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Open access to novel dual flow chamber technology for in vitro cell mechanotransduction, toxicity and pharamacokinetic studies.

Anderson EJ, Knothe Tate ML - Biomed Eng Online (2007)

Bottom Line: Bench-top testing of the novel chamber prototype shows improvements, in the ease of use as well as in performance, compared to the other commercial chambers.The predictability of the imparted stress improves both experiment repeatability as well as the accuracy of inter-study comparisons.Carefully controlling the stresses on cells is critical in effectively mimicking in vivo situations.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Mechanical & Aerospace Engineering, Case Western Reserve University, Cleveland, OH, USA. eric.anderson@case.edu

ABSTRACT

Background: A major stumbling block for researchers developing experimental models of mechanotransduction is the control of experimental variables, in particular the transmission of the mechanical forces at the cellular level. A previous evaluation of state of the art commercial perfusion chambers showed that flow regimes, applied to impart a defined mechanical stimulus to cells, are poorly controlled and that data from studies in which different chambers are utilized can not be compared, even if the target stress regimes are comparable.

Methods: This study provides a novel chamber design to provide both physiologically-based flow regimes, improvements in control of experimental variables, as well as ease of use compared to commercial chambers. This novel design achieves controlled stresses through five gasket designs and both single- and dual-flow regimes.

Results: The imparted shear stress within the gasket geometry is well controlled. Fifty percent of the entire area of the 10 x 21 mm universal gasket (Gasket I, designed to impart constant magnitude shear stresses in the center of the chamber where outcome measures are taken), is exposed to target stresses. In the 8 mm diameter circular area at the center of the chamber (where outcome measures are made), over 92% of the area is exposed to the target stress (+/- 2.5%). In addition, other gasket geometries provide specific gradients of stress that vary with distance from the chamber inlet. Bench-top testing of the novel chamber prototype shows improvements, in the ease of use as well as in performance, compared to the other commercial chambers. The design of the chamber eliminates flow deviations due to leakage and bubbles and allows actual flow profiles to better conform with those predicted in computational models.

Conclusion: The novel flow chamber design provides predictable and well defined mechanical forces at the surface of a cell monolayer, showing improvement over previously tested commercial chambers. The predictability of the imparted stress improves both experiment repeatability as well as the accuracy of inter-study comparisons. Carefully controlling the stresses on cells is critical in effectively mimicking in vivo situations. Overall, the improved perfusion flow chamber provides the needed resolution, standardization and in vitro model analogous to in vivo conditions to make the step towards greater use in research and the opportunity to enter the diagnostic and therapeutic market.

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Related in: MedlinePlus

Dual-flow profile schematic. The general schematic for apical and basal flow used in the design of the novel flow chamber, where the cell monolayer is housed between the two fluid reservoirs on either a porous membrane or solid substrate.
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Figure 1: Dual-flow profile schematic. The general schematic for apical and basal flow used in the design of the novel flow chamber, where the cell monolayer is housed between the two fluid reservoirs on either a porous membrane or solid substrate.

Mentions: To use the dual flow modality, cells seeded within and/or on one or both sides of a permeable membrane, or cells seeded in a scaffold disc, or cells in situ (in a slice of tissue), are interposed between the two gaskets and flow is applied. For example, cells are seeded on a porous membrane (0.2 micron Anapore membrane, Nunc International, Denmark) that is interposed between two gaskets. The cell monolayer is interposed between two identical fluid layers, each with an independent inlet and outlet (Figure 1). The dual configuration provides relatively even flow across both surfaces of the monolayer, where the thin membrane (thickness is chosen by the user) between the regimes suspends the specimen in a physiologic manner. However, as mentioned previously, the membrane can be interchanged with a variety of other substrates, depending on the specific needs of the study, e.g. a layer in which cells are seeded or embedded can be placed between the gaskets or a slice of tissue with cells in situ can also be interposed between the gaskets. Furthermore, single-flow can also be achieved with a solid coverslip in place of the membrane.


Open access to novel dual flow chamber technology for in vitro cell mechanotransduction, toxicity and pharamacokinetic studies.

Anderson EJ, Knothe Tate ML - Biomed Eng Online (2007)

Dual-flow profile schematic. The general schematic for apical and basal flow used in the design of the novel flow chamber, where the cell monolayer is housed between the two fluid reservoirs on either a porous membrane or solid substrate.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2235874&req=5

Figure 1: Dual-flow profile schematic. The general schematic for apical and basal flow used in the design of the novel flow chamber, where the cell monolayer is housed between the two fluid reservoirs on either a porous membrane or solid substrate.
Mentions: To use the dual flow modality, cells seeded within and/or on one or both sides of a permeable membrane, or cells seeded in a scaffold disc, or cells in situ (in a slice of tissue), are interposed between the two gaskets and flow is applied. For example, cells are seeded on a porous membrane (0.2 micron Anapore membrane, Nunc International, Denmark) that is interposed between two gaskets. The cell monolayer is interposed between two identical fluid layers, each with an independent inlet and outlet (Figure 1). The dual configuration provides relatively even flow across both surfaces of the monolayer, where the thin membrane (thickness is chosen by the user) between the regimes suspends the specimen in a physiologic manner. However, as mentioned previously, the membrane can be interchanged with a variety of other substrates, depending on the specific needs of the study, e.g. a layer in which cells are seeded or embedded can be placed between the gaskets or a slice of tissue with cells in situ can also be interposed between the gaskets. Furthermore, single-flow can also be achieved with a solid coverslip in place of the membrane.

Bottom Line: Bench-top testing of the novel chamber prototype shows improvements, in the ease of use as well as in performance, compared to the other commercial chambers.The predictability of the imparted stress improves both experiment repeatability as well as the accuracy of inter-study comparisons.Carefully controlling the stresses on cells is critical in effectively mimicking in vivo situations.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Mechanical & Aerospace Engineering, Case Western Reserve University, Cleveland, OH, USA. eric.anderson@case.edu

ABSTRACT

Background: A major stumbling block for researchers developing experimental models of mechanotransduction is the control of experimental variables, in particular the transmission of the mechanical forces at the cellular level. A previous evaluation of state of the art commercial perfusion chambers showed that flow regimes, applied to impart a defined mechanical stimulus to cells, are poorly controlled and that data from studies in which different chambers are utilized can not be compared, even if the target stress regimes are comparable.

Methods: This study provides a novel chamber design to provide both physiologically-based flow regimes, improvements in control of experimental variables, as well as ease of use compared to commercial chambers. This novel design achieves controlled stresses through five gasket designs and both single- and dual-flow regimes.

Results: The imparted shear stress within the gasket geometry is well controlled. Fifty percent of the entire area of the 10 x 21 mm universal gasket (Gasket I, designed to impart constant magnitude shear stresses in the center of the chamber where outcome measures are taken), is exposed to target stresses. In the 8 mm diameter circular area at the center of the chamber (where outcome measures are made), over 92% of the area is exposed to the target stress (+/- 2.5%). In addition, other gasket geometries provide specific gradients of stress that vary with distance from the chamber inlet. Bench-top testing of the novel chamber prototype shows improvements, in the ease of use as well as in performance, compared to the other commercial chambers. The design of the chamber eliminates flow deviations due to leakage and bubbles and allows actual flow profiles to better conform with those predicted in computational models.

Conclusion: The novel flow chamber design provides predictable and well defined mechanical forces at the surface of a cell monolayer, showing improvement over previously tested commercial chambers. The predictability of the imparted stress improves both experiment repeatability as well as the accuracy of inter-study comparisons. Carefully controlling the stresses on cells is critical in effectively mimicking in vivo situations. Overall, the improved perfusion flow chamber provides the needed resolution, standardization and in vitro model analogous to in vivo conditions to make the step towards greater use in research and the opportunity to enter the diagnostic and therapeutic market.

Show MeSH
Related in: MedlinePlus