Related Experiment Video

Updated: Aug 31, 2025

Author Spotlight: Validating Cancer Therapy Responses with Desmoplastic Spheroid Models

Author Spotlight: Validating Cancer Therapy Responses with Desmoplastic Spheroid Models

Published on: September 27, 2024

Increased Stiffness Downregulates Focal Adhesion Kinase Expression in Pancreatic Cancer Cells Cultured in 3D

Nausika Betriu¹, Anna Andreeva¹, Anna Alonso¹

¹Tissue Engineering Research Laboratory, Department of Bioengineering, IQS-School of Engineering, Ramon Llull University, 08017 Barcelona, Spain.

|August 26, 2022

Abstract:

The focal adhesion kinase (FAK) is a non-receptor tyrosine kinase that participates in integrin-mediated signal transduction and contributes to different biological processes, such as cell migration, survival, proliferation and angiogenesis. Moreover, FAK can be activated by autophosphorylation at position Y397 and trigger different signaling pathways in response to increased extracellular matrix stiffness. In addition, FAK is overexpressed and/or hyperactivated in many epithelial cancers, and its expression correlates with tumor malignancy and invasion potential. One of the characteristics of solid tumors is an over deposition of ECM components, which generates a stiff microenvironment that promotes, among other features, sustained cell proliferation and survival. Researchers are, therefore, increasingly developing cell culture models to mimic the increased stiffness associated with these kinds of tumors. In the present work, we have developed a new 3D in vitro model to study the effect of matrix stiffness in pancreatic ductal adenocarcinoma (PDAC) cells as this kind of tumor is characterized by a desmoplastic stroma and an increased stiffness compared to its normal counterpart. For that, we have used a synthetic self-assembling peptide nanofiber matrix, RAD16-I, which does not suffer a significant degradation in vitro, thus allowing to maintain the same local stiffness along culture time. We show that increased matrix stiffness in synthetic 3D RAD16-I gels, but not in collagen type I scaffolds, promotes FAK downregulation at a protein level in all the cell lines analyzed. Moreover, even though it has classically been described that stiff 3D matrices promote an increase in pFAK^Y397/FAK proteins, we found that this ratio in soft and stiff RAD16-I gels is cell-type-dependent. This study highlights how cell response to increased matrix stiffness greatly depends on the nature of the matrix used for 3D culture.

Insights

This study shows that matrix stiffness affects focal adhesion kinase (FAK) levels in pancreatic cancer cells. The type of 3D matrix used significantly influences how cells respond to stiffness, impacting FAK expression and activation.

Area of Science:

Biochemistry
Cell Biology
Biomaterials Science

Background:

Focal adhesion kinase (FAK) is a key regulator in integrin signaling, influencing cell migration, survival, proliferation, and angiogenesis.
FAK activation, particularly autophosphorylation at Y397, is linked to increased extracellular matrix (ECM) stiffness and is implicated in epithelial cancers.
Solid tumors often exhibit a stiff microenvironment due to ECM deposition, promoting tumor progression.

Purpose of the Study:

To develop and utilize a novel 3D in vitro model to investigate the impact of matrix stiffness on pancreatic ductal adenocarcinoma (PDAC) cells.
To assess the effect of a non-degradable synthetic peptide nanofiber matrix (RAD16-I) on FAK expression and activation in PDAC cells under varying stiffness conditions.

Main Methods:

Development of a 3D cell culture model using synthetic self-assembling peptide nanofiber matrix (RAD16-I) with stable stiffness.
Comparison of cell responses in RAD16-I gels versus collagen type I scaffolds.
Analysis of focal adhesion kinase (FAK) protein levels and pFAK^Y397/FAK ratios in PDAC cells cultured on matrices of varying stiffness.

Main Results:

Increased matrix stiffness in 3D RAD16-I gels led to FAK protein downregulation across all analyzed cell lines, unlike in collagen type I scaffolds.
The ratio of pFAK^Y397/FAK was found to be cell-type-dependent in both soft and stiff RAD16-I gels, challenging the classical view of stiff matrices universally increasing this ratio.
Cellular response to matrix stiffness is highly dependent on the specific biomaterial used for 3D culture.

Conclusions:

The study demonstrates that the nature of the 3D matrix significantly modulates cellular responses to microenvironmental stiffness.
FAK downregulation and cell-type-specific pFAK^Y397/FAK ratios highlight the complexity of FAK signaling in response to matrix properties.
This research underscores the importance of selecting appropriate biomaterials for 3D cell culture models to accurately mimic in vivo tumor microenvironments.

Read full analysis

Keywords:

FAK PDAC RAD16-I biomechanics pancreatic ductal adenocarcinoma self-assembling peptides stiffness

More Related Videos

A 3D Spheroid Model as a More Physiological System for Cancer-Associated Fibroblasts Differentiation and Invasion In Vitro Studies

A 3D Spheroid Model as a More Physiological System for Cancer-Associated Fibroblasts Differentiation and Invasion In Vitro Studies

Published on: August 8, 2019

Polyacrylamide Gels for Invadopodia and Traction Force Assays on Cancer Cells

Polyacrylamide Gels for Invadopodia and Traction Force Assays on Cancer Cells

Published on: January 4, 2015

Related Concept Videos

Intracellular Signaling Affects Focal Adhesions

Intracellular Signaling Affects Focal Adhesions

Integrins act both as extracellular input receivers and as intracellular processing activators. As their name suggests, integrins are entirely integrated into the membrane structure. Their hydrophobic membrane-spanning regions interact with the phospholipid bilayer's hydrophobic region. These membrane receptors provide extracellular attachment sites for effectors like hormones and growth factors. They activate intracellular response cascades when their effectors are bound and active.
Some...

Cell-matrix's Response to Mechanical Forces

Cell-matrix's Response to Mechanical Forces

In animal cells, the extracellular matrix allows cells within tissues to withstand external stresses and transmits signals from the outside of the cell to the inside. The extracellular matrix is extensive, and its composition varies between different types of tissues. For example, the reticular fibers and ground substance make up the ECM in loose connective tissue, while collagen and bone minerals make up the ECM of bone tissue.
Anchoring junctions mechanically attach a cell to the...

Articles linked to this work by shared authors, journal, and citation graph.

Knockouts of Sulfur Metabolism Genes Induce Chronic Inflammation and Immune Dysregulation in Drosophila melanogaster.

Antioxidants (Basel, Switzerland)·2026

Expression of the peanut diacylglycerol acyltransferase 3 increases the neutral lipid content and improves the fatty acid composition of Chlorella vulgaris.

Frontiers in plant science·2026

Mining of QTLs for Spring Bread Wheat Spike Productivity by Comparing Spring Wheat Cultivars Released in Different Decades of the Last Century.

Plants (Basel, Switzerland)·2024

DJ-1 protects proteins from acylation by catalyzing the hydrolysis of highly reactive cyclic 3-phosphoglyceric anhydride.

Nature communications·2024

Study of the Polysaccharide Production by the Microalga Vischeria punctata in Relation to Cultivation Conditions.

Life (Basel, Switzerland)·2022

Study of the Physicochemical and Biological Properties of the Lipid Complex of Marine Microalgae Isolated from the Coastal Areas of the Eastern Water Area of the Baltic Sea.

Molecules (Basel, Switzerland)·2022

See all related articles