Transplantation of Neurons Made from Bioreactor-Grown Human Neural Precursor Cells Restores Brains of Rats With Huntington’s Disease


Huntington’s disease (HD) is an inherited disorder of the central nervous system characterized by progressive dementia, involuntary movements, and emotional deterioration.  The brain is affected in patients with HD and the part of the brain that takes the biggest beating is the “neostriatum.”

The term “neostriatum” is almost certainly not a word that you hear terribly often in conversation.  Therefore I will try to explain what it is.  The outer layers of the brain are known as the cerebral cortex and they are composed of so-called “grey matter.”  The cerebral cortex consists of grey matter because it is loaded with cells known as neurons.  Beneath the cortical layer of the brain, lies a whole host of extensions of these neurons that reside in the cerebral cortex.  These extensions are called “axons,” and beneath the cerebral cortex lies white matter, which consists, largely, of bundles of axons.  Think of the neurons are plugs and the axons as extension cords.  The neurons are plugged into each other by means of extension cords that extend from the cerebral cortex.

Now the cerebral cortex is not the only game in town.  There are also clusters of neurons that lie beneath the cerebral cortex called “nuclei.”  One of these nuclei beneath the cerebral cortex plays an extremely important role in voluntary motion, and this structure is called the “basal ganglia.”  Here’s a picture to make things a little clearer:

Basal ganglia

As you can see in the figure, the striatum consists of two structures: the putamen and the caudate nucleus.  The striatum or striate nucleus receives neural inputs from the cerebral cortex and inputs this neural information to the basal ganglia.

From a functional perspective, the striatum helps coordinate motivation with body movement.  It facilitates and balances motivation with both higher-level and lower-level functions – for example, inhibiting one’s behavior in a complex social interaction and fine-motor functions involved in inhibiting small voluntary movement.

Some of the neural outputs from the striatum are excitatory – they stimulate other neurons.  Other signals are inhibitory – they prevent the neurons to which they are connected from becoming stimulated.  Inhibitory neurons release a chemical called “GABA.”  These GABA-using neurons are very important for the work of the striatum, and it is exactly these neurons that die off at the greatest rate in patients with HD.  Therefore, treatments for patients with HD have focused on replacing or protecting these GABA-using neurons.

Experimentally, you can induce an HD-like disease in rodents if you inject a chemical into their brains called quinolinic acid.  Quinolinic acid causes many of the GABA-using neurons in the striatum to pack up and die, and for this reason, this chemical is heavily used in the laboratories of scientists who study HD and HD treatments.

In a paper by Marcus McLeod and others who did their work in the laboratory of Ivar Mendez, who was at Dalhousie University in Nova Scotia, Canada, but has since moved to the University of Saskatchewan, GABA-using neurons were made from cultured human neural precursor cells (hNPCs) and then implanted into the brains of rats that had been injected with quinolinic acid.  The results were spectacularly successful.  This work was published in the journal Cell Transplantation.

A definite twist with this particular paper is the way the GABA-using neurons were grown in culture; they were grown in bioreactors.  Bioreactors are devices that support biological cells, processes, or organisms.  They keep the environment of the cells constant, and provide a far superior way to grow cells or tissues in the laboratory.  McLeod and his colleagues used human neural progenitor cells and grew them to large numbers in bioreactors.  These expanded hNPCs were then differentiated them into GABA-using neurons and then injected into the brains of rats who has been treated with quinolinic acid.

The rat model allows the scientist to inject only one side of the brain with quinolinic acid.  This leaves the intact side of the brain as a control tissue that can be compared with the injected one.  The injected rats showed the characteristic death of the GABA-using neurons and the behavioral features that result from the death of these neurons.  Such animals do not walk normally when they are led through a cylinder, and they have trouble finding their way through a maze.  The animals that received the transplantations of the GABA-using neurons, however, performed almost as well in these tests as normal rats; not quite as well, but almost as well.  The rats treated with quinolinic acid did quite poorly, as expected.

Upon post-mortem examination, the rats transplanted with GABA-using neurons shows a host of new GABA-using neurons in their striatums.  These cells also underwent further maturation after transplantation, and they also made connections with other neurons.

Now this paper shows that the injected cells not only survived the transplantations, but they also matured, made connections and promoted recovery of many of the behavioral symptoms of HD.  This procedure certainly has promise.

Having said all that, there are two caveats to these experiments.  The rodent model is a good model as far as it goes, but it seems clear that the actual human disease turns the environment of the brain into a very inhospitable place.  Transplanted cells in the case of human HD patients do not usually survive terribly well.  It seems to me that treatments like this must be coupled with other treatments that seek to improve the actual cerebral environment.  The second caveat to this experiment is that the neural progenitor cells were taken 10-week-old from aborted fetuses.  While these scientists did not perform the abortions that ended the lives of these babies, it is more than little troubling that this research was done using the corpses of those babies whose lives were prematurely ended.

Nevertheless, despite these caveats, this paper represents a definite advance in the regenerative strategies available to treat HD patients.

Personalized Stem Cells for Curing Parkinson’s Disease


Stem cell treatments for curing Parkinson’s disease have been one of the dreams of stem cell scientists ever since the first embryonic stem cells were derived from mouse embryos in 1981. Unfortunately, this proved to be one of the harder therapeutic nuts to crack. Several experiments have shown that while feasible, getting the recipe right has required a fair amount of tweaking.

brain-labels

Parkinson’s disease (PD) results from the progressive death of neurons in the midbrain that release a neurotransmitter called dopamine, To review briefly, the brain consists of the forebrain, midbrain and hindbrain. The forebrain consists of the two large cerebral hemispheres that occupy the vast majority of the space within your skull. In addition to the left and right cerebral hemispheres is the diencephalon that consists of the thalamus, subthalamus, hypothalamus, and epithalamus. The thalamus serves as a relay station for a whole variety of nerve fiber tracts, the hypothalamus regulates visceral activities by way of other brain regions and the autonomic nervous system. and the epithalamus connects the limbic system to the rest of the brain. The midbrain, which lies below the diencephalon, is part of the brain stem and dopamine produced in two regions of the midbrain, the substantia nigra and ventral tegmental area play roles in motivation and habituation, and refinement of the control of voluntary movement, The hindbrain consists of the metencephalon and the myelencephalon, both of which contain mutiple fiber tracts and nuclei for vital functions.

Midbrain 2

The death of dopamine-producing neurons in the pars compacta region of the substantia nigra region of the midbrain causes PD. The par compacta sends nerve fibers to the cerebral hemispheres, in particular to cluster of neurons called the basal ganglia. The basal ganglia do not initiate movement, but they refine movement and stabilize the limbs and other body parts while moving. Thus the basal ganglia normally exert a constant inhibitory influence on a wide range of movements. preventing movement at inappropriate times. When someone decides to move, this inhibition is reduced for the required motor system, thereby releasing it for activation. Dopamine releases this inhibition, and therefore high levels of dopamine tend to promote movement and low levels of dopamine demand greater exertion to generate any given movement. Thus the net effect of dopamine depletion is to produce hypokinesia, or less overall movement.

Basal ganglia

Now that we have some knowledge of PD and what causes it, we can examine how to cure it. Since the death of dopamine-secreting neurons causes PD, replacing death or moribund neurons should be possible. Several preclinical studies in laboratory animals and clinical studies with human patients has shown that this is possible.

Rodents can contract a synthetic form of PD if they are treated with a drug called 6-hydroxydopamine. This drug kills off their dopamine-secreting neurons and creates a PD-like disease. Embryonic stem cells can be differentiated in the laboratory into dopamine-secreting neurons, which can then be transplanted into the midbrain. In PD rats, this strategy has reversed the symptoms of PD, but tumor growth has been a nagging problem. The biggest problem is that isolating fully differentiated dopamine-secreting cells has proven difficult because of a lack of good, solid indicators that say to the scientists, “This one is a dopamine-secreting neuron and this one is not.” Thus, isolating nice, clean cultures of only dopamine-secreting cells has been kind of tough to do.

Fortunately, Doi and others in the Takahashi lab at the University of Kyoto showed that prolonged maturation culture system (42 days long) can eliminate most of the tumor-making cells. However, this culture system is laboriously long. Now, Takahashi and Doi and others have struck again in a paper published in Stem Cell Reports in which they used induced pluripotent stem cells to derive dopamine-secreting neurons to treat PD rats.  Because induced pluripotent stem cells are made from a patient’s own adult cells and are converted into embryonic-like stem cells by means of a combination of genetic engineering and cell culture techniques, they are patient-specific and do not require the dismembering of human embryos.

The novelty of this paper is that Doi and others used a protein that acts as an earmark for dopamine-secreting midbrain neurons and this protein is called CORIN. CORIN is a protease, which simply means that it clips other proteins into small pieces. Nevertheless, by using the CORIN protein, Takahashi, Doi and others successfully and efficiently isolated dopamine-secreting midbrain neurons from other cells in their cultures.  Additionally, Doi and the gang were able to differentiate the induced pluripotent stem cells into dopamine-secreting progenitor cells.  This means that the cells were poised to differentiate into dopamine-secreting neurons, but were not quite there yet.  This way, the cells would grow in culture, but upon transplantation, they would differentiate into dopamine-secreting neurons rather than form tumors.  High numbers of cells are required for clinical purposes and this technique allows the for production of large number of cells.

The technique used in this paper also produced the cells under conditions that were safe, scalable and potentially usable for clinical use. These high-quality cells never produced any tumors and produced definitive behavioral improvements in the implanted laboratory animals. The problems that remain are one of scale. The grafts of dopamine-secreting cells that survived in the midbrains of these mice were relatively small (about 1 square millimeter in size or the thickness of a dime).  This is probably due to the fact that the cells differentiate when transplanted rather than growing.  Therefore, this technique will need to be adapted to somehow increase the size of the graphs of dopamine-secreting neurons.  In some PD patients such small graphs will probably work just fine, but in others, probably not.  The other issue is that these implanted cells might be subjected to the same bad intracerebral environment as the original cells and die off quickly, thus abrogating any positive clinical effect they might have.  This is another issue that will need to be examined.

The work goes on, without the need to destroy any embryos.

See Daisuke Doi at al., Isolation of Human Induced Pluripotent Stem Cell-Derived Dopaminergic Progenitors by Cell Sorting for Successful Transplantation. Stem Cell Reports 2014, 2: 337-350.