Lab-on-a-chip tool for the investigation of biological barriers

Epithelial outer, and endothelial inner barriers of the body are important defense systems to maintain homeostasis, and play a crucial role in drug absorption and transport. Culture models of biological barriers are important tools to study physiological functions, transport mechanisms, drug delivery and pathological processes. Tight intercellular junctions are fundamental features of epithelial and endothelial barriers in vivo, which are reflected in high electrical resistance and low passive permeability for hydrophilic and hydrophobic compounds. These physical and physico-chemical parameters describe the barrier integrity and function accurately. In more detail, the trans-endothelial/epithelial electric resistance (TEER) represents the paracellular ion mobility which gives information about the tightness of the intercellular junctions. The passive permeability of small, charged or electroneutral molecules describe the para- and transcellular pathways. Another important contributor to the barrier function could be the high negative surface charge of the cell monolayers, yet this is the least explored field of the physical properties of the biological barriers. In the past 10 years, besides static models cultured on inserts, dynamic lab-on-a-chip (LOC)/organ-on-chip (OC) devices were developed to study cell-cell interactions, molecular pathways, pathological conditions and drug delivery in biological barriers. These models incorporate the use of fluid flow enabling the investigation of physiological-like functions such as receptor and mechanosensor expression, transport mechanisms, pathologies and drug delivery. LOC/OC devices became important tools since they provide controlled conditions for cellular signaling and external stimulus, and are able to track the development and changes in the barrier function. Confluent monolayers of adherent epithelial or endothelial cells grown on culture inserts are widely used static models for intestinal, lung and blood-brain barriers (BBB). The field of biochips modeling gut and blood-brain barriers has been rapidly evolving during the last fifteen years, while the focus of the devices being shifted according to the main interest of the corresponding study. Our aim was to develop a versatile LOC device that can be used for different kinds of biological barriers, and thus, for different kinds of approaches, as well. Also, the measurement of all crucial physical parameters, such as TEER and permeability assays, are included. The negative surface charge of the cell layers is an important element of the defense system of barriers. A quantitative description of the surface electric properties of cell layers forming biological barriers is essential for the broader understanding of their function in physiological processes and diseases. The role of the negatively charged glycocalyx of the vascular endothelial barrier, for example, is well known in the protection of the cardiovascular system, and important in microbiological infections. Despite the recent boom in LOC devices, no biochip to determine the surface charge of intact cell layers forming biological barriers has been published, yet

Simvastatin, edaravone and dexamethasone protect against kainate-induced brain endothelial cell damage

Excitotoxicity is a central pathological pathway in many neurological diseases with blood-brain barrier (BBB) dysfunction. Kainate, an exogenous excitotoxin, induces epilepsy and BBB damage in animal models, but the direct effect of kainate on brain endothelial cells has not been studied in detail. Our aim was to examine the direct effects of kainate on cultured cells of the BBB and to test three anti-inflammatory and antioxidant drugs used in clinical practice, simvastatin, edaravone and dexamethasone, to protect against kainate-induced changes.Primary rat brain endothelial cell, pericyte and astroglia cultures were used to study cell viability by impedance measurement. BBB permeability was measured on a model made from the co-culture of the three cell types. The production of nitrogen monoxide and reactive oxygen species was followed by fluorescent probes. The mRNA expression of kainate receptors and nitric oxide synthases were studied by PCR.Kainate damaged brain endothelial cells and made the immunostaining of junctional proteins claudin-5 and zonula occludens-1 discontinuous at the cell border indicating the opening of the barrier. The permeability of the BBB model for marker molecules fluorescein and albumin and the production of nitric oxide in brain endothelial cells were increased by kainate. Simvastatin, edaravone and dexamethasone protected against the reduced cell viability, increased permeability and the morphological changes in cellular junctions caused by kainate. Dexamethasone attenuated the elevated nitric oxide production and decreased the inducible nitric oxide synthase (NOS2/iNOS) mRNA expression increased by kainate treatment.Kainate directly damaged cultured brain endothelial cells. Simvastatin, edaravone and dexamethasone protected the BBB model against kainate-induced changes. Our results confirmed the potential clinical usefulness of these drugs to attenuate BBB damage

Protection of cultured brain endothelial cells from cytokine-induced damage by α-melanocyte stimulating hormone

L

Temporal instability of salience network activity in migraine with aura.

This study aims to investigate whether intra-network dynamic functional connectivity and causal interactions of the salience network is altered in the interictal term of migraine. 32 healthy controls, 37 migraineurs without aura and 20 migraineurs with aura were recruited. Participants underwent a T1-weighted scan and resting-state fMRI protocol inside a 1.5T MR scanner. We obtained average spatial maps of resting-state networks using group independent component analysis, which yielded subject-specific time series via a dual regression approach. Salience network ROIs (bilateral insulae and prefrontal cortices, dorsal anterior cingulate cortex) were obtained from the group average map via cluster-based thresholding. To describe intra-network connectivity, average and dynamic conditional correlation was calculated. Causal interactions between the default-mode, dorsal attention and salience network were characterised by spectral Granger's causality. Time-averaged correlation was lower between the right insula and prefrontal cortex in migraine without aura vs. with aura and healthy controls (p<0.038, p<0.037). Variance of dynamic conditional correlation was higher in migraine with aura vs. healthy controls and migraine with aura vs. without aura between the right insula and dorsal anterior cingulate cortex (p<0.011, p<0.026), and in migraine with aura vs. healthy controls between the dorsal anterior cingulate and left prefrontal cortex (p<0.021). Causality was weaker in the <0.05 Hz frequency range between the salience and dorsal attention networks in migraine with aura (p<0.032). Overall, migraineurs with aura exhibit more fluctuating connections in the salience network, which also affect network interactions, and could be connected to altered cortical excitability and increased sensory gain

Evaluation of transorbital sonography measures of optic nerve diameter in the context of global and regional brain volume in multiple sclerosis

Transorbital sonography (TOS) could be a swift and convenient method to detect the atrophy of the optic nerve, possibly providing a marker that might reflect other quantitative structural markers of multiple sclerosis (MS). Here we evaluate the utility of TOS as a complementary tool for assessing optic nerve atrophy, and investigate how TOS-derived measures correspond to volumetric brain markers in MS. We recruited 25 healthy controls (HC) and 45 patients with relapsing-remitting MS and performed B-mode ultrasonographic examination of the optic nerve. Patients additionally underwent MRI scans to obtain T1-weighted, FLAIR and STIR images. Optic nerve diameters (OND) were compared between HC, MS patients with and without history of optic neuritis (non-ON) using a mixed-effects ANOVA model. The relationship between within-subject-average OND and global and regional brain volumetric measures was investigated using FSL SIENAX, voxel-based morphometry and FSL FIRST. OND was significantly different between HC-MS (HC = 3.2 ± 0.4 mm, MS = 3 ± 0.4 mm; p < 0.019) and we found significant correlation between average OND and normalised whole brain (β = 0.42, p < 0.005), grey matter (β = 0.33, p < 0.035), white matter (β = 0.38, p < 0.012) and ventricular cerebrospinal fluid volume (β = - 0.36, p < 0.021) in the MS group. History of ON had no impact on the association between OND and volumetric data. In conclusion, OND is a promising surrogate marker in MS, that can be simply and reliably measured using TOS, and its derived measures correspond to brain volumetric measures. It should be further explored in larger and longitudinal studies

Connection between microstructural alterations detected by diffusion MRI and cognitive dysfunction in MS: A model-free analysis approach

Cognitive decline is a prominent symptom of MS. Clear connection between cognitive status and white matter microstructural changes has not been unequivocally observed to date.To characterise the relationship between white matter microstructure and cognitive performance a partial least squares (PLS) approach was used.53 RR MS patients' T1 and DTI images and BICAMS subtests were used in our analysis. Standard FSL pipeline was used to obtain diffusion parameters. A PLS approach was applied to reveal the diffusion parameter patterns responsible for the cognitive dysfunction.The first latent variable (LV) was mainly associated with demyelination, while the second and third explained axonal damage. While the first two LV represented mainly Brief Visuospatial Memory Test (BVMT) and Single Digit Modality Test (SDMT), the third LV depicted diffusion alterations mainly the verbal subtest. The first LVs spatial map showed demyelination in the corpus callosum. The second LVs spatial map showed the diffusion alterations in the thalamus. The third LV depicted diffusion alterations in the putative left superior longitudinal fascicle.Visual memory demanding tasks versus language functions depend on distinct patterns of diffusion parameters and the spatial organisation. Axial diffusivity alterations, a putative marker of irreversible axonal loss explained around 20% of variability in the cognitive functions

Modulation of cortical resting state functional connectivity during a visuospatial attention task in Parkinson's disease

Visual dysfunction is a recognized early symptom of Parkinson's disease (PD) that partly scales motor symptoms, yet its background is heterogeneous. With additional deficits in visuospatial attention, the two systems are hard to disentangle and it is not known whether impaired functional connectivity in the visual cortex is translative in nature or disrupted attentional modulation also contributes. In this study, we investigate functional connectivity modulation during a visuospatial attention task in patients with PD. In total, 15 PD and 16 age-matched healthy controls performed a visuospatial attention task while undergoing fMRI, in addition to a resting-state fMRI scan. Tensorial independent component analysis was used to investigate task-related network activity patterns. Independently, an atlas-based connectivity modulation analysis was performed using the task potency method. Spearman's rank correlation was calculated between task-related network expression, connectivity modulation, and clinical characteristics. Task-related networks including mostly visual, parietal, and prefrontal cortices were expressed to a significantly lesser degree in patients with PD (p < 0.027). Resting-state functional connectivity did not differ between the healthy and diseased cohorts. Connectivity between the precuneus and ventromedial prefrontal cortex was modulated to a higher degree in patients with PD (p < 0.004), while connections between the posterior parietal cortex and primary visual cortex, and also the superior frontal gyrus and opercular cortex were modulated to a lesser degree (p < 0.001 and p < 0.011). Task-related network expression and superior frontal gyrus-opercular cortex connectivity modulation were significantly associated with UPDRSIII motor scores and the Hoehn-Yahr stages (R = -0.72, p < 0.006 and R = -0.90, p < 0.001; R = -0.68, p < 0.01 and R = -0.71, p < 0.007). Task-related networks function differently in patients with PD in association with motor symptoms, whereas impaired modulation of visual and default-mode network connectivity was not correlated with motor function

Lidocaine turns the surface charge of biological membranes more positive and changes the permeability of blood-brain barrier culture models

The surface charge of brain endothelial cells forming the blood-brain barrier (BBB) is highly negative due to phospholipids in the plasma membrane and the glycocalyx. This negative charge is an important element of the defense systems of the BBB. Lidocaine, a cationic and lipophilic molecule which has anaesthetic and antiarrhytmic properties, exerts its actions by interacting with lipid membranes. Lidocaine when administered intravenously acts on vascular endothelial cells, but its direct effect on brain endothelial cells has not yet been studied. Our aim was to measure the effect of lidocaine on the charge of biological membranes and the barrier function of brain endothelial cells. We used the simplified membrane model, the bacteriorhodopsin (bR) containing purple membrane of Halobacterium salinarum and culture models of the BBB. We found that lidocaine turns the negative surface charge of purple membrane more positive and restores the function of the proton pump bR. Lidocaine also changed the zeta potential of brain endothelial cells in the same way. Short-term lidocaine treatment at a 10 μM therapeutically relevant concentration did not cause major BBB barrier dysfunction, substantial change in cell morphology or P-glycoprotein efflux pump inhibition. Lidocaine treatment decreased the flux of a cationic lipophilic molecule across the cell layer, but had no effect on the penetration of hydrophilic neutral or negatively charged markers. Our observations help to understand the biophysical background of the effect of lidocaine on biological membranes and draws the attention to the interaction of cationic drug molecules at the level of the BBB