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Quantitative analysis of 3D extracellular matrix remodelling by pancreatic stellate cells.

Robinson BK, Cortes E, Rice AJ, Sarper M, Del Río Hernández A - Biol Open (2016)

Bottom Line: We present an integrated methodology where cell-ECM interactions can be investigated in 3D environments via ECM remodelling.Pancreatic stellate cells (PSCs) are the key effectors of the stromal fibrosis associated to pancreatic cancer.We use PSCs to implement our methodology and demonstrate that PSC matrix remodelling capabilities depend on their contractile machinery and β1 integrin-mediated cell-ECM attachment.

View Article: PubMed Central - PubMed

Affiliation: Cellular and Molecular Biomechanics Laboratory, Department of Bioengineering, Faculty of Engineering, Imperial College London, South Kensington Campus, London SW7 2AZ, UK.

No MeSH data available.


Related in: MedlinePlus

Remodelling of 3D collagen-I matrices by PSCs. (A) Bright-field images of matrix remodelling, assessed by matrix contraction (left). Mean±s.e.m. percentage change due to matrix contraction per cell number (right). Acellular, n=10; 250K, n=8; 500K, n=16; 750K, n=6. *P<0.05, ***P<0.0001 (unpaired t-test). (B) Immunofluorescence of PSCs (red), fibronectin (blue) and SHG imaging of collagen-I (green) for remodelled matrices containing 250,000 and 750,000 cells (left). Scale bar: 50 μm. Mean±s.e.m. of collagen-I and fibronectin intensity density of immunofluorescence (250K, n=13; 500K, n=11; 750K, n=7) and second harmonic signal for remodelled matrices (acellular, n=10; 250K, n=25; 500K, n=10; 750K, n=29) (right). Student's t-test shows a significant difference in fibronectin intensity between 250,000 and 750,000 cell conditions (P=0.01).
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BIO017632F1: Remodelling of 3D collagen-I matrices by PSCs. (A) Bright-field images of matrix remodelling, assessed by matrix contraction (left). Mean±s.e.m. percentage change due to matrix contraction per cell number (right). Acellular, n=10; 250K, n=8; 500K, n=16; 750K, n=6. *P<0.05, ***P<0.0001 (unpaired t-test). (B) Immunofluorescence of PSCs (red), fibronectin (blue) and SHG imaging of collagen-I (green) for remodelled matrices containing 250,000 and 750,000 cells (left). Scale bar: 50 μm. Mean±s.e.m. of collagen-I and fibronectin intensity density of immunofluorescence (250K, n=13; 500K, n=11; 750K, n=7) and second harmonic signal for remodelled matrices (acellular, n=10; 250K, n=25; 500K, n=10; 750K, n=29) (right). Student's t-test shows a significant difference in fibronectin intensity between 250,000 and 750,000 cell conditions (P=0.01).

Mentions: To assess the ECM remodelling capacity of PSCs, we prepared collagen/matrigel 3D matrices containing PSCs with increasing cell concentration. Monitoring the matrix contraction by imaging matrices in 96 well plates at 24 h intervals for 72 h allowed the assessment of the relative dimensional changes under remodelling. Taking the initial and final time points of the contraction, the percentage change in the matrix area represented the contractile ability of each condition. Matrix contraction was proportional to the number of seeded cells, with the maximum contraction of 80% observed for the matrices embedded with 750,000 cells (Fig. 1A;Fig. S1A).Fig. 1.


Quantitative analysis of 3D extracellular matrix remodelling by pancreatic stellate cells.

Robinson BK, Cortes E, Rice AJ, Sarper M, Del Río Hernández A - Biol Open (2016)

Remodelling of 3D collagen-I matrices by PSCs. (A) Bright-field images of matrix remodelling, assessed by matrix contraction (left). Mean±s.e.m. percentage change due to matrix contraction per cell number (right). Acellular, n=10; 250K, n=8; 500K, n=16; 750K, n=6. *P<0.05, ***P<0.0001 (unpaired t-test). (B) Immunofluorescence of PSCs (red), fibronectin (blue) and SHG imaging of collagen-I (green) for remodelled matrices containing 250,000 and 750,000 cells (left). Scale bar: 50 μm. Mean±s.e.m. of collagen-I and fibronectin intensity density of immunofluorescence (250K, n=13; 500K, n=11; 750K, n=7) and second harmonic signal for remodelled matrices (acellular, n=10; 250K, n=25; 500K, n=10; 750K, n=29) (right). Student's t-test shows a significant difference in fibronectin intensity between 250,000 and 750,000 cell conditions (P=0.01).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4920190&req=5

BIO017632F1: Remodelling of 3D collagen-I matrices by PSCs. (A) Bright-field images of matrix remodelling, assessed by matrix contraction (left). Mean±s.e.m. percentage change due to matrix contraction per cell number (right). Acellular, n=10; 250K, n=8; 500K, n=16; 750K, n=6. *P<0.05, ***P<0.0001 (unpaired t-test). (B) Immunofluorescence of PSCs (red), fibronectin (blue) and SHG imaging of collagen-I (green) for remodelled matrices containing 250,000 and 750,000 cells (left). Scale bar: 50 μm. Mean±s.e.m. of collagen-I and fibronectin intensity density of immunofluorescence (250K, n=13; 500K, n=11; 750K, n=7) and second harmonic signal for remodelled matrices (acellular, n=10; 250K, n=25; 500K, n=10; 750K, n=29) (right). Student's t-test shows a significant difference in fibronectin intensity between 250,000 and 750,000 cell conditions (P=0.01).
Mentions: To assess the ECM remodelling capacity of PSCs, we prepared collagen/matrigel 3D matrices containing PSCs with increasing cell concentration. Monitoring the matrix contraction by imaging matrices in 96 well plates at 24 h intervals for 72 h allowed the assessment of the relative dimensional changes under remodelling. Taking the initial and final time points of the contraction, the percentage change in the matrix area represented the contractile ability of each condition. Matrix contraction was proportional to the number of seeded cells, with the maximum contraction of 80% observed for the matrices embedded with 750,000 cells (Fig. 1A;Fig. S1A).Fig. 1.

Bottom Line: We present an integrated methodology where cell-ECM interactions can be investigated in 3D environments via ECM remodelling.Pancreatic stellate cells (PSCs) are the key effectors of the stromal fibrosis associated to pancreatic cancer.We use PSCs to implement our methodology and demonstrate that PSC matrix remodelling capabilities depend on their contractile machinery and β1 integrin-mediated cell-ECM attachment.

View Article: PubMed Central - PubMed

Affiliation: Cellular and Molecular Biomechanics Laboratory, Department of Bioengineering, Faculty of Engineering, Imperial College London, South Kensington Campus, London SW7 2AZ, UK.

No MeSH data available.


Related in: MedlinePlus