Dr Shikha Nangia – The Blood-Brain Barrier: More than Just a Barrier

Neurodegenerative disorders present a major cause of death and disability worldwide. Treatments are typically expensive, non-efficient, and invasive. Although scientists are committed to finding better treatment strategies, the challenge of penetrating the blood-brain barrier remains. This highly selective envelope protects our brain from harmful substances but also prevents drugs from reaching the brain when needed. Dr Shikha Nangia at Syracuse University, USA, focuses on understanding the molecular structure of this complex interface to ultimately facilitate the transport of drugs across the blood-brain barrier.

The Blood-Brain Barrier: A Blessing and a Curse

The blood-brain barrier is a highly selective barrier that allows only vital nutrients (such as glucose and water) to enter the brain and filters toxic substances present in the bloodstream. Its integrity is crucial to our survival but in some pathological contexts, it may become leaky.

Neurodegenerative disorders such as Alzheimer’s disease and Parkinson’s disease are characterised by the progressive degeneration of neurons in various brain regions. Currently available drugs are not able to effectively penetrate the blood-brain barrier. Overcoming this difficulty would open a new and unchartered horizon for the development of treatment strategies that could benefit a range of neurogenerative diseases.

Dr Shikha Nangia at Syracuse University, USA, is determined to overcome this challenge. Together with her team, Dr Nangia is progressively unravelling the secrets of blood-brain barrier permeability using advanced computational techniques.

Gatekeepers of the Blood-Brain Barrier

Dr Nangia often uses the analogy of the hook-and-loop fastener known as ‘Velcro’ to explain how the blood-brain barrier functions. Between the brain and the blood, there is a single layer of cells that are strongly attached to each other. This layer is principally composed of blood vessel cells (called endothelial cells), stitched together by tight junctions acting as gatekeepers. These tight junctions are formed between membranes of two adjacent cells and are primarily made of proteins named ‘claudins’. Claudins were identified for the first time in 1998 by Japanese researchers and were named after the Latin verb claudere ‘to close’, already suggesting the barrier role of these proteins in the brain.

The claudins represent one of the main research areas for Dr Nangia as they have a crucial role in the permeability of the blood-brain barrier. More specifically, they are able to form a physical fence between cells that is responsible for filtering the influx of nutrients into the brain.

The difficulties in isolating the intact tight junction strands using experimental techniques had made research progress difficult. Therefore, Dr Nangia and her colleagues developed a computational approach to get around this technical challenge. They are simulating the blood-brain barrier interface in silico (performed on a computer), from the basic claudin structure to tight junction aggregates.

This approach allowed Dr Nangia and her team to publish in 2015 their observation that two claudins can spontaneously interact to form a dimer (a process known as dimerisation). Dr Nangia’s team showed that among all possible dimers, there were four dimers that were observed frequently. The predicted dimers matched with the experimental results. Dr Nangia noted that out of the four dimers, two dimers could form pores to allow the passage of small molecules in and out, contributing to the tight junction permeability. Intrigued, Dr Nangia and her team measured the stability of each dimer by calculating the force needed to separate the two claudins, and found that each dimer had different stability.

They then went a step further and proposed that the lipid composition of the cell membrane might also play a role in the dimer interaction. The results showed that interactions between two claudins are driven by the lipid environment of the cell membrane. This discovery has far-reaching consequences because it provides important insight into the transport properties of the tight junctions and into the physiological complexity of the blood-brain barrier itself. Furthermore, the relevance of these findings extends beyond the blood-brain barrier as claudins are a large family of proteins that establish tight junctions in numerous other interfaces throughout the body.

An Additional Signal to Influence Claudin Interactions

It has been well established that after they are expressed in a cell, proteins undergo a series of transformations known as ‘post-translational modifications’. These modifications slightly change the protein to influence either their function or their dynamic. For example, they can act as signals to guide the final protein to its designated location.

Previous studies have demonstrated that various post-translational modifications of claudin can directly influence tight junction architecture, notably through the addition of a single palmitic acid molecule (the basic component of palm oil) – already known to play a role in tissue permeability regulation. This process is referred to scientifically as palmitoylation, with the precise function being dependent upon the proteins involved.

In this context, Dr Nangia and her colleagues reported in 2019 that claudin palmitoylation can influence the capacity of claudins to form a dimer and as well as the dimer stability. This completed another important piece of the puzzle, as they came to believe on the basis of these findings that palmitoylation has a significant impact on the permeability of tight junctions.

This finding is again applicable to many other interfaces, as explained by Dr Nangia, ‘overall, this study contributes to the growing body of research focused on understanding the significance of post-translational lipid modification of proteins in cellular and subcellular membranes and its impact on critical cellular functions.’

PANEL: A Novel Approach to Study Protein-Protein Interactions

Computational approaches are often associated with having several limitations, including being highly time-consuming and expensive. In 2019, Dr Nangia and her team published details of their recently developed accessible, robust, and affordable approach to study protein interactions: PANEL, which stands for Protein AssociatioN Energy Landscape.

PANEL is based on calculating the non-bonded interaction energies between two proteins, which depend on the position and the orientation of the two proteins relative to each other in the cell membrane. The PANEL approach computes this energy for all the possible protein-protein orientations and ranks their stability. Dr Nangia further explains that the PANEL method can generate a comprehensive dataset for any interacting membrane protein – which is at least 100 times faster than other available methods. Indeed, Dr Nangia and her colleagues have used the PANEL plot to generate a comprehensive sampling of dimer conformations and provide a clear visualisation of the entire energy landscape demonstrating the dimer interaction energies.

Progressively, Dr Nangia and her colleagues are diving deeper into the tight junctions of the blood-brain barrier and they now have a clear idea of the claudin interactions necessary for maximal permeability. It is important, however, to remember that tight junctions provide dynamic interfaces that undergo continual change, and that many other physiological factors contribute to the final architecture.

Limitations and Perspectives

Using computational tools, Dr Nangia and her team have explored the depth of the blood-brain barrier, bringing fundamental findings to the scientific community. To put these findings in perspective, approximately 40 other families of proteins are known to be involved in the tight junction architecture. Though they do not directly contribute to the selectivity of the tight junctions, their structural function suggests that they also interact with claudins to strengthen the backbone of the blood-brain barrier.

Furthermore, several other studies have reported claudin aggregation into groups known as trimers, tetramers, and even hexamers, according to the number and arrangement of the protein subunits. This highlights the high level of complexity of the tight junctions in their regulation of the permeability of the blood-brain barrier.

Although many questions are yet to be answered, Dr Nangia’s approach and the PANEL method will enable future research to focus on developing strategies to facilitate drug delivery into the brain. The next critical step forward is now to identify small molecules that could be used to modulate tight junctions, allowing treatments to target the brain but in a safely controlled and temporary manner. As one might imagine, the pores cannot stay indefinitely opened, as this would leave the brain vulnerable to damage through exposure to innumerable toxins.

Ultimately, modulating the blood-brain barrier and controlling its permeability will allow drugs to reach the brain via the bloodstream. This is an unchartered territory that will give rise to new challenges such as drug safety for the brain and side effects, but one that offers enormous potential for progressing treatment for neurodegenerative diseases. Dr Nangia and her team are making giant strides forward towards making such much-needed treatments a real possibility in the coming years.