Stem Cells and Their Fat Neighbors

We recently published a PLOS ONE paper (Mitochondrial respiration regulates adipogenic differentiation of human mesenchymal stem cells) in which we studied how the metabolism of an adult stem cell can influence its ability to differentiate. Human bone marrow mesenchymal stem cells (also known as marrow stromal cells, marrow progenitor cells or MSCs) can be converted into fat (adipocytes), cartilage (chondrocytes) or bone (osteoblasts). The work performed by Yanmin Zhang and Glenn Marsboom in my lab showed that MSCs undergo a major metabolic shift towards increased mitochondrial oxidation when they become fat cells and that suppressing mitochondrial respiration can prevent their differentiation. The metabolic state of the adult stem cells is therefore not only an indicator of their “stemness”, it can be used to either promote or suppress their differentiation.


Dr. Peter Toth, one of the co-authors on the paper, helped us acquire some really beautiful images of the cells that I would like to share with the readers of the blog. The image below shows undifferentiated adult human bone marrow mesenchymal stem cells (MSCs) that were exposed to an adipogenic differentiation medium, i.e a combination of factors which induces the formation of fat cells (adipocytes). However, as with many stem cell differentiation protocols, not all stem cells turned into fat cells. The cells on the right have a typical fat-like structure in which cells are full of round lipid droplets. The neighboring cells on the left are MSCs that have not (yet?) become fat cells. We stained the cells with the fluorescent mitochondrial dye JC-1. Depolarized mitochondria appear green and hyperpolarized mitochondria red. As you can see, the cells on the left have a much higher mitochondrial membrane potential (significant amount of red among the green mitochondria) than their fat neighbors on the right (mostly green mitochondria, all of them located between lipid droplets). By capturing both cell types next to each other, we could show an illustrative example of how entwined metabolism and stem cell differentiation are. The morphology and metabolic state of neighboring cells in this image were quite different, despite the fact that all cells were subjected to the same cocktail of differentiation factors. The blue-appearing dye is DAPI and stains nuclei of cells so one can tell the cells apart. Each cell in this image has one blue nucleus.



The image was published with a PLoS ONE CC-BY license. Feel free to use it as an example of adult stem cell differentiation or how mitochondrial morphology and function can vary between stem cell and its differentiated progeny, as long as you attribute the original PLoS One paper. The image in the paper also has a scale bar and asterisks/arrows pointing out the specific cells.

Zhang Y, Marsboom G, Toth PT, & Rehman J (2013). Mitochondrial respiration regulates adipogenic differentiation of human mesenchymal stem cells. PLoS ONE 8(10): e77077;  PMID: 24204740; DOI: 10.1371/journal.pone.0077077

Stemming the Flow: Using Stem Cells To Treat Urinary Bladder Dysfunction

Neurogenic bladder is a disorder which occurs in spinal cord diseases such as spina bifida and is characterized by an inability of the nervous system to properly control the urinary bladder and the muscle tissue contained in the bladder wall. This can lead to spasms and a build-up of pressure in the bladder, often resulting in urinary incontinence. Children with spina bifida and neurogenic bladder often feel urges to urinate after drinking comparatively small amounts of liquid and they can also involuntarily leak urine. This is a source of a lot of emotional stress, especially in social settings such as when they are around friends or in school. If untreated, the long-standing and frequent pressure build-up in the bladder can have even more devastating effects such as infections or kidney damage.

Current treatments for neurogenic bladder involve surgeries which reconstruct and increase the size of the bladder by using tissue patches obtained from the bowel of the patient. Since such a gastrointestinal patch is derived from the patient’s own body, it is less likely to elicit an immune response and these intestinal tissue patches tend to be strong enough to withstand the pressures in the bladder. Unfortunately, the incompatibility of intestinal tissue and bladder tissue can lead to long-term complications, such as urinary tract infections, formation of urinary tract stones and in some rare cases even cancers. For this reason, researchers have been searching for newer safer patches which resemble the actual bladder wall.


A team of researchers at Northwestern University recently published a study which used stem cells of children with spina bifida to generate tissue patches that could be used for bladder surgery. In the paper “Cotransplantation with specific populations of spina bifida bone marrow stem/progenitor cells enhances urinary bladder regeneration” published in the Proceedings of the National Academy of Sciences (online publication on February 19, 2013), Arun Sharma and colleagues isolated two types of cells from the bone marrow of children with spina bifida: Mesenchymal stem cells (MSCs) and CD34+ cells (stem and progenitor cells which usually give rise to blood cells). They then coated a special polymer scaffold called POC with the cells and implanted this newly created patch into a rat bladder after performing a bladder augmentation surgery, similar to what is performed in patients with spina bifida. They then assessed the survival and formation of human muscle tissue on the implanted patch. When both human cell types (MSCs and CD34+) were combined, more than half of the implanted patch was covered with muscle tissue, four weeks after the implantation. If they only used CD34+ cells, they found that only a quarter of the patch was covered with muscle tissue. What is even more remarkable is that in addition to the newly formed muscle tissue, the implanted patch also showed evidence of some peripheral nerve growth and of blood vessel formation, both of which are found in healthy, normal bladder walls. These findings suggest that a patient’s own bone marrow stem cells can be used to help construct a tissue patch which could be used for bladder augmentation surgeries. The observation of some nerve growth in the implanted patch is also an exciting finding. One could conceivably try to re-connect the reconstructed bladder tissue with the main nervous system, but its success would largely depend on the severity of the neurologic disease.

One has to keep in mind that there are some key limitations to this study. The authors of the paper believe that the newly formed muscle tissue on the implanted patches was all derived from the patients’ bone marrow stem cells. However, there were no experiments performed to convincingly demonstrate this. The authors report that in previous studies, merely implanting the empty POC scaffold without any human stem cells resulted in 20% coverage with muscle tissue. This suggests that a big chunk of the newly formed muscle tissue is actually derived from the host rat and not from human stem cells. The authors also did not compare the effectiveness of this newly formed stem cell patch to the currently used intestinal patches, and there is no assessment of whether the newly formed muscle tissue on the reconstructed bladder is less prone to spasms and involuntary contractions. Lastly, all the in vivo testing of the tissue patches was performed in rats without neurogenic bladder and it is possible that the highly successful formation of muscle tissue may have been diminished if the animals had a neurologic disease.

A second study published in PLOS One took a different approach. In “Evaluation of Silk Biomaterials in Combination with Extracellular Matrix Coatings for Bladder Tissue Engineering with Primary and Pluripotent Cells” (online publication February 7, 2013), Debra Franck and colleagues describe how they coated a scaffold consisting of silk threads with extracellular matrix proteins such as fibronectin. Instead of using bone marrow stem cells, they converted induced pluripotent stem cells into the smooth muscle cells that are typically found inside the bladder wall and placed these newly differentiated cells on the silk scaffold. The induced pluripotent stem cells (iPSCs) used by Franck and colleagues can be generated from a patient’s own skin cells which reduces the risk of being rejected by a patient’s immune system. The advantage of this approach is that it starts out with a pure and truly pluripotent stem cell population, which is easier to direct and control than bone marrow stem cells. There are also a few important limitations to this second study. Franck and colleagues used mouse pluripotent stem cells and it is not clear that their approach would necessarily work with human pluripotent stem cells. They also did not test the function of these differentiated cells on the silk scaffold to check if they actually behaved like true bladder wall smooth muscle cells. Unlike the first study, Franck and colleagues did not evaluate the newly created patch in an animal model.

Both studies are purely experimental and much additional work is needed before they can be tested in humans, but both show promising new approaches to help improve bladder dysfunction. It is heartening to see that researchers are developing new cell-based therapies to help children and adults who suffer from neurogenic bladder. The results from these two experimental studies are still too preliminary to predict whether cell-based therapies can be successfully used in patients, but they represent important first steps.


Image credit: Taken from Franck D, Gil ES, Adam RM, Kaplan DL, Chung YG, et al. (2013) Evaluation of Silk Biomaterials in Combination with Extracellular Matrix Coatings for Bladder Tissue Engineering with Primary and Pluripotent Cells. PLoS ONE 8(2): e56237. doi:10.1371/journal.pone.0056237- Figure 6 B: Differentiated mouse induced pluripotent stem cells cultured on fibronectin-coated silk matrices show protein markers typically found in bladder smooth muscle cells.
Franck, D., Gil, E., Adam, R., Kaplan, D., Chung, Y., Estrada, C., & Mauney, J. (2013). Evaluation of Silk Biomaterials in Combination with Extracellular Matrix Coatings for Bladder Tissue Engineering with Primary and Pluripotent Cells PLoS ONE, 8 (2) DOI: 10.1371/journal.pone.0056237