The blood-brain barrier (BBB)

Endothelial cells, which are highly specialized to enable precise regulation of chemicals entering or leaving the brain, make up the blood-brain barrier (BBB). Endothelial cell interaction with cellular and non-cellular components controls the barrier's development and maintenance (Obermeier et al., 2013). Extracellular matrix, pericytes, and astrocytes in particular offer both structural and functional support, and the neurovascular unit affects how the cellular components interact with one another. Saunders et al. (2016) claim that the essential purpose of the BBB is to preserve the stability of the CNS's external environment, particularly with regard to its ionic elements, or, to put it another way, to facilitate the transmission of nerve impulses. It further regulates entry of a wide range of nutrients such as amino acids, vitamins, monocarboxylates and glucose to ensure that they are an inappropriate concentration in the brain as they are essential in its development and functionality (Pardridge, 2005). The barrier also protects the brain from toxic substances including drugs such as antibiotics and antibodies, which pose a detrimental issue in the field of medicine as it limits neurotherapeutics (Saunders et al., 2016)). The idea of BBB emerged from studies conducted by Ehrlich, a Germany scientist who used to inject dye into the circulation of adult animals in 1878 (Ribatti et al., 2006). However, the existence of this barrier was first hypothesized by Bield and Kraus in 1998 and Lewandowsky in 1900 after finding out that sodium ferrocyanide had no pharmacological impact on the Central nervous system when intravenously injected while neurological symptoms appeared after intraventricular application of the same substance was done. The following discussion is a literature review on development of the BBB.

How Seminal Studies Inspired Further Research

Initial conclusions drawn by Paul Ehrlich were that brain remained unstained due to a characteristic feature of CNS that makes it have low affinity for the dye. That discovery made scientists more curious including Goldmann to understand how that was possible and which precisely was that feature (Obermeier et al., 2013). However, surprisingly the subsequent studies revealed convincingly that what existed was a barrier, which restricted movement of materials from blood to the CNS. In fact, as it will be shown in the review below immediately after Bield and Kraus in 1998 and Lewandowsky in 1900 clarification and affirmation of BBB existence many other scientific studies emerged seeking to understand what BBB precisely is and things that are made up of and how it develops.

Some of the researchers focused on understanding whether the BBB works and what kind of substance can pass through the barrier. A good example is Goldman who was Ehrlich student, the man who evidently showed that BBB is real, and not imagination. Others sought to understand how the barrier develops from early stages of life (Obermeier et al., 2013). Many of them focused on the understanding formation of BBB from an early age, which has up to date brought numerous insight about the composition of the barrier. In fact, since the coining of the BBB concept discoveries of previous researchers have always inspired more studies as people continue trying to understand more information needed to improve medicine and treatment approaches.

Literature Review

Following the formulation of brain barrier, the second significant experiment according to Davson (1989) is Goldmann’s trypan blue experiments of 1909 to 1913. As explained by Davson, the researcher intravenously injected the dye into the brains of some rabbits and dogs of the total sample size while the others were injected through the ventricular system. Observations made were quite insightful as for those with dye injected into ventricular brain system had a colored brain while intravenously injected subject turned blue in most parts of the body except spinal cord and brain that remained unstained (Davson, 1989). Interestingly the choroid plexuses also got stained. The experiments eliminated the idea posed by Enrich stating that the reason for the lack of stain in the brain after parental injection of adult brain with vital dyes was a particular feature of CNS that caused low affinity to this dyes and supported the unclear BBB concept coined by Lewandowsky (Davson, 1989). Goodman further theorized that the transporter of materials was the cerebrospinal fluid, which enters the brain through choroid plexuses. Currently, this model is also known as way of the spinal fluid.

As discussed by Saunders (2014), more studies using trypan blue were conducted by Wislocki in 1920, who observed that similar to adults animals embryo stained in most parts of the body except in the brain. Even earlier than Wislocki studies, Weed in 1917 using potassium ferrocyanide and pig embryos conducted a similar study. The scientist observed that even in the early development stages after neural tube closure the CNS separates from the rest of the developing organs since it stained after injecting isotonic sodium ferrocyanide into spinal canal while the rest of the embryo remained uncolored. This findings are further supported by Cohen and Davis in 1938 in their experiment using chick embryos. According to Saunders (2014), these tests provided sufficient and convincing evidence to show existence of brain blood barrier right from embryonic stage of development yet scientist have persisted for over a century with the belief that embryos, fetus and even babies have no or immature BBB

More studies on the same were conducted by Olsson et al. (1968) who tested cerebral blood vessels permeability to albumin as development takes place. They utilized bovine serum that was labeled using fluorescent and injected it into the tail of an adult, young and newborn rats as well as into the embryos’ umbilical artery that was aged between fifteen days and twenty days by (Olsson et al., 1968). Distribution of the tracer was tracked using fluorescence microscopy. In brain of postnatal and embryos brains, they observed that fluorescent albumin was confined to the blood vessels lumen, whereas substantial extravascular passage was seen in subcutaneous tissue. The findings indicated that in cerebral spinal vessels of rat embryos the walls are impermeable to albumin as early as the 15th day after fertilization.

More studies on the same conducted by Cremer, Braun & Oldendorf (1976) who further studies permeability of BBB to several compounds in rats between the age of 15 days and nine weeks. 14c- labeled test material was injected together with two reference isotopes. According to their observations, they saw that monocarboxylic transport system had a more significant capacity compared to adults with more permeability to acetate and l- lactate. In all ages, they observed a constant ratio between (−) d-3-hydroxybutyrate and l-lactate values (Cremer, Braun & Oldendorf, 1976). For D-Glucose, permeability was seen to increase with age whereas that of the amino acids was same in both adult and young rats. This observation affirmed the existence of BBB in both adult and young age but also reveals there is a slight difference between BBB as a subject develops.

Another significant study is by Risau, Hallmann & Albrecht (1986) who researched differentiation-dependent expression of proteins in endothelium during BBB development. BBB is a unique property of endothelium cells of the brain. To study this, the investigators correlated expression of several proteins in endothelial cells of the brain with the development of BBB in chick, quail and mouse embryos. They found that alkaline phosphatase activity was present in all the samples using histochemical methods. In particular they observed this in 12 days (chick), 14(quail) and 17 (mouse) days of embryonic development (Risau, Hallmann & Albrecht, 1986). In quails and Mouse butyrylcholinesterase activity was detected but was absent in brain vasculature of the chick in days 15 (quail) and 17(mouse). Histochemically they also found γ-Glutamyltranspeptidase activity in mouse only at the beginning of day 15 of embryonic development. Using monoclonal antibodies, they also found that transferrin receptor was localized on endothelium of brain in 11th and 15th day of chick and mouse embryonic development respectively.

They then compared leptomeningeal blood vessels and brain endothelium of adult to embryonic brain using staining of all markers. They further correlated development of BBB with expression of this proteins by determining permeability of brain endothelium to protein horseradish peroxidase during embryogenesis of mouse (Risau, Hallmann & Albrecht, 1986). Impermeability to the test substance was found to occur around day 16 of embryonic development. When all the above results were taken together they scientists concluded that quantity of proteins are sequentially expressed in endothelial cells of brain correlates with the time of BBB formation in different species (Risau, Hallmann & Albrecht, 1986). This findings are important as provides more insight on how the BBB develops and opened up ne opportunity for scientist to study and understand the stages of barrier development and proteins involved.

The other significant study on development of BBB was carried out by Kniesel, Risau & Wolburg, and (1996) who investigated the complex tight junctions located between endothelia cells of brain capillaries. The investigators studied modulation of tight junction fine structure in endothelial cells of BBB during the cerebral cortex development of the rat using freeze- fracture technique. The researchers used adult rat, postnatal one day old rat and embryos aged 13, 15 and 18 (Kniesel, Risau & Wolburg, 1996). They observed that complexity of tight junction and association of their particles significantly increased between day 18 of embryonic development and day one after birth. This finding suggest transition process of BBB to mature state which further affirms that newborn infants have BBB unlike the previous believe about lack of the barrier at young age.

All these studies have brought in sight on what makes up BBB namely the tight junctions and endothelial cells, which reveal biochemical composition and unique morphological constituents of the body’s vasculature. Recent studies as indicated by Liebner, Czupalla & Wolburg, (2011) identified Wnt/b – catenin pathway as essential during brain angiogenesis, BBB and tight junction formation. Gaining insight about mechanism formation and development of BBB is crucial as it assist in improving treatment approaches of diseases that attacks brain.

Conclusion

More studies about development of BBB have been conducted and lately scientists have discovered that retinoic acid plays an important role in brain during embryogenesis. In the study about the same Mizee et al. (2013) analyzed human postmortem fetal brain tissue and found out that radial glial cells express the enzyme mainly responsible for retinoic acid synthesis (retinaldehyde dehydrogenase) while brain vasculature express RA-receptor β which is the most important retinoic acid receptor in CNS. All this discoveries about BBB and function are recent discoveries, otherwise since identification of brain blood barrier the concept has remained unknown over many years. As described by Chow & GU, (2015) the restrictive nature of the barrier hinders delivery of most drugs to CNS that have ability to cure many neurological disorders. Most studies in modern world are focused on discovering strategies of delivering drugs into the brain as more information unveil about the structural composition, formation and functionality of the BBB.

Pardridge (2012) suggests that scientist need to reengineer drugs for BBB transport based on the insight of transport system within the barrier. With such approach, small molecule drugs can be made in a manner that they have capability of accessing carrier mediated transport system while large molecule drug should be created with a molecular delivery system that access receptor mediated transport within the blood brain barrier. In fact, most drugs delivery strategies that are being recently developed target oligonucleotides, enzymes, glycoproteins, regulatory proteins and peptides. With the current technology, it is now possible to study live in vivo tracking of brain blood barrier using different brain on a chip in vitro models and Single photon emission tomography (Banks, 2009). Such technologies provide opportunities to scientist understand better BBB and discover various disruptions of BBB in sick condition together with ways of crossing the hurdles that occur in gene and drug delivery across the barrier.

























References

Banks, W. A. (2009). Characteristics of compounds that cross the blood-brain barrier. BMC neurology, 9(1), S3

Chow, B. W., & Gu, C. (2015). The molecular constituents of the blood–brain barrier. Trends in neurosciences, 38(10), 598-608.

Cremer, J. E., Braun, L. D., & Oldendorf, W. H. (1976). Changes during development in transport processes of the blood-brain barrier. Biochimica et Biophysica Acta (BBA)-Biomembranes, 448(4), 633-637.

Davson, H. (1989). History of the blood-brain barrier concept. In Implications of the blood-brain barrier and its manipulation (pp. 27-52). Springer US.

Kniesel, U., Risau, W., & Wolburg, H. (1996). Development of blood-brain barrier tight junctions in the rat cortex. Developmental brain research, 96(1), 229-240.

Liebner, S., Czupalla, C. J., & Wolburg, H. (2011). Current concepts of blood-brain barrier development. International Journal of Developmental Biology, 55(4-5), 467-476.

Mizee, M. R., Wooldrik, D., Lakeman, K. A., van het Hof, B., Drexhage, J. A., Geerts, D., ... & de Vries, H. E. (2013). Retinoic acid induces blood–brain barrier development. Journal of Neuroscience, 33(4), 1660-1671.

Obermeier, B., Daneman, R., & Ransohoff, R. M. (2013). Development, maintenance and disruption of the blood-brain barrier. Nature medicine, 19(12), 1584-1596.

Olsson, Y., Klatzo, I., Sourander, P., & Steinwall, O. (1968). Blood-brain barrier to albumin in embryonic new born and adult rats. Acta neuropathologica, 10(2), 117-122.

Pardridge, W. M. (2005). The blood-brain barrier: bottleneck in brain drug development. NeuroRx, 2(1), 3-14.

Pardridge, W. M. (2012). Drug transport across the blood–brain barrier. Journal of Cerebral Blood Flow & Metabolism, 32(11), 1959-1972.

Ribatti, D., Nico, B., Crivellato, E., & Artico, M. (2006). Development of the blood‐brain barrier: A historical point of view. The Anatomical Record, 289(1), 3-

Risau, W., Hallmann, R., & Albrecht, U. (1986). Differentiation-dependent expression of proteins in brain endothelium during development of the blood-brain barrier. Developmental biology, 117(2), 537-545.

Saunders, N. R., Dreifuss, J. J., Dziegielewska, K. M., Johansson, P. A., Habgood, M. D., Møllgård, K., & Bauer, H. C. (2014). The rights and wrongs of blood-brain barrier permeability studies: a walk through 100 years of history. Frontiers in neuroscience, 8.

Saunders, N. R., Habgood, M. D., Møllgård, K., & Dziegielewska, K. M. (2016). The biological significance of brain barrier mechanisms: help or hindrance in drug delivery to the central nervous system?. F1000Research, 5.