Neuroinflammatory Dysfunction of the Blood-Brain Barrier and Basement Membrane Dysplasia Play a Role in the Development of Drug-Resistant Epilepsy

Abstract

Drug-resistance epilepsy (DRE) is a key problem in neurology. It is possible that damage to the blood-brain barrier (BBB) may affect resistance in DRE. The aim of this work was to assess the damage and dysfunction in the BBB in the area of epileptic foci in patients with DRE under conditions of neuroinflammation. The changes to the BBB in temporal lobe epilepsy (by immunohistochemistry and transmission electron microscopy), levels of neuroinflammatory proteins, and cytokine levels in the blood (by multiplex analysis) were studied. Increased levels of vascular endothelial growth factor (VEGF) and growth-regulated protein (GRO), and decreased levels of epidermal growth factor (EGF) in plasma, combined with overexpression of the VEGF-A receptor by endotheliocytes were detected. Malformation-like growths of the basement membrane of the capillaries of the brain complicate the delivery of antiepileptic drugs (AEDs). Dysplasia of the basement membrane is the result of inadequate reparative processes in chronic inflammation. In conclusion, it should be noted that damage to the microcirculatory network of the brain should be considered one of the leading factors contributing to DRE.

Keywords: VEGF; basement membrane; blood–brain barrier; cytokines; drug-resistant epilepsy; dysplasia; electron microscopy; immunohistochemistry; multiplex analysis; neuroinflammation.

Figure 1

Morphological changes in the white matter of the brain: (a) moderately expanded perivascular space (thick arrow), a pronounced VEGF-positive reaction (IHC staining), and (thin arrows) a patient with DRE; (b) no VEGF-positive reaction in the Control 1 group of patients without epilepsy; (c) optical density of VEGF-positive cells in the DRE group (orange bar) vs. Control 1 group (green bar) (***—p < 0.001). IHC—immunohistochemistry; DRE—drug-resistant epilepsy; VEGF—vascular endothelial growth factor. VEGF-A antibody staining, hematoxylin staining, magnification ×400.

Figure 2

The level of antibodies in the blood serum of patients with DRE compared with the control group: (a) NMDA-2A receptor; (b) GABA A receptor; (c) GAD65/67; (d) Glu. DRE group vs. Control 1 group (*—p < 0.001).

Figure 3

Fragments of capillaries with unevenly thickened areas of the basement membrane (BM). (a) Bumpy, in some places sharply expanded BM limits pericytes (Pc) with hyperchromic cytoplasm. From the outside, powerful bundles of gliofilaments (GlF) are adjacent to the capillary. (b) Loose, swollen BM has foci of encrustation with destroyed membranes and calcifications (arrows). (c) Cross-section of the capillary: pronounced expansion of the BM also occurs due to the germination of collagen fibers (CF) in it, causing asymmetry of the vessel; fibrosis is observed in some Pc. (d) A longitudinal section of the capillary in which the transverse growth of the BM and its powerful fibrosis led to a critical narrowing of the lumen. CL—capillary lumen; Ec —endotheliocyte. Electronograms: (a)—×8200; (b)—×11,500; (c,d)—×9900.

Figure 3

Fragments of capillaries with unevenly thickened areas of the basement membrane (BM). (a) Bumpy, in some places sharply expanded BM limits pericytes (Pc) with hyperchromic cytoplasm. From the outside, powerful bundles of gliofilaments (GlF) are adjacent to the capillary. (b) Loose, swollen BM has foci of encrustation with destroyed membranes and calcifications (arrows). (c) Cross-section of the capillary: pronounced expansion of the BM also occurs due to the germination of collagen fibers (CF) in it, causing asymmetry of the vessel; fibrosis is observed in some Pc. (d) A longitudinal section of the capillary in which the transverse growth of the BM and its powerful fibrosis led to a critical narrowing of the lumen. CL—capillary lumen; Ec —endotheliocyte. Electronograms: (a)—×8200; (b)—×11,500; (c,d)—×9900.

Figure 4

Malformed growth of the basement membrane of the capillaries of the brain. (a) A small capillary with erythrocytes (Er) in the capillary lumen (CL) is surrounded by a basement membrane (BM), uneven in the thickness and density of its matrix. The arrows indicate the BM processes that are growing into the surrounding brain tissue. Endotheliocytes (Ec) and pericytes (Pc) are dystrophic, with a minimal set of cytoplasmic organelles. (b) A small capillary with a narrowed CL and a large Pc containing lipofuscin granules (LF) is surrounded by a network of branched BM. (c) A fragment of capillaries with Er in the lumen and a large edematous nucleus of the endotheliocyte (NEc). The BM is multilayered, and its outer zone creates a picture of a radiant crown with numerous parallel, sometimes branched processes. They are not always close together, creating pericytic cavities, which indicates a dynamically developing process of formation of vascular malformations. (d) A malformation-like tangle of multiple, branched, sometimes merged into a single shapeless conglomerate, processes of the BM. GlF—gliofilaments; NPc—nucleus of the pericyte. Electronograms: (a)—×11,500; (b)—×16,500; (c,d)—×6000.

Figure 4

Malformed growth of the basement membrane of the capillaries of the brain. (a) A small capillary with erythrocytes (Er) in the capillary lumen (CL) is surrounded by a basement membrane (BM), uneven in the thickness and density of its matrix. The arrows indicate the BM processes that are growing into the surrounding brain tissue. Endotheliocytes (Ec) and pericytes (Pc) are dystrophic, with a minimal set of cytoplasmic organelles. (b) A small capillary with a narrowed CL and a large Pc containing lipofuscin granules (LF) is surrounded by a network of branched BM. (c) A fragment of capillaries with Er in the lumen and a large edematous nucleus of the endotheliocyte (NEc). The BM is multilayered, and its outer zone creates a picture of a radiant crown with numerous parallel, sometimes branched processes. They are not always close together, creating pericytic cavities, which indicates a dynamically developing process of formation of vascular malformations. (d) A malformation-like tangle of multiple, branched, sometimes merged into a single shapeless conglomerate, processes of the BM. GlF—gliofilaments; NPc—nucleus of the pericyte. Electronograms: (a)—×11,500; (b)—×16,500; (c,d)—×6000.