Construction of in vitro Blood-Brain Barrier (BBB) model using MDCK-1 cell line
INTRODUCTION: The BBB constitutes the major obstacle to drug delivery to the brain. To test BBB’s permeability using the paracellular, non-specific transport pathway of a newlyformulated hydrophilic particle for neuromodulating application, BBB model using the EOO (epithelial cell/absent cell/abs...
| Main Authors: | , , |
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| Format: | Conference or Workshop Item |
| Language: | English English |
| Published: |
2014
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| Subjects: | |
| Online Access: | http://irep.iium.edu.my/39613/ http://irep.iium.edu.my/39613/1/Poster_17.pdf http://irep.iium.edu.my/39613/4/39613.pdf |
| Summary: | INTRODUCTION: The BBB constitutes the major obstacle to drug delivery to the brain. To
test BBB’s permeability using the paracellular, non-specific transport pathway of a newlyformulated
hydrophilic particle for neuromodulating application, BBB model using the EOO
(epithelial cell/absent cell/absent cell) arrangement was constructed. MATERIALS AND
METHODS: Briefly, Madin-Darby Canine Kidney (MDCK) 1 cells were cultured in
Dulbecco’s Minimum Essential Medium (DMEM) complete growth media and plated at the
density of 1 x 105 on 0.4 μm PET filter of the Millipore hanging cell culture inserts
supplemented with 0.3 ml of the complete growth medium in the luminal space and 1.1 ml of
the complete growth media in the abluminal space. The inserts were placed in the 24-well
plates to replicate the parenchymal and microvascular space of the BBB. Media were
changed for every 2 alternate days, cells were allowed to resume growth and differentiate
within 4-9 days, and finally subjected to the permeation study involving the ‘sink condition’
where the complete growth media were substituted with Hanks Balance Salt Solution (HBSS)
pH 7.4 (0.2 ml in the luminal space; 1.1 ml in the abluminal space) for pre-equilibration for 1
hour. Cells were moderately shaken in the incubator shaker for 30 minutes at 100rpm, and
trans-epithelial electrical resistance (TEER) reading was measured every 30 minutes up to 3
hours. Then, cells were washed with HBSS and substituted with the complete culture media
before being stored overnight in the incubator (37ºC, 95% relative humidity, 5% CO2). The
recovery of the TEER was measured 24 hours post-treatment using similar measurement used
at the start of the experiment. RESULTS AND DISCUSSION: In this study, the TEER values
of the MDCK cells were in the range of ~600 to ~900 Ω x cm2
. CONCLUSION: Hence the
integrity of MDCK 1 BBB was successfully sustained to meet standard models. |
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