Ultrastructural Remodeling of the Blood-Brain Barrier and Neurovascular Unit by Lipopolysaccharide-Induced Neuroinflammation.

The blood-brain barrier (BBB) is an interface primarily comprised of brain endothelial cells (BECs), separating the central nervous system (CNS) from the systemic circulation while carefully regulating the transport of molecules and inflammatory cells, and maintaining the required steady-state environment. Inflammation modulates many BBB functions, but the ultrastructural cytoarchitectural changes of the BBB with inflammation are understudied. Inflammation was induced in male 8-10-week-old CD-1 mice with intraperitoneal lipopolysaccharide (LPS), using a regimen (3 mg/kg at 0, 6, and 24 h) that caused robust BBB disruption but had minimal lethality at the study timepoint of 28 h. Perfusion-fixed brains were collected and the frontal cortical layer III regions were analyzed using a transmission electron microscopy (TEM). The LPS-treated mice had pronounced ultrastructural remodeling changes in BECs that included plasma membrane ruffling, increased numbers of extracellular microvesicles, small exosome formation, aberrant BEC mitochondria, increased BEC transcytosis, while tight junctions appeared to be unaltered. Aberrant pericytes were contracted with rounded nuclei and a loss of their elongated cytoplasmic processes. Surveilling microglial cells were attracted to the neurovascular unit (NVU) of BECs, and astrocyte detachment and separation were associated with the formation of a perivascular space and pericapillary edema. The LPS treatment resulted in numerous ultrastructural aberrant remodeling changes to the neurovascular unit's BECs, microglia, pericytes, and astrocytes. In summary, a disturbance of the NVU morphology is a consequence of LPS treatment.

Why Are Perivascular Spaces Important?

Perivascular spaces (PVS) and their enlargement (EPVS) have been gaining interest as EPVS can be visualized non-invasively by magnetic resonance imaging (MRI) when viewing T-2-weighted images. EPVS are most commonly observed in the regions of the basal ganglia and the centrum semiovale; however, they have also been identified in the frontal cortex and hippocampal regions. EPVS are known to be increased in aging and hypertension, and are considered to be a biomarker of cerebral small vessel disease (SVD). Interest in EPVS has been significantly increased because these PVS are now considered to be an essential conduit necessary for the glymphatic pathway to provide the necessary efflux of metabolic waste. Metabolic waste includes misfolded proteins of amyloid beta and tau that are known to accumulate in late-onset Alzheimer's disease (LOAD) within the interstitial fluid that is delivered to the subarachnoid space and eventually the cerebral spinal fluid (CSF). The CSF acts as a sink for accumulating neurotoxicities and allows clinical screening to potentially detect if LOAD may be developing early on in its clinical progression via spinal fluid examination. EPVS are thought to occur by obstruction of the PVS that associates with excessive neuroinflammation, oxidative stress, and vascular stiffening that impairs flow due to a dampening of the arterial and arteriolar pulsatility that aids in the convective flow of the metabolic debris within the glymphatic effluxing system. Additionally, increased EPVS has also been associated with Parkinson's disease and non-age-related multiple sclerosis (MS).

Reactive Microgliosis in Sepsis-Associated and Acute Hepatic Encephalopathies: An Ultrastructural Study.

Sepsis and acute liver failure are associated with severe endogenous intoxication. Microglia, which are the resident immune brain cells, play diverse roles in central nervous system development, surveillance, and defense, as well as contributing to neuroinflammatory reactions. In particular, microglia are fundamental to the pathophysiology of reactive toxic encephalopathies. We analyzed microglial ultrastructure, morphotypes, and phagocytosis in the sensorimotor cortex of cecal ligation and puncture (CLP) and acetaminophen-induced liver failure (AILF) Wistar rats. A CLP model induced a gradual shift of ~50% of surveillant microglia to amoeboid hypertrophic-like and gitter cell-like reactive phenotypes with active phagocytosis and frequent contacts with damaged neurons. In contrast, AILF microglia exhibited amoeboid, rod-like, and hypertrophic-like reactive morphotypes with minimal indications for efficient phagocytosis, and were mostly in contact with edematous astrocytes. Close interactions of reactive microglia with neurons, astrocytes, and blood-brain barrier components reflect an active contribution of these cells to the tissue adaptation and cellular remodeling to toxic brain damage. Partial disability of reactive microglia may affect the integrity and metabolism in all tissue compartments, leading to failure of the compensatory mechanisms in acute endogenous toxic encephalopathies.

Astroglia in Sepsis Associated Encephalopathy.

Cellular pathophysiology of sepsis associated encephalopathy (SAE) remains poorly characterised. Brain pathology in SAE, which is manifested by impaired perception, consciousness and cognition, results from multifactorial events, including high levels of systemic cytokines, microbial components and endotoxins, which all damage the brain barriers, instigate neuroinflammation and cause homeostatic failure. Astrocytes, being the principal homeostatic cells of the central nervous system contribute to the brain defence against infection. Forming multifunctional anatomical barriers, astroglial cells maintain brain-systemic interfaces and restrict the damage to the nervous tissue. Astrocytes detect, produce and integrate inflammatory signals between immune cells and cells of brain parenchyma, thus regulating brain immune response. In SAE astrocytes are present in both reactive and astrogliopathic states; balance between these states define evolution of pathology and neurological outcomes. In humans pathophysiology of SAE is complicated by frequent presence of comorbidities, as well as age-related remodelling of the brain tissue with senescence of astroglia; these confounding factors further impact upon SAE progression and neurological deficits.

Mobilisation and redistribution of multivesicular bodies to the endfeet of reactive astrocytes in acute endogenous toxic encephalopathies.

Endogenous toxicity caused by systemic inflammation as well as by acute liver failure triggers a wide range of dysfunctional disorders in the brain ranging from delirium and acute psychosis to coma. Astrocytes, the main homeostatic cells of the central nervous system (CNS), play a key role in pathophysiology of neurotoxic insults. We examined the cecal ligation and puncture (CLP) and acetaminophen-induced liver failure (AILF) of Wistar rats, and analysed ultrastructure of astrocytes in the brain cortex and subcortical white matter of sensorimotor zone with transmission electron microscopy. Both models showed significant similarities in reactive changes of astroglial endosomal machinery. In survived animals (with relative prevalence in the CLP-model), at 24 h after intervention we found an increase in number of multivesicular bodies (MVBs) in astroglial perikarya and astroglial processes. In particular, the number of MVBs substantially (3 times of control values) increased in the perivascular astroglial endfeet. Increased number of MVBs in astrocytes was associated with the lesser degree of intracellular oedema and with signs of compensated oedematous tissue changes. In deceased animals, up to 24 h after intervention, single MVBs were localised mainly in astroglial perikarya, and their number was not significantly changed compared to control. Activation of astroglial endosomal-exosomal machinery in both models reflects the uniform pattern of reactive changes of astroglia in these two systemic conditions and may represent activation of astroglial defence in sepsis-associated encephalopathy (SAE) and acute hepatic encephalopathy (AHE). Our data highlight the special role of astroglial adaptive activity in the counterbalancing of an impaired brain homeostasis under action of endogenous toxins. Accumulation of MVBs in astrocytic processes indicates the activation of their intercellular and gliovascular interactions through endo- and exocytosis in SAE and AHE.