In recent years, fluorescent dyes have been frequently used for monitoring mitochondrial membrane potential to evaluate mitochondrial viability and function. However, the reproducibility of the results across laboratories strongly depends upon following well validated and reliable protocols along with the appropriate controls. Herein, we provide a practical user guide for monitoring mitochondrial membrane potential in whole cells using a fluorescent cationic probe. The data analysis of mitochondrial membrane potential we provide is one associated with the impact of xenobiotics such as tobacco smoking on blood-brain barrier endothelial cells including both mouse primary (mBMEC) and a mouse-based endothelial cell line (bEnd3) in a side by side comparison.

Electrophiles and reactive oxygen species (ROS) play a major role in modulating cellular defense mechanisms as well as physiological functions, and intracellular signaling. However, excessive ROS generation (endogenous and exogenous) can create a state of redox imbalance leading to cellular and tissue damage (Ma and He, 2012) [1]. A growing body of research data strongly suggests that imbalanced ROS and electrophile overproduction are among the major prodromal factors in the onset and progression of several cerebrovascular and neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS), stroke, Alzheimer's disease (AD), Parkinson's disease (PD), and aging (Ma and He, 2012; Ramsey et al., 2017; Salminen et al., 2012; Sandberg et al., 2014; Sarlette et al., 2008; Tanji et al., 2013) [1–6]. Cells offset oxidative stress by the action of housekeeping antioxidative enzymes (such as superoxide dismutase, catalase, glutathione peroxidase) as well direct and indirect antioxidants (Dinkova-Kostova and Talalay, 2010) [7]. The DNA sequence responsible for modulating the antioxidative and cytoprotective responses of the cells has been identified as the antioxidant response element (ARE), while the nuclear factor erythroid 2-related factor (NRF2) is the major regulator of the xenobiotic-activated receptor (XAR) responsible for activating the ARE-pathway, thus defined as the NRF2-ARE system (Ma and He, 2012) [1]. In addition, the interplay between the NRF2-ARE system and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-ĸB, a protein complex that controls cytokine production and cell survival), has been further investigated in relation to neurodegenerative and neuroinflammatory disorders. On these premises, we provide a review analysis of current understanding of the NRF2-NF-ĸB interplay, their specific role in major CNS disorders, and consequent therapeutic implication for the treatment of neurodegenerative and cerebrovascular diseases.

The development of realistic in vitro blood-brain barrier (BBB) models that recapitulate the physiological parameters and molecular aspect of the neurovascular unit (NVU) is of fundamental importance not only in CNS drug discovery but also in translational research. Successful modeling of the NVU would provide an invaluable tool to aid in dissecting out the pathological factors, mechanism of action (and corresponding targets) prodromal to the onset of CNS disorders. The field of BBB in vitro modeling has seen many radical changes in the last few years with the introduction on novel technologies and methods to improve over existing models and develop new ones. Therefore, the goal of this review is to provide the readers with updated technical and operational details concerning current BBB platforms with special focus on stem cell technology used to establish a functional BBB model in vitro. Furthermore, we provide a detailed update on rapidly advancing 3D printing technologies used for engineering BBB models which use is now fast expanding among researchers.