Poster S5 - Advanced AD-BBB-on-a-Chip microfluidic device

Friday, October 25, 2024

3:30 PM – 4:30 PM EST

Location: Exhibit Hall E, F & G

Presenting Author(s)

Yi Zuo, PhD (she/her/hers)

Postdoc
Johns Hopkins University
Baltimore, Maryland, United States

Co-Author(s)

NZ

Nan Zhao

Assistant Research Scientist
Johns Hopkins University, United States
Zikai Xia (he/him/his)

Graduate student
Johns Hopkins University
Baltimore, Maryland, United States
Da Huo, MSc (he/him/his)

Graduate student
Johns Hopkins University, Maryland, United States
Jinchong Xu, MD, PhD

Assistant Professor
Johns Hopkins University, United States
PS

Peter Searson

Joseph R. and Lynn C. Reynolds Professor
Johns Hopkins University, United States

Primary Investigator(s)

LG

Luo Gu, PhD

Assistant Professor
Johns Hopkins University, United States

Introduction: The development of accurate cellular models for Alzheimer's disease (AD) remains a significant challenge due to the limitations of traditional 2D cell cultures and animal models. To address this, we have engineered an advanced 3D AD-BBB-on-a-chip microfluidic device that incorporates both AD brain organoids and a functional blood-brain barrier (BBB) model. This platform uniquely integrates a tunable viscoelastic alginate hydrogel system to replicate the mechanical properties of the AD brain, a critical aspect often overlooked in previous 3D models. By co-culturing iPSC-derived AD brain organoids with 3D BBB microvessels, our model provides a comprehensive platform for investigating AD pathogenesis. The microfluidic design allows for independent manipulation and analysis of both the brain and vascular components, facilitating a deeper understanding of their interactions and responses to potential therapeutic interventions. By recapitulating key aspects of AD pathology and the brain microenvironment, our 3D AD-BBB-on-a-chip model can help to elucidate disease mechanisms, accelerate drug discovery, and facilitate the development of more effective treatments for AD in the future.

Materials and

Methods: This study encompasses a comprehensive range of methodologies. Alginate hydrogels were synthesized and coupled with RGD, followed by purification through dialysis and lyophilization. Hydrogels were crosslinked with calcium sulfate and subjected to compression testing to determine mechanical properties.
Brain organoids were generated from iPSCs, including both wild-type and AD models. These organoids were cultured in ultra-low attachment plates, embedded in Matrigel, and maintained in a specialized medium. They were evaluated for AD-specific features after a month of culture. Human umbilical vein endothelial cells (HUVECs-GFP) were cultured in endothelial growth medium and maintained under standard conditions for use in microvessel experiments.
Microfluidic devices were fabricated from PDMS using a custom aluminum mold, integrated with brain organoids, and seeded with endothelial cells. The permeability of these microvessels was assessed using fluorescence microscopy.
Immunocytochemistry of the blood-brain barrier (BBB) involved fixing, permeabilizing, blocking, and staining samples with primary and secondary antibodies, followed by imaging with a confocal microscope. For brain organoid immunohistochemistry, samples were cryosectioned, stained for neuronal, glial markers, and Abeta, and imaged using confocal microscopy.

Results, Conclusions, and Discussions: The study utilized viscoelastic alginate hydrogels to replicate the mechanical properties of AD brain tissue. By adjusting alginate polymer molecular weight and calcium ion crosslinker concentration, the hydrogels achieved stiffness and stress relaxation similar to AD brain tissue. Compression tests confirmed Young's modulus of 2kPa for the hydrogels, closely matching AD brain tissue. The advanced AD-BBB-Chip microfluidic device successfully co-cultured brain organoids and blood-brain barrier (BBB) microvessels, maintaining stability for at least six days. Brain organoids showed neuron and astrocyte differentiation, while microvessels formed tight junctions. Permeability tests validated barrier function.
The research demonstrates that viscoelastic hydrogels provide a suitable mechanical environment for brain organoids, closely mimicking AD brain tissue. The AD-BBB-on-a-Chip platform effectively co-cultures brain organoids with BBB microvessels, maintaining cellular structure and function over time. This system allows for the study of cellular interactions and drug testing in a controlled environment, providing insights into AD pathology and potential treatments. The platform's adaptability to other neurodegenerative diseases broadens its applicability in neurological research.
This human cell-based 3D platform offers a substantial advantage over traditional 2D culture and animal models. Fast-relaxing hydrogels provide a dynamic environment that closely mimics brain tissue, significantly influencing cell behavior and differentiation. The AD-BBB- Chip device's successful integration of brain organoids and BBB microvessels provides a robust model for studying AD and potentially testing drug candidates. This platform's versatility underscores its potential in modeling various neurological conditions, enhancing the understanding of disease mechanisms, and accelerating the development of effective therapies.

Acknowledgements (Optional):