Effects of Aqueous Boundary Layers and Paracellular Transport on the Efflux Ratio as a Measure of Active Transport Across Cell Layers
The efflux ratio (ER), determined by Caco-2/MDCK assays, is the standard in vitro metric to establish qualitatively whether a compound is a substrate of an efflux transporter. However, others have also enabled the utilisation of this metric quantitatively by deriving a relationship that expresses th...
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| Format: | Article |
| Language: | English |
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MDPI AG
2024-01-01
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| Series: | Pharmaceutics |
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| Online Access: | https://www.mdpi.com/1999-4923/16/1/132 |
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| author | Soné Kotze Andrea Ebert Kai-Uwe Goss |
| author_facet | Soné Kotze Andrea Ebert Kai-Uwe Goss |
| author_sort | Soné Kotze |
| collection | DOAJ |
| description | The efflux ratio (ER), determined by Caco-2/MDCK assays, is the standard in vitro metric to establish qualitatively whether a compound is a substrate of an efflux transporter. However, others have also enabled the utilisation of this metric quantitatively by deriving a relationship that expresses the ER as a function of the intrinsic membrane permeability of the membrane (<inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msub><mi>P</mi><mn>0</mn></msub></semantics></math></inline-formula>) as well as the permeability of carrier-mediated efflux (<inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msub><mi>P</mi><mrow><mi>p</mi><mi>g</mi><mi>p</mi></mrow></msub></semantics></math></inline-formula>). As of yet, <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msub><mi>P</mi><mrow><mi>p</mi><mi>g</mi><mi>p</mi></mrow></msub></semantics></math></inline-formula> cannot be measured directly from transport experiments or otherwise, but the ER relationship provides easy access to this value if <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msub><mi>P</mi><mn>0</mn></msub></semantics></math></inline-formula> is known. However, previous derivations of this relationship failed to consider the influence of additional transport resistances such as the aqueous boundary layers (ABLs) and the filter on which the monolayer is grown. Since single fluxes in either direction can be heavily affected by these experimental artefacts, it is crucial to consider the potential impact on the ER. We present a model that includes these factors and show both mathematically and experimentally that this simple ER relationship also holds for the more realistic scenario that does not neglect the ABLs/filter. Furthermore, we also show mathematically how paracellular transport affects the ER, and we experimentally confirm that paracellular dominance reduces the ER to unity and can mask potential efflux. |
| first_indexed | 2024-03-08T10:37:43Z |
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| id | doaj.art-aad6f2d8c1d947fd91a5720ae14b21f5 |
| institution | Directory Open Access Journal |
| issn | 1999-4923 |
| language | English |
| last_indexed | 2024-03-08T10:37:43Z |
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| spelling | doaj.art-aad6f2d8c1d947fd91a5720ae14b21f52024-01-26T18:08:10ZengMDPI AGPharmaceutics1999-49232024-01-0116113210.3390/pharmaceutics16010132Effects of Aqueous Boundary Layers and Paracellular Transport on the Efflux Ratio as a Measure of Active Transport Across Cell LayersSoné Kotze0Andrea Ebert1Kai-Uwe Goss2Department of Analytical Environmental Chemistry, Helmholtz Centre for Environmental Research (UFZ), Permoserstraße 15, 04318 Leipzig, GermanyDepartment of Analytical Environmental Chemistry, Helmholtz Centre for Environmental Research (UFZ), Permoserstraße 15, 04318 Leipzig, GermanyDepartment of Analytical Environmental Chemistry, Helmholtz Centre for Environmental Research (UFZ), Permoserstraße 15, 04318 Leipzig, GermanyThe efflux ratio (ER), determined by Caco-2/MDCK assays, is the standard in vitro metric to establish qualitatively whether a compound is a substrate of an efflux transporter. However, others have also enabled the utilisation of this metric quantitatively by deriving a relationship that expresses the ER as a function of the intrinsic membrane permeability of the membrane (<inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msub><mi>P</mi><mn>0</mn></msub></semantics></math></inline-formula>) as well as the permeability of carrier-mediated efflux (<inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msub><mi>P</mi><mrow><mi>p</mi><mi>g</mi><mi>p</mi></mrow></msub></semantics></math></inline-formula>). As of yet, <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msub><mi>P</mi><mrow><mi>p</mi><mi>g</mi><mi>p</mi></mrow></msub></semantics></math></inline-formula> cannot be measured directly from transport experiments or otherwise, but the ER relationship provides easy access to this value if <inline-formula><math xmlns="http://www.w3.org/1998/Math/MathML" display="inline"><semantics><msub><mi>P</mi><mn>0</mn></msub></semantics></math></inline-formula> is known. However, previous derivations of this relationship failed to consider the influence of additional transport resistances such as the aqueous boundary layers (ABLs) and the filter on which the monolayer is grown. Since single fluxes in either direction can be heavily affected by these experimental artefacts, it is crucial to consider the potential impact on the ER. We present a model that includes these factors and show both mathematically and experimentally that this simple ER relationship also holds for the more realistic scenario that does not neglect the ABLs/filter. Furthermore, we also show mathematically how paracellular transport affects the ER, and we experimentally confirm that paracellular dominance reduces the ER to unity and can mask potential efflux.https://www.mdpi.com/1999-4923/16/1/132efflux transportersMDCKCaco-2active transportP-glycoproteinpermeability |
| spellingShingle | Soné Kotze Andrea Ebert Kai-Uwe Goss Effects of Aqueous Boundary Layers and Paracellular Transport on the Efflux Ratio as a Measure of Active Transport Across Cell Layers Pharmaceutics efflux transporters MDCK Caco-2 active transport P-glycoprotein permeability |
| title | Effects of Aqueous Boundary Layers and Paracellular Transport on the Efflux Ratio as a Measure of Active Transport Across Cell Layers |
| title_full | Effects of Aqueous Boundary Layers and Paracellular Transport on the Efflux Ratio as a Measure of Active Transport Across Cell Layers |
| title_fullStr | Effects of Aqueous Boundary Layers and Paracellular Transport on the Efflux Ratio as a Measure of Active Transport Across Cell Layers |
| title_full_unstemmed | Effects of Aqueous Boundary Layers and Paracellular Transport on the Efflux Ratio as a Measure of Active Transport Across Cell Layers |
| title_short | Effects of Aqueous Boundary Layers and Paracellular Transport on the Efflux Ratio as a Measure of Active Transport Across Cell Layers |
| title_sort | effects of aqueous boundary layers and paracellular transport on the efflux ratio as a measure of active transport across cell layers |
| topic | efflux transporters MDCK Caco-2 active transport P-glycoprotein permeability |
| url | https://www.mdpi.com/1999-4923/16/1/132 |
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