Side by side comparison between dynamic versus static models of blood–brain barrier in vitro: A permeability study

Santaguida, Stefano; Janigro, Damir; Hossain, Mohammed; Oby, Emily R.; Rapp, Edward; Cucullo, Luca

doi:10.1016/j.brainres.2006.06.027

Cited by 180 publications

(196 citation statements)

References 45 publications

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“…Cocultures involving three-cell types are highly restrictive and appear difficult to implement routinely. To date, the most complex in vitro BBB models are the dynamic in vitro models (DIV-BBB) which include vessel-like organization with astrocytes and include a flow of medium that mimics the blood flow 75,79,80,81,82,83,84,85 . When cerebral endothelial cells are exposed to a flow, the generated shear stress activates mechanotransducers at the cell surface, which modulate the expression of different genes involved in endothelial cell physiology such as cell division, differentiation, migration and apoptosis 80 .…”

Section: Molecular and Functional Characterizationmentioning

confidence: 99%

Setting-up an <em>In Vitro</em> Model of Rat Blood-brain Barrier (BBB): A Focus on BBB Impermeability and Receptor-mediated Transport

Molino¹,

Jabès²,

Lacassagne

et al. 2014

JoVE

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Citation: Molino, Y., Jabès, F., Lacassagne, E., Gaudin, N., Khrestchatisky, M. Setting-up an In Vitro Model of Rat Blood-brain Barrier (BBB): A Focus on BBB Impermeability and Receptor-mediated Transport. J. Vis. Exp. (88), e51278, doi:10.3791/51278 (2014). AbstractThe blood brain barrier (BBB) specifically regulates molecular and cellular flux between the blood and the nervous tissue. Our aim was to develop and characterize a highly reproducible rat syngeneic in vitro model of the BBB using co-cultures of primary rat brain endothelial cells (RBEC) and astrocytes to study receptors involved in transcytosis across the endothelial cell monolayer. Astrocytes were isolated by mechanical dissection following trypsin digestion and were frozen for later co-culture. RBEC were isolated from 5-week-old rat cortices. The brains were cleaned of meninges and white matter, and mechanically dissociated following enzymatic digestion. Thereafter, the tissue homogenate was centrifuged in bovine serum albumin to separate vessel fragments from nervous tissue. The vessel fragments underwent a second enzymatic digestion to free endothelial cells from their extracellular matrix. The remaining contaminating cells such as pericytes were further eliminated by plating the microvessel fragments in puromycin-containing medium. They were then passaged onto filters for co-culture with astrocytes grown on the bottom of the wells. RBEC expressed high levels of tight junction (TJ) proteins such as occludin, claudin-5 and ZO-1 with a typical localization at the cell borders. The transendothelial electrical resistance (TEER) of brain endothelial monolayers, indicating the tightness of TJs reached 300 ohm·cm 2 on average. The endothelial permeability coefficients (Pe) for lucifer yellow (LY) was highly reproducible with an average of 0.26 ± 0.11 x 10 -3 cm/min. Brain endothelial cells organized in monolayers expressed the efflux transporter P-glycoprotein (P-gp), showed a polarized transport of rhodamine 123, a ligand for P-gp, and showed specific transport of transferrin-Cy3 and DiILDL across the endothelial cell monolayer. In conclusion, we provide a protocol for setting up an in vitro BBB model that is highly reproducible due to the quality assurance methods, and that is suitable for research on BBB transporters and receptors.

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Section: Molecular and Functional Characterizationmentioning

confidence: 99%

Setting-up an <em>In Vitro</em> Model of Rat Blood-brain Barrier (BBB): A Focus on BBB Impermeability and Receptor-mediated Transport

Molino¹,

Jabès²,

Lacassagne

et al. 2014

JoVE

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“…In conjunction with TEER monitoring, depletion of the main carbohydrate component of the growth medium (glucose) and accumulation of metabolically produced lactic acid were used as indicators of cell growth and the assessment of a viable in vitro BBB (Cucullo et al, 2007;Santaguida et al, 2006). Both the luminal and abluminal compartments (in both DIV-BBB and Transwell) were sampled at 2-day intervals.…”

Section: Cell Metabolism: Lactate Production and Glucose Consumptionmentioning

confidence: 99%

“…Shear stress affects endothelial cell differentiation, tight junction formation, the expression of junction-related proteins (Arisaka et al, 1995;Yoshida et al, 1995), and induces mitotic arrest (Desai et al, 2002;Lin et al, 2000). An index of the endothelial 'tightness' of the monolayer is the TEER measured across the BBB (Santaguida et al, 2006). Our data clearly show that the exposure to flow, as found in vivo, allows endothelial cells cultured in the DIV-BBB flow apparatus (in the presence of abluminal glia) to achieve a TEER over 15-fold higher than in static conditions.…”

Section: Role Of Shear Stress In Bbb Development and Cell Metabolismmentioning

confidence: 99%

Immortalized Human Brain Endothelial Cells and Flow-Based Vascular Modeling: A Marriage of Convenience for Rational Neurovascular Studies

Cucullo

Couraud

Weksler

et al. 2007

J Cereb Blood Flow Metab

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232

219

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In evaluating drugs that enter or are excluded from the brain, novel pharmaceutical strategies are needed. For this reason, we have developed a humanized Dynamic In vitro Blood-Brain Barrier model (hDIV-BBB) based on a novel human brain vascular endothelial cell line (HCMEC/D3), which closely mimics the BBB in vivo. In this system, HCMEC/D3 was grown in the lumen of hollow microporous fibers and exposed to a physiological pulsatile flow. Comparison with well-established humanized DIV-BBB models (based on human brain and non-brain vascular endothelial cells co-cultured with abluminal astrocytes) demonstrated that HCMEC/D3 cells cultured under flow conditions maintain in vitro physiological permeability barrier properties of the BBB in situ even in the absence of abluminal astrocytes. Measurements of glucose metabolism demonstrated that HCMEC/D3 cells retain an aerobic metabolic pathway. Permeability to sucrose and two relevant central nervous system drugs showed that the HCMEC/D3 cells grown under dynamic conditions closely mimic the physiological permeability properties of the BBB in situ (slope = 0.93). Osmotic disruption of the BBB was also successfully achieved. Peak BBB opening in the DIV-BBB lasted from 20 to 30 mins and was completely reversible. Furthermore, the sequence of flow cessation/ reperfusion in the presence of leukocytes led to BBB failure as demonstrated by a biphasic decrease in transendothelial electrical resistance. Additionally, BBB failure was paralleled by the intraluminal release of proinflammatory factors (interleukin-6 and interleukin-1b) and matrix metalloproteinase-9 (MMP-9). Pretreatment with ibuprofen (0.125 mmol/L) prevented BBB failure by decreasing the inflammatory response after flow cessation/reperfusion.

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“…I). A possible reason resides on the velocity field and the relevant shear stress field acting on cells that are able to better mimic the local physiological brain microenvironment (50)(51)(52)(53)(54)(55). The morphology of cultured cells, the cell glucose consumption, the lactate production, the transendothelial electrical resistance in HFs and other parameters, including the potassium transport and the HF permeability to drugs, were investigated.…”

Section: The Blood Brain Barriermentioning

confidence: 99%

Hollow Fiber Bioreactor Technology for Tissue Engineering Applications

et al. 2016

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Hollow fiber bioreactors are the focus of scientific research aiming to mimic physiological vascular networks and engineer organs and tissues in vitro. The reason for this lies in the interesting features of this bioreactor type, including excellent mass transport properties. Indeed, hollow fiber bioreactors allow limitations to be overcome in nutrient transport by diffusion, which is often an obstacle to engineer sizable constructs in vitro. This work reviews the existing literature relevant to hollow fiber bioreactors in organ and tissue engineering applications. To this purpose, we first classify the hollow fiber bioreactors into 2 categories: cylindrical and rectangular. For each category, we summarize their main applications both at the tissue and at the organ level, focusing on experimental models and computational studies as predictive tools for designing innovative, dynamic culture systems. Finally, we discuss future perspectives on hollow fiber bioreactors as in vitro models for tissue and organ engineering applications.

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Side by side comparison between dynamic versus static models of blood–brain barrier in vitro: A permeability study

Cited by 180 publications

References 45 publications

Setting-up an <em>In Vitro</em> Model of Rat Blood-brain Barrier (BBB): A Focus on BBB Impermeability and Receptor-mediated Transport

Setting-up an <em>In Vitro</em> Model of Rat Blood-brain Barrier (BBB): A Focus on BBB Impermeability and Receptor-mediated Transport

Immortalized Human Brain Endothelial Cells and Flow-Based Vascular Modeling: A Marriage of Convenience for Rational Neurovascular Studies

Hollow Fiber Bioreactor Technology for Tissue Engineering Applications

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