RWTH Aachen University Medical School, University of Bonn, Max Planck Institute for Molecular Medicine, University of Cologne

ISSCR poster: a silky cell-penetrating protein for producing iPS cells

Cell-permeable peptides and proteins offer great potential as a non-integrative system for the creation of induced pluripotent stem cells (iPSCs). This poster reported the potential application of the recently discovered cell-penetrating ability of 30K protein, which is found in the plasma of silkworms. The researchers found that if this 30K protein is added to culture media, it makes its way into the cell. Additionally, the researchers fused green fluorescent protein (GFP) to the 30K protein and demonstrated that the fusion protein is transported into the cell: HeLa cells cultured with media containing 30K protein-tagged GFP have observable GFP inside the cells, though not clearly in the nucleus. Next, the researchers combined 30K protein with each of four reprogramming factors (recombinant proteins Oct3/4, Sox2, c-Myc, and Klf4) and added them to culture media. The combination proteins were transported inside HEK293T, HeLa, and human fibroblast cells. And concentrations of the 30K conjugated proteins can be tuned over time: when media that does not contain 30K protein is added to the culture, the protein is exported out of the cells, implying that the 4 factors could also be removed from the cell with a simple change of media. The authors anticipate that using 30K protein in this way could lead to better methods to create iPSCs without requiring genetic modification.

Poster #906

Title: Attempt for Generation of Induced Pluripotent Stem Cells Using Recombinant 4 Factors Conjugated to Cell-Permeable 30K Protein Without Gene Integration (#906)

Authors: Park, Ju Hyun, Park, Hee Ho, Park, Tai Hyun

Seoul National University, Korea

These account is by Teisha Rowland, a student at UC Santa Barbara who uses hESCs and iPSCs. She runs a blog called allthingsstemcell.com

Note from Niche editor This post comes as a response to my solicitation in June calling for people to submit their accounts of ISSCR 2009. I’d asked people to describe what most interested them and to disclose any conflicts of interest. I’m very grateful for these volunteers’ help making more information available.

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