Method to deliver molecules within embryonic stem cells improves differentiation

Embryonic stem cell therapies have been proposed for regenerative medicine and tissue replacement after injury or disease. However, the inability of stem cells to efficiently develop into the desired specific cell type – such as muscle, skin, blood vessels, bone or neurons – now limits the potential clinical utility of this therapy.

New research shows that delivering molecules within aggregates of embryonic stem cells via biodegradable microspheres enhances the efficiency and purity of differentiation, which is the process the cells undergo to become more specialized. Details of the microsphere-mediated delivery method, which is funded by the National Science Foundation, were presented on April 9 at the 235th American Chemical Society national meeting.

“Directing embryonic stem cells to efficiently differentiate into a specific cell type has been challenging to this point,” said Todd McDevitt, an assistant professor in the Wallace H. Coulter Department of Biomedical Engineering at Georgia Tech and Emory University. “In my lab, we’re trying to better define and then control the environmental cues that regulate the fate and function of the stem cells.”

Because physical interactions between stem cells is critical during normal embryonic development, most laboratory growth methods allow the cells to aggregate in three-dimensional clumps called “embryoid bodies” in order to differentiate. After individual cells aggregate together, hollow internal structures begin to develop and the aggregate becomes larger and more complex over time.

“Many researchers add soluble factors to the culture dish medium to direct differentiation, but this does not accurately mimic the time and location of signaling events present in normal development, and may contribute to heterogeneous differentiation,” said McDevitt. “Our method focuses on incorporating the differentiation factors directly into the cell aggregates in order to have a more controlled mechanism of presentation.”

The research team – which also includes graduate students Richard Carpenedo and Andrés Bratt-Leal and undergraduate students Ross Marklein and Scott Seaman – fabricated biodegradable polymer microspheres that could contain growth factors, proteins or other small molecules.

McDevitt’s team tested the impact of the poly(lactic-co-glycolic acid) (PLGA) microspheres on embryonic stem cell differentiation under different conditions by varying the microsphere-to-cell ratio and speed at which the aggregate cells were mixed with the microspheres. They also included a fluorescent dye in the microspheres so the degree of incorporation of the microspheres within the embryoid bodies could be assessed using fluorescent microscopy and spectroscopy.

The results revealed that the microspheres were incorporated into embryoid bodies under a variety of mixing conditions, but that slower rotary speeds and higher microsphere-to-cell ratios resulted in a greater degree of incorporation.

Next, the researchers compared differentiation of untreated cells, cells mixed with empty microspheres, cells mixed with retinoic acid-loaded microspheres, and cells treated with soluble retinoic acid. Retinoic acid was chosen initially because it is a potent inducer of embryonic stem cell differentiation.

After ten days, approximately 90 percent of the embryoid bodies mixed with retinoic acid-loaded microspheres began to display the hollow structure signifying differentiation, compared to 6 percent of the untreated bodies, 10 percent of the bodies coated with soluble retinoic acid, and 30 percent of the bodies mixed with empty microspheres. In addition, thirty percent of the embryoid bodies mixed with retinoic acid-loaded microspheres were completely hollow in the center, compared to nearly zero percent for the other groups.

“These results suggest that if you can control the signaling by presenting molecules locally on the inside of the embryoid body from biodegradable microspheres, you can effectively change the course and synchrony of differentiation,” said McDevitt.

To examine the cells in more detail, McDevitt teamed with Georgia Tech School of Biology chair John McDonald and research scientist Nathan Bowen to conduct microarray gene expression studies to determine cell phenotype.

The results revealed enhanced expression of fibroblast growth factor 5 (FGF-5) – a marker for primitive ectoderm – in the embryoid bodies mixed with retinoic acid-loaded microspheres compared to the other treatment groups after 10 days. The researchers also confirmed increased or inhibited expression of many additional markers.

“The importance of these findings is that we’ve shown that biomaterial-based approaches to regulate stem cell microenvironments can significantly improve differentiation methods,” said McDevitt. “Our ultimate goal is to improve the efficiency of this differentiation process into specific cell types for cell replacement therapies.”

Source: Georgia Institute of Technology

Related stories:

Coatings to help medical implants connect with neurons
Plastic coatings could someday help neural implants treat conditions as diverse as Parkinson's disease and macular degeneration.
'Smart' materials get smarter with ability to better control shape and size
A dynamic way to alter the shape and size of microscopic three-dimensional structures built out of proteins has been developed by biological chemist Jason Shear and his former graduate student Bryan Kaehr at The University of Texas at Austin.
New vaccine approach prevents/reverses diabetes in lab study
Results of study are published in Diabetes, a journal of the American Diabetes Association
Microspheres carrying targeted nucleic acid molecules fabricated in the laboratory have been shown to prevent and even reverse new-onset cases of type 1 diabetes in animal models. The results of these studies were reported by diabetes researchers at the John G. Rangos Sr. Research Center at Children’s Hospital of Pittsburgh of UPMC and Baxter Healthcare Corporation.
Single-cell sensitive biological sensor works in liquid
Microscopic, magnetized balls of Styrofoam have been turned into inexpensive biological sensors in a University of Michigan laboratory.
Seaweed Transformed Into Stem Cell Technology
Engineers at Rensselaer Polytechnic Institute have transformed a polymer found in common brown seaweed into a device that can support the growth and release of stem cells at the sight of a bodily injury or at the source of a disease.
MIT works toward novel therapeutic device
MIT and University of Rochester researchers report important advances toward a therapeutic device that has the potential to capture cells as they flow through the blood stream and treat them. Among other applications, such a device could zapp cancer cells spreading to other tissues, or signal stem cells to differentiate.
New clues to air circulation in the atmosphere
Air circulates above the Earth in four distinct cells, with two either side of the equator, says new research. The new observational study describes how air rises and falls in the atmosphere above the Earth's surface, creating the world's weather.
Accumulated bits of a cell's own DNA can trigger autoimmune disease
A security system wired within every cell to detect the presence of rogue viral DNA can sometimes go awry, triggering an autoimmune response to single-stranded bits of the cell's own DNA, according to a report in the August 22nd issue of the journal Cell, a Cell Press publication. The source of that single-stranded DNA is so-called endogenous retroelements—genetic elements accounting for a substantial portion of the genome that can move to new locations using a "copy and paste" mechanism, according to the researchers.

News discussion:

General Science news