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Surface-Bound Molecular Gradients for the High-Throughput Screening of Cell Responses.

Lagunas A, Martínez E, Samitier J - Front Bioeng Biotechnol (2015)

Bottom Line: Chemical gradient surfaces are described as surfaces with a gradually varying composition along their length.Here, we review some of the most recent examples in which gradients have been used to evaluate the effect of a varying ligand concentration in cell adhesion, morphology, growth, and differentiation of cells, including some of our recent findings.They show the importance of the organization of ligands at the nanoscale, which is highlighted by abrupt changes in cell behavior at critical concentration thresholds.

View Article: PubMed Central - PubMed

Affiliation: Networking Biomedical Research Center in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN) , Madrid , Spain ; Nanobioengineering Group, Institute for Bioengineering of Catalonia (IBEC) , Barcelona , Spain.

ABSTRACT
Chemical gradient surfaces are described as surfaces with a gradually varying composition along their length. Continuous chemical gradients have recently been proposed as an alternative to discrete microarrays for the high-throughput screening of the effects of ligand concentration in cells. Here, we review some of the most recent examples in which gradients have been used to evaluate the effect of a varying ligand concentration in cell adhesion, morphology, growth, and differentiation of cells, including some of our recent findings. They show the importance of the organization of ligands at the nanoscale, which is highlighted by abrupt changes in cell behavior at critical concentration thresholds.

No MeSH data available.


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Evolution of differentiation markers in cells cultured onto BMP-2 gradients. (A) Plot of the percentage of cells in the osteogenic commitment showing ALP positive staining versus BMP-2 surface density. Non-linear effects are found below 2.1 pmol/cm2 and the percentage of cells in the osteogenic commitment reaches saturation from this value onward. Control experiments performed on streptavidin-modified gradients show differentiation values below 10%, independent of the BMP-2 dose. Dashed lines are an eye guide. (B) Differentiation of C2C12 myoblasts on BMP-2 gradients. Immuno-fluorescent imaging reveals a decrease of troponin T positive cells (undergoing myogenic differentiation) with increasing BMP-2 concentration and an increase in the ALP expression [(A): Lagunas et al., 2013; (B): Almodóvar et al., 2014].
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Figure 2: Evolution of differentiation markers in cells cultured onto BMP-2 gradients. (A) Plot of the percentage of cells in the osteogenic commitment showing ALP positive staining versus BMP-2 surface density. Non-linear effects are found below 2.1 pmol/cm2 and the percentage of cells in the osteogenic commitment reaches saturation from this value onward. Control experiments performed on streptavidin-modified gradients show differentiation values below 10%, independent of the BMP-2 dose. Dashed lines are an eye guide. (B) Differentiation of C2C12 myoblasts on BMP-2 gradients. Immuno-fluorescent imaging reveals a decrease of troponin T positive cells (undergoing myogenic differentiation) with increasing BMP-2 concentration and an increase in the ALP expression [(A): Lagunas et al., 2013; (B): Almodóvar et al., 2014].

Mentions: In our group, we took advantage of a biotin–streptavidin gradient platform to study the concentration effects of BMP-2 on C2C12 cell differentiation (Figure 2A; Lagunas et al., 2013). We fabricated gradients of biotinylated BMP-2 bound to PMMA substrates with low slope (0.9 pmol/cm3) and an overall surface density ranging 1.4–2.3 pmol/cm2. We observed a non-linear dependence of the osterix (OSX) nuclear translocation (an osteoblast-specific transcription factor) with an abrupt increase above a threshold density of 1.7 pmol/cm2. A similar behavior was seen for the expression of alkaline phosphatase (ALP) enzyme, also related to osteogenic cell commitment. We attributed such non-linear behavior to the non-even distribution of the ligand (BMP-2) on the surface at the nanometer scale: BMP-2 clustered distribution synergistically enhances the probability of rebinding events by providing a large number of adjacent binding sites in the vicinity of the receptor, thereby increasing signaling activation (Lagunas et al., 2012). Such a non-linear C2C12 cell behavior in the presence of BMP-2 has been also reported in a recent work performed by Picart and co-workers (Figure 2B; Almodóvar et al., 2014). They used microfluidics to create gradients of BMP-2 and BMP-7 growth factors on layer-by-layer films composed of poly(l-lysine) and hyaluronan. The gradients have slopes of 0.58 μg/cm3 (22.3 pmol/cm3) for BMP-2 and 1.24 μg/cm3 (25 pmol/cm3) for BMP-7, and are linear over a distance of 20 mm. The effects of the gradients on the trans-differentiation capacity of the C2C12 cells to the osteogenic lineage were assessed. This platform was used then to produce gradients of both factors in a parallel or opposite fashion and the data found suggest an additive or synergistic effect between BMP-2 and BMP-7.


Surface-Bound Molecular Gradients for the High-Throughput Screening of Cell Responses.

Lagunas A, Martínez E, Samitier J - Front Bioeng Biotechnol (2015)

Evolution of differentiation markers in cells cultured onto BMP-2 gradients. (A) Plot of the percentage of cells in the osteogenic commitment showing ALP positive staining versus BMP-2 surface density. Non-linear effects are found below 2.1 pmol/cm2 and the percentage of cells in the osteogenic commitment reaches saturation from this value onward. Control experiments performed on streptavidin-modified gradients show differentiation values below 10%, independent of the BMP-2 dose. Dashed lines are an eye guide. (B) Differentiation of C2C12 myoblasts on BMP-2 gradients. Immuno-fluorescent imaging reveals a decrease of troponin T positive cells (undergoing myogenic differentiation) with increasing BMP-2 concentration and an increase in the ALP expression [(A): Lagunas et al., 2013; (B): Almodóvar et al., 2014].
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Figure 2: Evolution of differentiation markers in cells cultured onto BMP-2 gradients. (A) Plot of the percentage of cells in the osteogenic commitment showing ALP positive staining versus BMP-2 surface density. Non-linear effects are found below 2.1 pmol/cm2 and the percentage of cells in the osteogenic commitment reaches saturation from this value onward. Control experiments performed on streptavidin-modified gradients show differentiation values below 10%, independent of the BMP-2 dose. Dashed lines are an eye guide. (B) Differentiation of C2C12 myoblasts on BMP-2 gradients. Immuno-fluorescent imaging reveals a decrease of troponin T positive cells (undergoing myogenic differentiation) with increasing BMP-2 concentration and an increase in the ALP expression [(A): Lagunas et al., 2013; (B): Almodóvar et al., 2014].
Mentions: In our group, we took advantage of a biotin–streptavidin gradient platform to study the concentration effects of BMP-2 on C2C12 cell differentiation (Figure 2A; Lagunas et al., 2013). We fabricated gradients of biotinylated BMP-2 bound to PMMA substrates with low slope (0.9 pmol/cm3) and an overall surface density ranging 1.4–2.3 pmol/cm2. We observed a non-linear dependence of the osterix (OSX) nuclear translocation (an osteoblast-specific transcription factor) with an abrupt increase above a threshold density of 1.7 pmol/cm2. A similar behavior was seen for the expression of alkaline phosphatase (ALP) enzyme, also related to osteogenic cell commitment. We attributed such non-linear behavior to the non-even distribution of the ligand (BMP-2) on the surface at the nanometer scale: BMP-2 clustered distribution synergistically enhances the probability of rebinding events by providing a large number of adjacent binding sites in the vicinity of the receptor, thereby increasing signaling activation (Lagunas et al., 2012). Such a non-linear C2C12 cell behavior in the presence of BMP-2 has been also reported in a recent work performed by Picart and co-workers (Figure 2B; Almodóvar et al., 2014). They used microfluidics to create gradients of BMP-2 and BMP-7 growth factors on layer-by-layer films composed of poly(l-lysine) and hyaluronan. The gradients have slopes of 0.58 μg/cm3 (22.3 pmol/cm3) for BMP-2 and 1.24 μg/cm3 (25 pmol/cm3) for BMP-7, and are linear over a distance of 20 mm. The effects of the gradients on the trans-differentiation capacity of the C2C12 cells to the osteogenic lineage were assessed. This platform was used then to produce gradients of both factors in a parallel or opposite fashion and the data found suggest an additive or synergistic effect between BMP-2 and BMP-7.

Bottom Line: Chemical gradient surfaces are described as surfaces with a gradually varying composition along their length.Here, we review some of the most recent examples in which gradients have been used to evaluate the effect of a varying ligand concentration in cell adhesion, morphology, growth, and differentiation of cells, including some of our recent findings.They show the importance of the organization of ligands at the nanoscale, which is highlighted by abrupt changes in cell behavior at critical concentration thresholds.

View Article: PubMed Central - PubMed

Affiliation: Networking Biomedical Research Center in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN) , Madrid , Spain ; Nanobioengineering Group, Institute for Bioengineering of Catalonia (IBEC) , Barcelona , Spain.

ABSTRACT
Chemical gradient surfaces are described as surfaces with a gradually varying composition along their length. Continuous chemical gradients have recently been proposed as an alternative to discrete microarrays for the high-throughput screening of the effects of ligand concentration in cells. Here, we review some of the most recent examples in which gradients have been used to evaluate the effect of a varying ligand concentration in cell adhesion, morphology, growth, and differentiation of cells, including some of our recent findings. They show the importance of the organization of ligands at the nanoscale, which is highlighted by abrupt changes in cell behavior at critical concentration thresholds.

No MeSH data available.


Related in: MedlinePlus