[Blood-Brain Barrier Transwell Modeling].

In vitro blood-brain barrier (BBB) modeling with the use of the brain endothelial cells grown on a transwell membrane is widely used to investigate BBB disorders and factors intended to ameliorate these pathologies. Endothelial cells, due to tight junction proteins, ensure selective permeability for a number of substances. The low integrity (i.e., high permeability) of the BBB model, as compared to the physiological one, complicates evaluation of the effects caused by different agents. Thus, the selection of conditions to improve barrier integrity is an essential task. In this study, mouse brain endothelial cells bEnd.3 are used in experiments on transwell modeling. To determine which factors enhance BBB integrity, the effects of the cultivation medium, the number of cells during seeding, the state of the transwell membrane, and cultivation in the presence or in the absence of primary mouse neurons and matrigel as a matrix on the passage of a fluorescent label through the cell monolayer were assessed. The effect of fetal bovine serum on the tight junction protein claudin-5 was analyzed by immunocytochemistry. The obtained cultivation parameter data facilitate the solution to the problem of low integrity of the BBB transwell model and bring the model closer to the physiologically relevant indicators.

Anti-amyloid Therapy of Alzheimer's Disease: Current State and Prospects.

Drug development for the treatment of Alzheimer's disease (AD) has been for a long time focused on agents that were expected to support endogenous ß-amyloid (Aß) in a monomeric state and destroy soluble Aß oligomers and insoluble Aß aggregates. However, this strategy has failed over the last 20 years and was eventually abandoned. In this review, we propose a new approach to the anti-amyloid AD therapy based on the latest achievements in understanding molecular causes of cerebral amyloidosis in AD animal models.

[Isomerization of Asp7 increases the toxic effects of amyloid ß and its phosphorylated form in SH-SY5Y neuroblastoma cells].

The generation of amyloid ß (Aß) toxic oligomers during the formation of senile plaques and amyloid fibrils is thought to play a central role in the onset and progression of Alzheimer's disease. Aß production is a physiological process, but the factors that trigger a transition to pathogenic Aß aggregation remain unknown. Posttranslational modifications of Aß could potentially induce the transition. The effects of Aß and its modified forms containing isomerized Asp7, phosphorylated Ser8, or both, were studied in SH-SY5Y human neuroblastoma cells. Asp7 isomerization of was shown to increase cytotoxicity of both the intact and phosphorylated Aß. An increase in cytotoxicity was not associated with an increased internalization of the isomerized Asp7-containing Aß or an influence on the function of mitochondria or reduced glutathione and reactive oxygen species levels. The nitric oxide (NO) level was identified as a determinant of the cytotoxic effect of isomerized Asp7-containing peptides, a decrease in NO level correlating with an increase in cytotoxicity.

[Heat-shock protein HSP70 protects neuroblastoma cells SK-N-SH from the neurotoxic effects hydrogen peroxide and the ß-amyloid peptide].

Neuronal cell death in Alzheimer's disease is associated with the development of oxidative stress caused by the reactive oxygen species (ROS), which can be generated as a result of the effect of beta-amyloid peptides. One of the sources of ROS is hydrogen peroxide, inducing the apoptosis and necrosis of neural tissue cells. The mechanism of hydrogen peroxide apoptotic action includes launching signaling pathways that involve protein kinases PI3K, p38MAPK, JNK and ERK. Oxidative stress leads to increased synthesis of heat-shock proteins in the cells including HSP70. It was shown that the exogenous HSP70 could reduce generation of ROS in cells. In this study, we determined how HSP70 affected apoptosis and necrosis in human neuroblastoma cells SK-N-SH, induced by hydrogen peroxide and ß-amyloid peptide Aß(1-42). It was shown that HSP70 reduces the cytotoxic effects of hydrogen peroxide and beta-amyloid, and protein kinases PI3K and JNK play an important role in the mechanism of HSP70 protective effect on the peroxide induced apoptosis in SK-N-SH cells.