Background Excessive inflammation might activate and injure the blood-brain barrier (BBB), a common feature of many central nervous system (CNS) disorders. We previously developed an in vitro BBB injury model in which the organophosphate paraoxon (PX) affects the BBB endothelium by attenuating junctional protein expression leading to weakened barrier integrity. The objective of this study was to investigate the inflammatory cellular response at the BBB to elucidate critical pathways that might lead to effective treatment in CNS pathologies in which the BBB is compromised. We hypothesized that caspase-1, a core component of the inflammasome complex, might have important role in BBB function since accumulating evidence indicates its involvement in brain inflammation and pathophysiology. Methods An in vitro human BBB model was employed to investigate BBB functions related to inflammation, primarily adhesion and transmigration of peripheral blood mononuclear cells (PBMCs). Caspase-1 pathway was studied by measurements of its activation state and its role in PBMCs adhesion, transmigration, and BBB permeability were investigated using the specific caspase-1 inhibitor, VX-765. Expression level of adhesion and junctional molecules and the secretion of pro-inflammatory cytokines were measured in vitro and in vivo at the BBB endothelium after exposure to PX. The potential repair effect of blocking caspase-1 and downstream molecules was evaluated by immunocytochemistry, ELISA, and Nanostring technology. Results PX affected the BBB in vitro by elevating the expression of the adhesion molecules E-selectin and ICAM-1 leading to increased adhesion of PBMCs to endothelial monolayer, followed by elevated transendothelial-migration which was ICAM-1 and LFA-1 dependent. Blocking caspase-8 and 9 rescued the viability of the endothelial cells but not the elevated transmigration of PBMCs. Inhibition of caspase-1, on the other hand, robustly restored all of barrier insults tested including PBMCs adhesion and transmigration, permeability, and VE-cadherin protein levels. The in vitro inflammatory response induced by PX and the role of caspase-1 in BBB injury were corroborated in vivo in isolated blood vessels from hippocampi of mice exposed to PX and treated with VX-765. Conclusions These results shed light on the important role of caspase-1 in BBB insult in general and specifically in the inflamed endothelium, and suggest therapeutic potential for various CNS disorders, by targeting caspase-1 in the injured BBB.
Living in a social environment requires the ability to respond to specific social stimuli and to incorporate information obtained from prior interactions into future ones. One of the mechanisms that facilitates social interaction is pheromone-based communication. In Drosophila melanogaster, the male-specific pheromone cis-vaccenyl acetate (cVA) elicits different responses in male and female flies, and functions to modulate behavior in a context and experience-dependent manner. Although it is the most studied pheromone in flies, the mechanisms that determine the complexity of the response, its intensity and final output with respect to social context, sex and prior interaction, are still not well understood. Here we explored the functional link between social interaction and pheromone-based communication and discovered an odorant binding protein that links social interaction to sex specific changes in cVA related responses. Odorant binding protein 69a (Obp69a) is expressed in auxiliary cells and secreted into the olfactory sensilla. Its expression is inversely regulated in male and female flies by social interactions: cVA exposure reduces its levels in male flies and increases its levels in female flies. Increasing or decreasing Obp69a levels by genetic means establishes a functional link between Obp69a levels and the extent of male aggression and female receptivity. We show that activation of cVA-sensing neurons is sufficeint to regulate Obp69a levels in the absence of cVA, and requires active neurotransmission between the sensory neuron to the second order olfactory neuron. The cross-talk between sensory neurons and non-neuronal auxiliary cells at the olfactory sensilla, represents an additional component in the machinery that promotes behavioral plasticity to the same sensory stimuli in male and female flies.
The objective of this study was to investigate the molecular response to damage at the blood brain barrier (BBB) and to elucidate critical pathways that might lead to effective treatment in central nervous system (CNS) pathologies in which the BBB is compromised. We have used a human, stem-cell derived in-vitro BBB injury model to gain a better understanding of the mechanisms controlling BBB integrity. Chemical injury induced by exposure to an organophosphate resulted in rapid lipid peroxidation, initiating a ferroptosis-like process. Additionally, mitochondrial ROS formation (MRF) and increase in mitochondrial membrane permeability were induced, leading to apoptotic cell death. Yet, these processes did not directly result in damage to barrier functionality, since blocking them did not reverse the increased permeability. We found that the iron chelator, Desferal© significantly decreased MRF and apoptosis subsequent to barrier insult, while also rescuing barrier integrity by inhibiting the labile iron pool increase, inducing HIF2α expression and preventing the degradation of Ve-cadherin specifically on the endothelial cell surface. Moreover, the novel nitroxide JP4-039 significantly rescued both injury-induced endothelium cell toxicity and barrier functionality. Elucidating a regulatory pathway that maintains BBB integrity illuminates a potential therapeutic approach to protect the BBB degradation that is evident in many neurological diseases.
In highly polyandrous species, where females mate with multiple males within a single fertility period, there is typically a high level of sperm competition. To cope with this challenge, males apply various behavioral and physiological strategies to maximize their paternity rates. Previous studies in Drosophila melanogaster established a link between the composition of the social environment and the reproductive success of individual male flies. While most studies until now focused on the adaptive responses of male flies to the presence of rival males, little is known about whether the outcomes of sexual interactions with female partners affect male-male social interactions in a competitive environment such as the social group. Here we show that repeated failures to mate promote a coordinated physiological and behavioral responses that can serve to increase paternity chances over mating rivals. We exposed male flies to sexual deprivation or successful mating and analyzed the behavioral repertoires of individuals within groups and the structure of their emerging social networks. We discovered that failures to mate and successful mating generate distinct emergent group interactions and structures, where sexually deprived males form low density social networks and actively minimize their encounters with other group members, while increasing their aggressive behavior. In addition, sexually deprived male flies elevate the production of seminal fluid proteins (known to facilitate post-mating responses in females) and extend mating duration upon mating with receptive females, altogether leading to reduced re-mating rates. Our results demonstrate the existence of a flexible mating strategy that may provide a short-term fitness advantage over competing rivals and pave the path for using simple model organisms to dissect the neurobiology of social plasticity as coping strategy to living in a highly dynamic environment as the social domain.
The blood-brain barrier serves as the guardian of the CNS, tightly regulating the movement of ions, molecules, and cells between the circulatory system and brain. We have used an in-vitro BBB injury model to gain a better understanding of the mechanisms controlling BBB integrity. This model exposes a co-culture of human stem-cell derived brain-like endothelial cells and brain pericytes that mimic the BBB, to the organophosphate paraoxon. This exposure results in rapid lipid peroxidation, initiating a ferroptosis-like process. Additionally, mitochondrial ROS formation (MRF) and increase in mitochondrial membrane permeability, were also detected leading to apoptotic cell death. Yet, these processes do not directly result in damage to barrier functionality since blocking them does not reverse the increased permeability. We found that the iron chelator, Desferal© significantly decreases MRF and apoptosis subsequent to paraoxon exposure, while also rescuing barrier integrity by inhibiting the liable iron pool increase, inducing HIF2α expression and preventing the degradation of Ve-cadherin on the cell surface. Moreover, the novel nitroxide JP4-039 significantly rescues both injury-induced endothelium cell toxicity and barrier functionality.
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