Hydroxyapatite (HAP, Ca 10 (PO 4 ) 6 (OH) 2 ) nanoparticles with controlled materials properties ... more Hydroxyapatite (HAP, Ca 10 (PO 4 ) 6 (OH) 2 ) nanoparticles with controlled materials properties have been synthesized through a two-step hydrothermal aging method to investigate fibronectin (Fn) adsorption. Two distinct populations of HAP nanoparticles have been generated: HAP1 particles had smaller size, plate-like shape, lower crystallinity, and more negative ζ potential than HAP2 particles. We then developed two-dimensional platforms containing HAP and Fn and analyzed both the amount and the conformation of Fn via Forster resonance energy transfer (FRET) at various HAP concentrations. Our FRET analysis reveals that larger amounts of more compact Fn molecules were adsorbed onto HAP1 than onto HAP2 particles. Additionally, our data show that the amount of compact Fn adsorbed increased with increasing HAP concentration due to the formation of nanoparticle agglomerates. We propose that both the surface chemistry of single nanoparticles and the size and morphology of HAP agglomerates play significant roles in the interaction of Fn with HAP. Collectively, our findings suggest that the HAP-induced conformational changes of Fn, a critical mechanotransducer protein involved in the communication of cells with their environment, will ultimately affect downstream cellular behaviors. These results have important implications for our understanding of organic−inorganic interactions in physiological and pathological biomineralization processes such as HAP-related inflammation.
Fibronectin (Fn) forms a fibrillar network that controls cell behavior in both physiological and ... more Fibronectin (Fn) forms a fibrillar network that controls cell behavior in both physiological and diseased conditions including cancer. Indeed, breast cancer-associated stromal cells not only increase the quantity of deposited Fn but also modify its conformation. However, (i) the interplay between mechanical and conformational properties of early tumor-associated Fn networks and (ii) its effect on tumor vascularization remain unclear. Here, we first used the Surface Forces Apparatus to reveal that 3T3-L1 preadipocytes exposed to tumor-secreted factors generate a stiffer Fn matrix relative to control cells. We then show that this early matrix stiffening correlates with increased molecular unfolding in Fn fibers, as determined by Förster Resonance Energy Transfer. Finally, we assessed the resulting changes in adhesion and proangiogenic factor (VEGF) secretion of newly seeded 3T3-L1s, and we examined altered integrin specificity as a potential mechanism of modified cell–matrix interactions through integrin blockers. Our data indicate that tumor-conditioned Fn decreases adhesion while enhancing VEGF secretion by preadipocytes, and that an integrin switch is responsible for such changes. Collectively, our findings suggest that simultaneous stiffening and unfolding of initially deposited tumor-conditioned Fn alters both adhesion and proangiogenic behavior of surrounding stromal cells, likely promoting vascularization and growth of the breast tumor. This work enhances our knowledge of cell – Fn matrix interactions that may be exploited for other biomaterials-based applications, including advanced tissue engineering approaches.
Fibronectin (FN) is a glycoprotein found in the superficial
zone of cartilage; however, its role ... more Fibronectin (FN) is a glycoprotein found in the superficial zone of cartilage; however, its role in the lubrication and the wear protection of articular joints is unknown. In this work, we have investigated the molecular interactions between FN and various components of the synovial fluid such as lubricin (LUB), hyaluronan (HA), and serum albumin (SA), which are all believed to contribute to joint lubrication. Using a Surface Forces Apparatus, we have measured the normal (adhesion/repulsion) and lateral (friction) forces across layers of individual synovial fluid components physisorbed onto FN-coated mica substrates. Our chief findings are (i) FN strongly tethers LUB and HA to mica, as indicated by high and reversible long-range repulsive normal interactions between surfaces, and (ii) FN and LUB synergistically enhance wear protection of surfaces during shear, as suggested by the structural robustness of FN+LUB layers under pressures up to about 4 MPa. These findings provide new insights into the role of FN in the lubricating properties of synovial fluid components sheared between ideal substrates and represent a significant step forward in our understanding of cartilage damage involved in diseases such as osteoarthritis.
Pre-adipocytes exposed to tumor soluble factors derived from MDA-MB-231 breast cancer cells, an i... more Pre-adipocytes exposed to tumor soluble factors derived from MDA-MB-231 breast cancer cells, an in vitro model of cancer-activated fibroblasts, deposited an initial extracellular matrix comprised primarily of fibronectin (Fn). This tumorassociated Fn was measured to be 87% more unfolded than that of Fn deposited by control cells and correlated to being 62% stiffer by the Förster Resonance Energy Transfer and Surface Forces Apparatus techniques, respectively. The initial dysregulated ECM lead to the rapid deposition of thick type I collagen (Col I) fibers. Our findings suggest that initial unfolded and stiff tumor-associated Fn ECMs promote altered cell-matrix interactions to fuel tumor progression.
Hydroxyapatite (HAP, Ca 10 (PO 4 ) 6 (OH) 2 ) nanoparticles with controlled materials properties ... more Hydroxyapatite (HAP, Ca 10 (PO 4 ) 6 (OH) 2 ) nanoparticles with controlled materials properties have been synthesized through a two-step hydrothermal aging method to investigate fibronectin (Fn) adsorption. Two distinct populations of HAP nanoparticles have been generated: HAP1 particles had smaller size, plate-like shape, lower crystallinity, and more negative ζ potential than HAP2 particles. We then developed two-dimensional platforms containing HAP and Fn and analyzed both the amount and the conformation of Fn via Forster resonance energy transfer (FRET) at various HAP concentrations. Our FRET analysis reveals that larger amounts of more compact Fn molecules were adsorbed onto HAP1 than onto HAP2 particles. Additionally, our data show that the amount of compact Fn adsorbed increased with increasing HAP concentration due to the formation of nanoparticle agglomerates. We propose that both the surface chemistry of single nanoparticles and the size and morphology of HAP agglomerates play significant roles in the interaction of Fn with HAP. Collectively, our findings suggest that the HAP-induced conformational changes of Fn, a critical mechanotransducer protein involved in the communication of cells with their environment, will ultimately affect downstream cellular behaviors. These results have important implications for our understanding of organic−inorganic interactions in physiological and pathological biomineralization processes such as HAP-related inflammation.
Fibronectin (Fn) forms a fibrillar network that controls cell behavior in both physiological and ... more Fibronectin (Fn) forms a fibrillar network that controls cell behavior in both physiological and diseased conditions including cancer. Indeed, breast cancer-associated stromal cells not only increase the quantity of deposited Fn but also modify its conformation. However, (i) the interplay between mechanical and conformational properties of early tumor-associated Fn networks and (ii) its effect on tumor vascularization remain unclear. Here, we first used the Surface Forces Apparatus to reveal that 3T3-L1 preadipocytes exposed to tumor-secreted factors generate a stiffer Fn matrix relative to control cells. We then show that this early matrix stiffening correlates with increased molecular unfolding in Fn fibers, as determined by Förster Resonance Energy Transfer. Finally, we assessed the resulting changes in adhesion and proangiogenic factor (VEGF) secretion of newly seeded 3T3-L1s, and we examined altered integrin specificity as a potential mechanism of modified cell–matrix interactions through integrin blockers. Our data indicate that tumor-conditioned Fn decreases adhesion while enhancing VEGF secretion by preadipocytes, and that an integrin switch is responsible for such changes. Collectively, our findings suggest that simultaneous stiffening and unfolding of initially deposited tumor-conditioned Fn alters both adhesion and proangiogenic behavior of surrounding stromal cells, likely promoting vascularization and growth of the breast tumor. This work enhances our knowledge of cell – Fn matrix interactions that may be exploited for other biomaterials-based applications, including advanced tissue engineering approaches.
Fibronectin (FN) is a glycoprotein found in the superficial
zone of cartilage; however, its role ... more Fibronectin (FN) is a glycoprotein found in the superficial zone of cartilage; however, its role in the lubrication and the wear protection of articular joints is unknown. In this work, we have investigated the molecular interactions between FN and various components of the synovial fluid such as lubricin (LUB), hyaluronan (HA), and serum albumin (SA), which are all believed to contribute to joint lubrication. Using a Surface Forces Apparatus, we have measured the normal (adhesion/repulsion) and lateral (friction) forces across layers of individual synovial fluid components physisorbed onto FN-coated mica substrates. Our chief findings are (i) FN strongly tethers LUB and HA to mica, as indicated by high and reversible long-range repulsive normal interactions between surfaces, and (ii) FN and LUB synergistically enhance wear protection of surfaces during shear, as suggested by the structural robustness of FN+LUB layers under pressures up to about 4 MPa. These findings provide new insights into the role of FN in the lubricating properties of synovial fluid components sheared between ideal substrates and represent a significant step forward in our understanding of cartilage damage involved in diseases such as osteoarthritis.
Pre-adipocytes exposed to tumor soluble factors derived from MDA-MB-231 breast cancer cells, an i... more Pre-adipocytes exposed to tumor soluble factors derived from MDA-MB-231 breast cancer cells, an in vitro model of cancer-activated fibroblasts, deposited an initial extracellular matrix comprised primarily of fibronectin (Fn). This tumorassociated Fn was measured to be 87% more unfolded than that of Fn deposited by control cells and correlated to being 62% stiffer by the Förster Resonance Energy Transfer and Surface Forces Apparatus techniques, respectively. The initial dysregulated ECM lead to the rapid deposition of thick type I collagen (Col I) fibers. Our findings suggest that initial unfolded and stiff tumor-associated Fn ECMs promote altered cell-matrix interactions to fuel tumor progression.
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Papers by Fei Wu
zone of cartilage; however, its role in the lubrication and the wear protection
of articular joints is unknown. In this work, we have investigated the molecular interactions between FN and various components of the synovial fluid such as lubricin (LUB), hyaluronan (HA), and serum albumin (SA), which are all believed to contribute to joint lubrication. Using a Surface Forces Apparatus, we have measured the normal (adhesion/repulsion) and lateral (friction) forces across layers of individual synovial fluid components physisorbed onto FN-coated mica substrates. Our chief findings are (i) FN strongly tethers LUB and HA to mica, as indicated by high and reversible long-range repulsive normal interactions between surfaces, and (ii) FN and LUB synergistically enhance wear protection of surfaces during shear, as suggested by the structural robustness of FN+LUB layers under pressures up to about 4 MPa. These findings provide new insights into the role of FN in the lubricating properties of synovial fluid components sheared between ideal substrates and represent a significant step forward in our understanding of cartilage damage involved in diseases such as osteoarthritis.
zone of cartilage; however, its role in the lubrication and the wear protection
of articular joints is unknown. In this work, we have investigated the molecular interactions between FN and various components of the synovial fluid such as lubricin (LUB), hyaluronan (HA), and serum albumin (SA), which are all believed to contribute to joint lubrication. Using a Surface Forces Apparatus, we have measured the normal (adhesion/repulsion) and lateral (friction) forces across layers of individual synovial fluid components physisorbed onto FN-coated mica substrates. Our chief findings are (i) FN strongly tethers LUB and HA to mica, as indicated by high and reversible long-range repulsive normal interactions between surfaces, and (ii) FN and LUB synergistically enhance wear protection of surfaces during shear, as suggested by the structural robustness of FN+LUB layers under pressures up to about 4 MPa. These findings provide new insights into the role of FN in the lubricating properties of synovial fluid components sheared between ideal substrates and represent a significant step forward in our understanding of cartilage damage involved in diseases such as osteoarthritis.