The Blood-Brain Barrier inNeurological Disease

Evan Wehr, Damir Janigro, PhD, Alex Chan, Mohammed Hossain, M.D.Janigro Lab, Lerner Research Institute Cleveland Clinic Science Internship

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Background of the Blood-Brain Barrier

• It’s located at the end of all of the brain capillaries.

• Only 1% of all biomedical materials can freely cross over a normal functioning blood-brain barrier (BBB).

• If a subject (a person) has cerebrovascular inflammation, the BBB permeability increases.

• Cerebrovascular inflammation is caused by most neurological diseases.

• It’s location can make in vivo cumbersome (as well as treatment for patients).

Brainparenchyma

Capillary

About the DIV-BBB Cartridge

• Developed in the Janigro lab.

• Has the potential to make in vitro possible (more feasible than in vivo)

• Each cartridge has a different level of strain. Strain is determined by how much the fibers (in the ECS) are stretched (0%, 51%, and 79%).

• The capillaries have holes and stretching makes less holes, yet all holes larger.

TheDIV-BBB Cartridge

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Fibers(Capillaries)

Forming the Study

There were three cartridges, with three different stretches

0% Stretch 51% Stretch 79% Stretch

Outer Surface:

Inner Surface:

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Background

• It was unknown if these cartridges would replicate the BBBs attributes.

• Astrocyte cells would be run through the lumen regions and samples would be searched for cells.

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Fibers(Capillaries)

• The cells found would be taken note of.

Note: Astrocyte cells were used for practicality.

Hypothesis

It was predicted that if astrocyte media / cells were run through the cartridge’s lumen region,the following would happen in each cartridge:

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• 0% stretch: cells will not cross over to the ECS.

• 51% stretch: cells will not cross

over to the ECS

• 79% stretch: cells will cross over to the ECS

Fibers(Capillaries)

Methodology

•Cleaned cartridges with sterile water.

•Ran astro media / cells through the lumen region of each cartridge.

•Took four samples from every port.

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Fibers(Capillaries) • Viewed all of the samples from

the ports on the 5x5 field of a hemocytometer with a microscope.

• Counted how may cells were in each sample.

Results

• The hypothesis was proven true for the 0% stretched cartridge and the 79% stretched cartridge, while in the 51% stretched cartridge, it did not hold true

• The 51% stretched cartridge allowed 2,500 cells/ml to cross over while the 79% stretched cartridge allowed 90,000 cells/ml cross over. In the 0% stretched cartridge, nothing crossed over.

• In vitro testing is possible using the DIV-BBB cartridge.

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Expanding Research

• Tubing can be attached to the lumen ports of the cartridge to allow material to be pumped through it for a long duration of time (48 hours).

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Fibers(Capillaries)

Expanding Research

• There is a TEER (Trans-Endothelial Electrical Resistance) measurement machine that can be hooked up to the electrodes on the bottom of the cartridge. The endothelial cells produce electrical resistance. The TEER machine can register how much electrical resistance there is.

• Flocel can record the data gathered by the TEER machine over a long duration of time.

• If used with the tubing set up, experiments can be run for long durations of time and data can be collected without taking samples.

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Fibers(Capillaries)

Recommendations

• Run trypan blue (a dye) through the cartridge’s lumen to see if it will cross over to the ECS, which it shouldn't. Also, with a biochemistry analyzer, take a sugar reading from the lumen and the ECS before and after loading the dye. This will determine the amount of sugar that the cells used (how hard they are “working”).

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Thank You

• I would like to thank the following for their help on this project. Without them, none of this would be possible.

• Office of Civic Education Initiatives

• Dr. Damir Janigro, PhD

• Mohammed Hossain, M.D.

• Alex Chan

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