The interstitial fluid (ISF) surrounds every cell in the brain, allowing it to be the main carrier for proteins trafficking to and from cells. ISF is of great significance in developing brain microvascular networks (MVN), a specialized vascular structure consisting of astrocytes, pericytes and brain endothelial cells. Brain endothelial cells contribute to forming a restrictive blood-brain barrier (BBB) between the blood and the extracellular space of the brain. The BBB-related protein and gene expression are also enhanced by the juxtacrine and paracrine signals provided by astrocytes and pericytes. The BBB is impermeable to about 100% of large-molecule drugs and about 98% of small-molecule therapeutics, and its dysfunction is a marker of diseased brain.
It is speculated that developing accurate in vitro models of human brain MVNs could provide a better understanding of the underlying MVNs mechanism of MVNs for modeling neurodegenerative diseases and innovative drug delivery techniques. Microfluidic devices are common platforms for developing in vitro models because they accurately control the process parameters. However, most of the available microfluidic devices designs for studying human BBB and cellular interactions fail to provide an accurate representation of the brain microvasculature morphology because they implement artificial membranes and endothelial cell-lined fluidic channels. Consequently, most in vitro models used in brain MVNs do not consider the impact of interstitial flow during microvessel generation. This is a critical shortcoming considering the vital role of interstitial flow in providing mechanical cues, delivering nutrients and removing waste products, among other functions.
To address these limitations, Northeastern University researchers: Mr. Max Winkelman, Dr. Diana Kim, Mr. Shravani Kakarla, Dr. Alexander Grath, Mr. Nathaniel Silvia and Professor Guohao Dai developed brain MVNs through both angiogenic and vasculogenic processes within by culturing primary human brain endothelial cells, astrocytes, and pericytes within microfluidic devices. The microvessels were cultured in a 3D fibrin matrix with interstitial flow conditions (flow) and without interstitial flow (static). The MVNs were evaluated based on different features, including their morphological characteristics, longevity, protein expression, and cellular interactions, among others. The authors predicted that the interstitial flow could enhance the formation of brain MVN, interconnectivity and the endothelium barrier function. Their work is currently published in the journal, Lab on a Chip.
The authors showed that the bulk interstitial fluid flow benefited both brain endothelial cells angiogenesis and vasculogenesis. The resulting brain MVNs exhibited favorable cellular interactions suitable for expressing BBB-related proteins. The MVNs cultured under flow conditions were perfusable and achieved enhanced vascular morphological characteristics and endothelial barrier function as well as improved longevity and connectivity. In contrast, those cultured under static conditions did not have connectivity or any perfuse ability. Although no significant changes were reported in the pericytes microvessel coverage, a slight increase in astrocytes coverage was reported under flow culture. Additionally, the immunofluorescence intensity of lamina proteins, laminin and collagen IV almost doubled in the flow culture.
In summary, the research team successfully demonstrated the importance of interstitial flow in developing human brain MVNs within microfluidic devices. The results highlighted the coupling of biological and physical cues responsible for regulating the maturation of brain microvessel. The characteristic features of the MVNs make the presented experimental design suitable for in vitro model for human brain microvasculature and can be replicated in other studies with ease. For instance, the resulting brain MVNs could be useful in determining drug efficacy for the central nervous system during preclinical studies. Furthermore, modification of the model could enable the study of neurodegenerative diseases associated with BBB dysfunction. In a statement to Medicine Innovates, Professor Guohao Dai explained their study provides a practical approach for increasing the success of realizing perfusable vasculature in microfluidic devices for various applications. In depth characterization of ISF using microfluidic device approach reported in the study are important in understanding the neurobiology of healthy and diseased brain as well as important for drug applications, disease treatment, and prevention
Winkelman, M., Kim, D., Kakarla, S., Grath, A., Silvia, N., & Dai, G. (2022). Interstitial flow enhances the formation, connectivity, and function of 3D brain microvascular networks generated within a microfluidic device. Lab on a Chip, 22(1), 170-192.