Using Stem Cells to Model the Blood-Brain Barrier

Our central nervous system include the brain and spinal cord. The central nervous system (CNS) is surrounded by a series of tough coverings called meninges that protect it and is bathed and fed by a circulating fluid called cerebrospinal fluid (CSF). The blood vessels that circulate blood through the CNS are composed of specialized cells that are very tightly apposed. These specialized blood vessels prevent molecules from spreading from the body to the CNS. In order for something to enter the CNS, the blood vessel-making cells (endothelia) must possess specific receptors that bind the desired molecule and allow it to pass into the CNS. Over 100 years ago, scientists found that if dye was injected into the bloodstream of a laboratory animal, the dye would enter everywhere except the CNS. This shows that there is a barrier that prevents the passage of all but a select set of molecules into the CNS and this barrier is called the blood-brain barrier (BBB).

The BBB is selectively permeable, which means that it allows some materials to cross into the CNS, but prevents others from doing so. In most parts of the body, the smallest blood vessels, called capillaries, are lined with endothelial cells. Endothelial tissue has small spaces between each individual cell so substances can move readily between the inside and the outside of the vessel. However, in the brain, the endothelial cells fit tightly together and substances cannot pass out of the bloodstream into the CNS. Some molecules, such as glucose, are transported out of the blood by special mechanisms.

Generally speaking. the BBB does not allow passage of large molecules, and non-fat soluble molecules also do not enter the brain. However, fat soluble molecules, such as barbituate drugs, can rapidly enter the brain. Also molecules that have a high electrical charge, if they enter the CNS at all, only do so rather slowly.

The BBB also prevents cancer drugs from entering the CNS, and this is one of the main reasons cancers of the CNS are difficult to treat. Designing cancer drugs that can enter the CNS and bypass the BBB is also challenging.

Fortunately, a new study from the University of Wisconsin, Madison, has shown that embryonic stem cells can be coaxed into differentiating into structures that greatly resemble the BBB. This might provide drug companies with a new model system to study the movement of experimental drugs into the CNS.

Eric Shusta is professor of chemical and biological engineering, is one of he senior authors of this new study. Since his laboratory has succeeded in differentiating embryonic stem cells into endothelial cells with BBB characteristics, he thinks that this “has the potential to streamline drug discovery for neurological disease. You can look at tens of thousands of drug candidates and just ask the question if they have a chance to get into the brain. There is a broad interest from the pharmaceutical industry.”

The endothelial cells generated in Shusta’s lab exhibit the active and passive regulatory characteristics of endothelial cells from the brains of a living animal.

Shusta and his team were able to induce embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) into forming BBB-like structures. The ability to drive iPSCs to form BBB-like structures is significant, since scientists could use cells from patients with particular neurological diseases to make BBB-like structures and then tailor drug treatments that will take into account the capacity of the patient’s BBB to admit or exclude particular drugs.

From an industrial standpoint, because these cells can be grown in culture and mass-produced, the can be used for diverse high-throughput screens for molecules that may have therapeutic value for neurological conditions or to identify neurotoxic properties of existing drugs.

According to Shusta: “The nice thing about deriving endothelial cells from induced pluripotent stem cells is that you can make disease-specific models of brain disease that incorporate the BBB. The cells you create will carry the genetic information of the condition you want to study.”

The BBB is also complete at birth but fragile. Former experiments led scientist to believe that the BBB was immature at birth, but these experiments used conditions that were destructive for the BBB. Therefore, high levels of particular molecules that are not a problem for an adult, such as bilirubin, can cause profound problems in a baby. Kernicterus is a form of mental retardation that results from high blood bilirubin levels in a newborn. The bilirubin accumulates in the brain and causes brain damage. This is a phenomenon that results from the BBB in neonates being overloaded with bilirubin. Certain medical conditions increase the risk of kernicterus. For example, premature birth, Rh incompatibility, polycythemia (too many red blood cells), certain drugs such as sulphonamides, which displace bilirubin from serum albumin, Crigler-Najjar syndrome, Gilbert’s syndrome or G6PD deficiency all predispose babies to kernicterus. A model system in which BBB cells from patients with these diseases are cultured and grown in the lab to provide profound and potentially life-saving insights into these diseases and how they affect the BBBs of newborn babies.

Making the BBB in culture also led to another important insight, The formation of the BBB requires the activity of brain-specific cells such as neurons. Shusta explained that neurons develop at the same time as the endothelial cells. Therefore, the developing neurons seem to secrete chemical cues that help determine functional specificity to the growing endothelial cells. Presently, Shusta and his group do not know what those chemicals are, but with this in the dish model, it will be relatively easy to go back and look.

Finally, in quoting from the abstract of this paper, “The resulting endothelial cells have many BBB attributes, including well-organized tight junctions, appropriate expression of nutrient transporters and polarized efflux transporter activity. Notably, they respond to astrocytes, acquiring substantial barrier properties as measured by transendothelial electrical resistance (1,450 ± 140 Ω cm2), and they possess molecular permeability that correlates well with in vivo rodent blood-brain transfer coefficients.” In other words their model BBB looks like a BBB and functionally acts like one. The possibilities for this model system are truly tremendous.