Research - Institute of Biophysics - Molecular Neurobiology Research Unit - Physiology and Pathology of the Blood-Brain Barrier Research Group

István KRIZBAI
scientific adviser

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Elek Attila FARKAS research associate
Imola WILHELM senior research associate
Csilla FAZAKAS research associate
János HASKÓ junior research associate
Ádám NYÚL-TÓTH junior research associate
Mihály KOZMA Ph.D. student
Ádám MÉSZÁROS Ph.D. student
Kinga MOLNÁR Ph.D. student

PHYSIOLOGY AND PATHOLOGY OF THE NEUROVASCULAR UNIT

The neurovascular unit (NVU) plays a key role in the maintenance of the homeostasis of the central nervous system (CNS). One of its main functions is the formation of the blood-brain barrier (BBB). The most important cellular components of the BBB are cerebral endothelial cells (interconnected by a continuous line of tight and adherens junctions), pericytes and astrocytes.

Due to its complex functions the BBB plays an important role in clinical practice. First, there is increasing evidence that the BBB is involved in the pathomechanism of a large number of CNS diseases like stroke, brain trauma and tumors as well as neurodegenerative disorders. Second, due to the relative impermeability of the barrier, many drugs are unable to reach the CNS in therapeutically relevant concentration, making the BBB one of the major impediments in the treatment of CNS disorders.

Our research is focused on the elucidation of molecular mechanisms regulating NVU function under physiological and pathological conditions. Recently we have shown that in different CNS disorders like cerebral ischemia or inflammatory disorders complex signaling events are initiated accompanied by changes in the integrity of the junctional complex. By using different in vitro models of the BBB and in vivo systems, presently we are investigating the role of the NVU in inflammatory processes and transmigration of tumor cells through the BBB.


Model system

In order to be able to directly investigate molecular mechanisms regulating BBB function, we use an in vitro model based on the culture of cerebral endothelial cells. The model can be used for basic research and applied research for the study of the interaction of drugs with the BBB and the transport of different drugs through the BBB. Our methods of investigation include Western-blot, phosphorylation analysis, proteomics, real-time PCR, RNA silencing, zymography, immunofluorescence, etc.



Figure 1: A: Cellular structure of the neurovascular unit. B: In vitro model of the BBB.



Role of the NVU in inflammatory processes

Inflammatory processes are associated to a large number of physiological and pathological conditions of the CNS including stroke, brain trauma and tumors, neurodegenerative disorders as well as ageing. We have characterized the expression profile of toll-like receptors in the cerebral endothelium and we identified the signaling pathway by which activation of TLR2/6 could lead to an increase in the permeability of the BBB. Furthermore, we have demonstrated that cerebral endothelial cells express a whole set of inflammasome components whose expression is increased in response to inflammatory mediators. Moreover cerebral endothelial cells are able to assemble functional inflammasomes which leads to interleukin-1-beta production. This may confer a yet unexplored role to the BBB in neuroimmune and neuroinflammatory processes.



Figure 2: Mecanisms of inflammasome activation in cerebral endothelial cells.



Interaction of metastatic tumour cells with the BBB

Brain metastases are clinically diagnosed in the majority of patients having secondary localizations of melanoma. The prognosis for these patients is very poor and treatment can be difficult because of rapid progression of the disease. Since the brain does not have a classical lymphatic system, tumor cells have to transmigrate through the BBB in order to reach the brain parenchyma. During this process the tumor cells either must disrupt intercellular junctions or use the transcellular route. We are currently investigating the molecular mechanisms of these phenomena. For this purpose besides in vitro techniques in vivo imaging using two-photon microscopy is applied.



Figure 3: A: Adhesion of melanoma cells to the cerebral endothelium. B: A2058 melanoma cells (green) in contact with brain endothelial cells (red: ZO-1 staining). C,D: in vivo two-photon microscopy images of tumor cells


Selected publications

Ramirez S, Hasko J, Skuba A, Fan S, McCormick R, Dykstra H, Reichenbach N, Krizbai I, Mahadevan A, Zhang M, Son YJ, Tuma R, Persidsky Y (2012). Activation of cannabioid receptor 2 attenuates leukocyte-endothelial interactions and blood-brain barrier dysfunction under inflammatory conditions. J. Neurosci. 32:4004-16

Wilhelm I, Fazakas C, Molnár J, Haskó J, Végh AG, Cervenak L, Nagyőszi P, Nyúl-Tóth A, Farkas AE, Bauer H, Guillemin GJ, Bauer HC, Váró G, Krizbai IA (2014). Role of Rho/ROCK signaling in the interaction of melanoma cells with the blood-brain barrier. Pigment Cell Melanoma Res. 27:113-23

Wilhelm I, Krizbai IA (2014). In vitro models of the blood-brain barrier for the study of drug delivery to the brain. Mol Pharm. 11:1949-63

Nagyőszi P, Nyúl-Tóth Á, Fazakas C, Wilhelm I, Kozma M, Molnár J, Haskó J, Krizbai IA (2015). Regulation of NOD-like receptors and inflammasome activation in cerebral endothelial cells. J Neurochem. 135:551-64

Krizbai IA, Gasparics Á, Nagyőszi P, Fazakas C, Molnár J, Wilhelm I, Bencs R, Rosivall L, Sebe A (2015). Endothelial-mesenchymal transition of brain endothelial cells: possible role during metastatic extravasation. PLoS One. 10(3):e0119655

Nyúl-Tóth Á, Suciu M, Molnár J, Fazakas C, Haskó J, Herman H, Farkas AE, Kaszaki J, Hermenean A, Wilhelm I, Krizbai IA (2016). Differences in the molecular structure of the blood-brain barrier in the cerebral cortex and white matter: an in silico, in vitro and ex vivo study. Am J Physiol Heart Circ Physiol. 310:H1702-14