Micro-Grooved Surfaces Influence Stem Cell Differentiation


Martin Knight and his colleagues from the Queen Mary’s School of Engineering and Materials Science and the Institute of Bioengineering in London, UK have shown that growing adult stem cells on micro-grooved surfaces disrupts a particular biochemical pathway that specified the length of a cellular structure called the “primary cilium.” Disruption of the primary cilium ultimately controls the subsequent behavior of these stem cells.

Primary cilia are about one thousand times narrower than a human hair. They are found in most cells and even though they were thought to be irrelevant at one time, this is clearly not the case.

Primary Cilium

The primary cilium acts as a sensory structure that responds to mechanical and chemical stimuli in the environment, and then communicates that external signal to the interior of the cell.  Most of the basic research on this structure was done using a single-celled alga called Chlamydomonas.

Martin Knight and his team, however, are certain that primary cilia in adult stem cells play a definite role in controlling cell differentiation.  Knight said, “Our research shows that they [primary cilia] play a key role in stem cell differentiation.  We found it’s possible to control stem cell specialization by manipulating primary cilia elongation, and that this occurs when stem cells are grown on these special grooved surfaces.”

When mesenchymal stromal cells were grown on grooved surfaces, the tension inside the cells was altered, and this remodeled the cytoskeleton of the cells.  Cytoskeleton refers to a rigid group of protein inside of cells that act as “rebar.” for the cell.  If you have ever worked with concrete, you will know that structural use of concrete requires the use of reinforcing metal bars to prevent the concrete from crumbling under the force of its own weight.  In the same way, cytoskeletal proteins reinforce the cell, give it shape, help it move, and help it resist shear forces.  Remodeling of the cytoskeleton can greatly change the behavior of the cell.

The primary cilium is important for stem cell differentiation.  Growing mesenchymal stromal cells on micro-grooved surfaces disrupts the primary cilium and prevents stem cell differentiation.  This simple culture technique can help maintain stem cells in an undifferentiated state until they have expanded enough for therapeutic purposes.

Once again we that there are ways to milk adult stem cells for all they are worth.  Destroying embryos is simply not necessary to save the lives of patients.

Controlling Transplanted Stem Cells from the Inside Out


Scientists have worked very hard to understand how to control stem cell differentiation.  However, despite how well you direct stem cell behavior in culture, once those stem cells have been transplanted, they will often do as they wish.  Sometimes, transplanted stem cells surprise people.

Several publications describe stem cells that, once transplanted undergo “heterotropic differentiation.” Heterotropic differentiation refers to tissues that form in the wrong place. For example, one lab found that transplantation of mesenchymal stem cells into mouse hearts after a heart attack produced bone (don’t believe me – see Martin Breitbach and others, “Potential risks of bone marrow cell transplantation into infarcted hearts.” Blood 2007 110:1362-1369).  Bone in the heart – that can’t be good. Therefore, new ways to control the differentiation of cells once they have been transplanted are a desirable goal for stem cell research.

From this motivation comes a weird but wonderful paper from Jeffrey Karp and James Ankrum of Brigham and Women’s Hospital and MIT, respectively, that loads stem cells with microparticles that give the transplanted stem cell continuous cues that tell them how to behave over the course of days or weeks as the particles degrade.

“Regardless of where the cell in the body, it’s going to be receiving its cues from the inside,” said Karp. “This is a completely different strategy than the current method of placing cells onto drug-doped microcarriers or scaffolds, which is limiting because the cells need to remain in close proximity to those materials in order to function. Also these types of materials are too large to be infused into the bloodstream.”

Controlling cells in culture is relatively easy. If cells take up the right molecules, they will change their behavior. This level of control, however, is lost after the cell is transplanted. Sometimes implanted cells readily respond to the environment within the body,. but other times, their behavior is erratic and unpredictable. Karp’s strategy, which her called “particle engineering,” corrects this problem by turning cells into pre-programmable units. The internalized particles stably remain inside the transplanted cell and instruct it precisely how to act. It can direct cells to release anti-inflammatory factors, or regenerate lost tissue and heal lesions or wounds.

“Once those particles are internalized into the cells, which can take on the order of 6-24 hours, we can deliver the transplant immediately or even cryopreserve the cells,” said Karp. “When the cells are thawed at the patient’s bedside, they can be administrated and the agents will start to be released inside the cells to control differentiation, immune modulation or matrix production, for example.”

It could take more than a decade for this type of cell therapy to be a common medical practice, but to speed up the pace of this research, Karp published the study to encourage others in the scientific community to apply the technique to their various fields. Karp’s paper also illustrates the range of different cell types that can be controlled by particle engineering, including stem cells, cells of the immune system, and pancreatic cells.

“With this versatile platform, which leveraged Harvard and MIT experts in drug delivery, cell engineering, and biology, we’ve demonstrated the ability to track cells in the body, control stem cell differentiation, and even change the way cells interact with immune cells, said Ankrum, who is a former graduate student in Karp’s laboratory. “We’re excited to see what applications other researchers will imagine using this platform.”

Microparticles and Local Control of Stem Cells


Using stem cells to grow three-dimensional structures, such as organs or damaged body parts, requires that scientists have the ability to control the growth and behavior of those cells. Also, adapting such a technology to an off-the-shelf kind of process so that it does not cost an arm and a leg is also important.

A research project by scientists from Atlanta, Georgia has used gelatin-based microparticles to deliver growth factors to specific areas of aggregates of stem cells that are differentiating. This localized delivery of growth factors provides spatial control of cell differentiation, which enables the creation of complex, three-dimensional tissues. The local delivery of growth factors also decreases the amount of growth factor used and, consequently, the cost of the procedure.

This particular microparticle technique was used on mouse embryonic stem cells and it proved to provide better control over the kinetics of cell differentiation since it delivered that promote cell differentiation or inhibit it.

Todd McDevitt, associate professor in the Wallace H. Coulter Department of Biomedical Engineering at Georgia Tech and Emory University, said, “By trapping these growth factors within microparticle materials first, we are concentrating the signal they provide to the stem cells. We can then put the microparticle materials physically inside the multicellular aggregate system that we use for differentiation for the stem cells. We have good evidence that this technique can work, and that we can use it to provide advantages in several areas.”

The differentiation of stem cells is largely controlled by external cues, including protein growth factors that direct cell proliferation, and differentiation that are available in the three-dimensional environment in which the cells live. In most experiments, stem cells are grown in liquid culture and growth factor is equally accessible to the growth factors. This makes the cultures quite homogeneous. But delivering the growth factors via microparticles gives better control of the spatial and temporal presentation of these growth factors to the stem cells. This gives scientists the means to make heterogeneous structures from stem cell cultures.

When embryonic stem cells grow in culture, they tend to clump together. When the growth medium is withdrawn or if growth factors that induce differentiation are added, the cells form an “embryoid body” that is stuff with cells differentiating into all kinds of cell types. When McDevitt and his co-workers added microparticles with the growth factors BMP4 (bone morphogen protein 4) or Noggin (which inhibits BMP4 signaling), they centrifuged the cells and found that the microparticles found their way into the interior of the embryoid bodies.

When they examined the embryoid bodies, with confocal microscopy they found that BMP4 directed the cells to make mesodermal and endodermal derived cell types. However, because the microparticles were in direct contact with the cells, they needed 12 times less growth factor than was required by solution-based techniques.

“One of the major , in a practical sense, is that we are using much less growth factor,” said McDevitt. “From a bioprocessing standpoint, a lot of the cost involved in making stem cell products is related to the cost of the molecules that must be added to make the stem cells differentiate.”

Beyond more focuses signaling, the microparticles also provided localized control that was not available through other techniques. It allowed researchers to create spatial differences in the aggregates and this is an important possible first step toward forming more complex structures with different tissue types such as vascularization and stromal cells.

“To build tissues, we need to be able to take stem cells and use them to make many cell types which are grouped together in particular spatial patterns,” explained Andres M. Bratt-Leal, the paper’s first author and a former graduate student in McDevitt’s lab. “This spatial patterning is what gives the ability to perform higher order functions.”

Once the stem cell aggregates were made and treated with growth factor-endowed microparticles, McDevitt and his colleagues saw spheres of cells with differentiating cells.

“We can see the microparticles had effects on one population that were different from the population that didn’t have the particles,” said McDevitt. “This may allow us to emulate aspects of how development occurs. We can ask questions about how tissues are naturally patterned. With this material incorporation we have the ability to better control the environment in which these cells develop.”

The microparticles could provide better control over the kinetics of cell differentiation; slowing it down with molecules that antagonize differentiation or speed up with other molecules that promote stem cell differentiation.

Despite the fact that McDevitt and his colleagues used mouse embryonic stem cells in this paper, he and his co-workers are already testing this technology on human embryonic stem cells, and the results have been comparable.

“Our findings will provide a significant new tool for tissue engineering, bioprocessing of stem cells and for better studying early development processes such as axis formation in embryos,” said Bratt-Leal. “During development, particular tissues are formed by gradients of signaling molecules. We can now better mimic these signal gradients using our system.”