A German research team at the University of Duisburg-Essen has published a study in the latest issue of STEM CELLS Translational Medicine that shows tiny membrane-enclosed structures that travel between cells work as well as adult stem cells to help the brain recover from a stroke.
Extracellular vesicles (EVs), which are small, membrane-enclosed structures that pass between cells, which are also referred to as exosomes, were given to one group of stroke-impaired mice and adult stem cells from bone marrow to another. After monitoring these mice for four weeks, both groups experienced the same degree of neurological repair. Besides promoting brain recovery in the mice, the EVs also down-regulated the post-stroke immune responses and provided long-term neurological protection.
This study could lead to a new clinical treatment for ischemic strokes, since exosomes carry far fewer risks than adult stem cell transplants, according to the co-leaders of this research, neurologist Thorsten Doeppner, and Bernd Giebel, a transfusion medicine specialist.
“We predict that with stringent proof-of-concept strategies, it might be possible to translate this therapy from rodents to humans, since EVs are better suited to clinical use than stem cell transplants,” said Doeppner and Giebel.
Scientists think that EVs carry biological signals between cells and direct a wide range of processes. Exosomes are under a good deal of scientific investigation for the role they could play in cancer, infectious diseases, and neurological disorders.
Other studies have shown that exosome administration can be beneficial after a stroke, but the Duisburg-Essen study is the first to supply evidence through a side-by-side analysis that they act as a key agent in repairing the brain.
“The fact that intravenous EV delivery alone was enough to protect the post-stroke brain and help it recover highlights the clinical potential of EVs in future stroke treatment,” Doeppner and Giebel said.
This study included contributions from ten different researchers from Duisburg-Essen’s Department of Neurology and Institute for Transfusion Medicine. The study was supported by the university, Volkswagen Foundation and German Research Council.
“The current research, combined with the previous demonstration that EVs are well tolerated in men, suggests the potential for using this treatment in conjunction with clot-busting therapies for treatment of stroke,” said Anthony Atala, M.D., editor of STEM CELLS Translational Medicine and Director of the Wake Forest Institute for Regenerative Medicine.
If something lodges in the blood vessels that feed the brain – say a blood clot, piece of bone marrow after a bone has been broken, or tissue debris from damaged tissue – the brain undergoes a loss of blood flow. Since the brain received its oxygen and nutrients from the bloodstream, blockage of the vessels that feed the brain can lead to the death of brain cells.
Such a phenomenon is called a stroke or a Trans-Ischemic Attack. However, if the heart stops, blood flow to the brain ceases; not because of blockage of the blood vessels that feed the brain, but because the pump that propels through the bloodstream has stopped and, therefore, blood flow stops. Such a condition is known as global cerebral ischemia or GCI.
GCI is one of the most challenging clinical issues encountered during cardiac arrest and, unfortunately, typically indicates a poor prognosis. Severe neurological damage develops in 33%–50% of GCI patients who have survived a cardiac arrest that was documented by a medical professional. In those rare cases of survival after cardiac arrest that was not documented by a medical professional, the percentage of neurological defects is 100%. I hope this convinces you that CGI is a problem.
In order to treat GCI, physicians usually induce hypothermia, which lowers and maintains the core body temperature at 32°C–34°C. Presently, this is the only treatment regime that has been demonstrated to improve neurological recovery. Unfortunately, there are many technical difficulties in the application of this therapy. Special equipment is required, and complications such as blood clots and infection are perennial problems. Is there a better way to treat GCI?
Sang Won Suh from the Hallym University College of Medicine in South Korea and his colleagues have used fat-based mesenchymal stem cells to treat laboratory animals that have suffered GCI. The results of their study are encouraging.
Suh and his coworkers used Sprague-Dawley rats for this study. They anesthetized the rats and then clamped their carotid arteries to reduce blood flow to the brain for seven minutes. This effectively simulates GCI in these laboratory animals. After the clamps were removed, some animals were given one million fat-based mesenchymal stem cells, and others were simply restored by means of unclamping the carotid arteries plus fluid reconstitution. The rats were subjected to behavioral tests three days before the procedure and seven days after it. These tests consisted of placing adhesive tape the forepaws of the animals and then measuring the day it tool for the animals to remove to adhesive tape. After the seventh day post-procedure, the rats were put down and their brains were examined for cell death, structure, blood vessel densities, and degree of inflammation.
When the brains of these animals were examined, it was clear that the animals that had received fat-based mesenchymal stem cells suffered much less cell death than the untreated animals.
In the figure above you can see a Fluoro-Jade B staining of these brains. FJB stains detect dying cells. As you can see, the brain from the rats that experienced GCI without any stem cell treatments had lots of dying cells in their brains. The “sham” operated rats – rats that were operated, but their carotid arteries were not clamped – had no cell death in their brains. The animals that had their carotid arteries clamped, but were given fat-based mesenchymal stem cells had a little cell death. The graph above shows the vast differences between the stem cell-treated and the non-stem cell-treated groups. Truly these are significant results. Other experiments that detected Now this is no a surprise, since Ohtaki and others showed a very similar result in 2008 (OhtakiH, et al. Proc Natl Acad Sci USA 105:14638–14643). Suh, and his group, however, took these experiments further to determine why these cells prevented cell death in the brain after GCI.
When Suh and his team examined the leakage of large proteins into the brain, they saw something quite remarkable; the mesenchymal stem cell-treated rats only leaked a little protein into their brains compared to the non-stem cell-treated rats.
The presence of the brown color indicated the presence of a protein in the brain that normally does not find its way to the brain unless the integrity of the blood-brain barrier is compromised. As you can see, the non-treated animals have a truckload of this protein in their brains, which indicates that their blood-brain barriers are very leaky. On the contrary, the stem cell-treated brains are not nearly as leaky and the sham operated brains are not leaky at all.
These results suggest that the stem cells help maintain the structural integrity of the blood-brain barrier in GCI patients and this prevents nasty things from the bloodstream, such as immune cells and so on from accessing the brain and ravaging it. To test this hypothesis, Suh and others examined the brains for the presence of neutrophils, which are white blood cells that show up when inflammation occurs. These cells are not found in the brain unless the blood-brain barrier is damaged. Sure enough, brains from the sham-operated rats showed no signs of neutrophils, brains from the non-stem cell-treated rats were chock full of neutrophils, and the brains from the stem cell-treated rats only had a few neutrophils.
A conclusion from this paper states: “Administration of MSCs decreased the delayed neuronal damage in a transient global cerebral ischemia model by prevention of BBB disruption, endothelial damage, and neutrophil infiltration.”
Clearly this merits more work. Larger animal models will need to be examined, and also it would be nice to know if administration of exosomes from mesenchymal stem cells can elicit a similar biological response. However his is a very hopeful beginning to what might become a fruitful bit of clinical research.
When blood flow to the brain ceases as the result of a blood clot, trauma, or injury, the brain suffers from a shortage of oxygen. Such an incident is known as a stroke and it can result in the death of neurons and the loss of those functions to which the dead neurons contributed. Treatment for stroke is largely supportive, but regenerative treatments that replace the dead neurons would be the most ideal treatment.
A research consortium at Lund University in Lund, Sweden has found that neurons made from induced pluripotent stem cells integrate into the brains of mice that had suffered strokes. This experiment takes a closer step towards the development of a regenerative treatment for strokes.
In the aftermath of a stroke, nerve cells in the brain die. At the Lund Stem Cell Center, the research groups of Zaal Kokaia and Olle Lindvall teamed up to develop a stem cell-based method to treat stroke patients.
After a stroke, the cerebal cortex tends to take the bulk of the damage and neuron loss from the cerebral cortex underlies many of the symptoms following a stroke, such a paralysis and speech problems. The method developed by the Lund Institute scientists should make it possible to generate nerve cells for transplantation from the patient’s own skin cells.
First, the Lund team isolated skin fibroblasts from the afflicted mice and used genetic engineering techniques to convert them into induced pluripotent stem cells (iPSCs), which have many of the differentiation capabilities of embryonic stem cells. These iPSC lines were differentiated into cortical neurons, which tend to populate the cerebral cortex. However, transplanting fully differentiated neurons into the brain tend to not work terribly well because the mature neurons are unable to divide and have poor abilities to connect with other cells. Therefore, the neuron progenitor cells that will give rise to cortical neurons are a better candidate for transplantation.
After generating long-term self-renewing neuroepithelial-like stem cells from iPSCs in the laboratory, the Lund group scientists showed that these stem cells could give rise to neural progenitors that expressed the types of genes found in mature cortical neurons. When these neural progenitor cells were transplanted into rats that had suffered strokes, two months after transplantation, the cortically fated cells showed less proliferation and more efficient differentiation into mature neurons with the right shape, size, and structure of cortical neurons and expressed the same proteins as cortical neurons. These tranplanted cells also extended more axons than those cells that were not fated to form cortical neurons. Transplantation of both the cortical neuron-fated and non-cortical neuron-fated cells caused recovery of the impaired function in the stepping test in comparison to controls. At 5 months after stroke, there was no tumor formation and the grafted cells had all the electrophysiological properties of mature neurons and showed full evidence that they had integrated into the existing neural circuitry.
These results are very promising and represent a very early but important step towards a stem cell-based treatment for stroke in patients. Further experimental studies are necessary if these experiments are to be translated into the clinic in a responsible way.