Cells activate signalling through ligand-receptor bonds by sensing the mechanical properties of t... more Cells activate signalling through ligand-receptor bonds by sensing the mechanical properties of the surrounding extracellular matrix (ECM). Ligands, indeed, have to withstand the pulling force elicited by cell receptors through focal adhesions (FAs). On this basis, we developed functional ligands to be simply adsorbed on surfaces and constituted by a two-domain peptide: one derived from ECM proteins and available to receptors to offer biochemical cues, and another adsorbed on material to withstand the tension upon receptor engagement. Tuneable compliance of the anchoring domain of the peptide ligand was verified by single peptide analysis through molecular dynamics and adsorption measurements. We showed that the highest adsorbed peptides combined with integrin cell-binding motifs allow for the cell recognition and polarization with larger mature FA areas. On the contrary, the lowest adsorbed se- quences did not provide mechanical resistance to the integrin pulling action, leading to more rounded cells with smaller FA areas. This evidence demonstrates that cell mechanosensory can discriminate li- gands on surfaces and should be considered as a criterion in ligand design for material bioactivation.
There is a growing interest in identifying biomacromolecules such as proteins and peptides to fun... more There is a growing interest in identifying biomacromolecules such as proteins and peptides to functionalize metallic surfaces through noncovalent binding. One method for functionalizing materials without fundamentally changing their inherent structure is using biorecognition moieties. Here, we proved a general route to select a biomolecule adhesive motif for surface functionalization by comprehensively screening phage displayed peptides. In particular, we selected a genetically engineered M13 bacteriophage and a linear dodecapeptide derived from its pIII domain for recognizing gold surfaces in a specific and selective manner. In the phage context, we demonstrated the adhesive motif was capable to adsorb on gold in a preferential way with a morphological and viscoelastic signature of the adsorbed layer as evidenced by QCM-D and AFM investigations. Out of the phage context, the linear dodecapeptide is reproducibly found to adhere to the gold surface, and by quantitative SPR measurements, high affinity constants (K eq $ 10 6 M À1 , binding energy $À8 kcal/mol) were determined.
There is a growing interest in identifying biomacromolecules such as proteins and peptides to fun... more There is a growing interest in identifying biomacromolecules such as proteins and peptides to functionalize metallic surfaces through noncovalent binding. One method for functionalizing materials without fundamentally changing their inherent structure is using biorecognition moieties. Here, we proved a general route to select a biomolecule adhesive motif for surface functionalization by comprehensively screening phage displayed peptides. In particular, we selected a genetically engineered M13 bacteriophage and a linear dodecapeptide derived from its pIII domain for recognizing gold surfaces in a specific and selective manner. In the phage context, we demonstrated the adhesive motif was capable to adsorb on gold in a preferential way with a morphological and viscoelastic signature of the adsorbed layer as evidenced by QCM-D and AFM investigations. Out of the phage context, the linear dodecapeptide is reproducibly found to adhere to the gold surface, and by quantitative SPR measurements, high affinity constants (K eq $ 10 6 M À1 , binding energy $À8 kcal/mol) were determined.
Targeted therapies represent a challenge in modern medicine. In this contest, we propose a rapid ... more Targeted therapies represent a challenge in modern medicine. In this contest, we propose a rapid and reliable methodology based on Isothermal Titration Calorimetry (ITC) coupled with confluent cell layers cultured around biocompatible templating microparticles to quantify the number of overexpressing receptors on cell membrane and study the energetics of receptor-ligand binding in near-physiological conditions. In the in vitro model here proposed we used the bEnd3 cell line as brain endothelial cells to mimic the blood brain barrier (BBB) cultured on dextran microbeads ranging from 67μm to 80μm in size (Cytodex) and the primary human umbilical vein cells (HUVEC) for comparison. The revealed affinity between transferrin (Tf) and transferrin receptor (TfR) in both systems is very high, Kd values are in the order of nM. Conversely, the value of TfRs/cell reveals a 100-fold increase in the number of TfRs per bEnd3 cells compared to HUVEC cells. The presented methodology can represent a novel and helpful strategy to identify targets, to address drug design and selectively deliver therapeutics that can cross biological barriers such as the blood brain barrier.
Imaging through turbid media is a challenging topic. A liquid is considered turbid when dispersed... more Imaging through turbid media is a challenging topic. A liquid is considered turbid when dispersed particles provoke strong light scattering, thus destroying the image formation by any standard optical system. Generally, colloidal solutions belong to the class of turbid media since dispersed particles have dimensions ranging between 0.2 μm and 2 μm. However, in microfluidics, another relevant issue has to be considered in the case of flowing liquid made of a multitude of occluding objects, e.g. red blood cells (RBCs) flowing in veins. In such a case instead of severe scattering processes unpredictable phase delays occur resulting in a wavefront distortion, thus disturbing or even hindering the image formation of objects behind such obstructing layer. In fact RBCs can be considered to be thin transparent phase objects. Here we show that sharp amplitude imaging and phase-contrast mapping of cells hidden behind biological occluding objects, namely RBCs, is possible in harsh noise conditions and with a large field-of view by Multi-Look Digital Holography microscopy (ML-DH). Noteworthy, we demonstrate that ML-DH benefits from the presence of the RBCs, providing enhancement in terms of numerical resolution and noise suppression thus obtaining images whose quality is higher than the quality achievable in the case of a liquid without occlusive objects.
Ni microparticles in nonmagnetic elastic matrices have a magnetic behavior which depends on parti... more Ni microparticles in nonmagnetic elastic matrices have a magnetic behavior which depends on particles percentage, temperature, intensity of magnetizing field and the induced strain. In particular, the elastic properties of the matrix give nonconventional effects of deformation on magnetization. When a compression is applied and the magnetizing field is high, i.e., near saturation, it becomes evident the importance of magnetic particles density increment. On the other hand, if the magnetizing field is little, below 1/4 of saturation field, the effect of elastic matrix deformation on particles' orientation and the consequent change of the magnetization intensity is prominent. We named the last peculiar behavior as ``elastomagnetic'' effect and a simple model to explain it is reported. A threshold field as a function of the particles concentration is experimentally determined which gives the prominence of one or the other of the above described mechanisms. The experimental results and the theoretical model explain that even when the intrinsic magnetoelastic effect is negligible one can find magnetoelastic effects due to coexistence of phases with different geometry, elastic and/or magnetic properties.
Mastering the interaction between cells and extracellular environment is a fundamental prerequisi... more Mastering the interaction between cells and extracellular environment is a fundamental prerequisite in order to engineer functional biomaterial interfaces able to instruct cells with specific commands. Such advanced biomaterials might find relevant application in prosthesis design, tissue engineering, diagnostics and stem cell biology. Because of the highly complex, dynamic, and multifaceted context, a thorough understanding of the cell-material crosstalk has not been achieved yet; however, a variety of material features including biological cues, topography, and mechanical properties have been proved to impact the strength and the nature of the cell-material interaction, eventually affecting cell fate and functions. Although the nature of these three signals may appear very different, they are equated by their participation in the same material-cytoskeleton crosstalk pathway as they regulate cell adhesion events. In this work we present recent and relevant findings on the material-induced cell responses, with a particular emphasis on how the presentation of biochemical/biophysical signals modulates cell behavior. Finally, we summarize and discuss the literature data to draw out unifying elements concerning cell recognition of and reaction to signals displayed by material surfaces.
Hyaluronic acid (HA) is a polysaccharide widely used in biomedical applications, due to its eleva... more Hyaluronic acid (HA) is a polysaccharide widely used in biomedical applications, due to its elevated biocompatibility and the peculiar viscoelastic properties of its solutions. Although the viscoelastic behaviour of HA solutions has been extensively studied in the literature it has been often reported in the range of low frequency (1-100 Hz) and high salt concentration, whereas the main rheological peculiarities of this molecule are expected at high frequency (>100 Hz) and low salt concentration. In this work we studied the viscoelastic properties of low molecular weight HA (155 kDa) in wide range of concentrations (0.01-20 mg/ml) at low ionic strength and over an extended frequency range (0.1-1000 Hz) using both optical tweezers and conventional rheometry. Good agreement between the high frequency dynamic behaviour (optical tweezers) and the viscoelastic properties at low frequency (rheometry) was found. We also found that, in apparent contradiction with polyelectrolyte solution theory, HA solution behaves as liquid-like viscoelastic fluid (G''>G') even at concentrations higher than the entanglement concentration where a weak-gel behavior should be expected.
In designing scaffolds for tissue regeneration, the principal objective is to recapitulate extrac... more In designing scaffolds for tissue regeneration, the principal objective is to recapitulate extracellular matrix (ECM) function in a temporally coordinated and spatially organised structure. A key issue is to encode required biological signals within the scaffold so that all aspects of cell response-adhesion and migration, proliferation and phenotype choice-can be controlled. In achieving this objective nanotechnology, bottom-up design approach and solid free-form fabrication (SFF) will play key roles, along with self-assembly processes. For scaffold materials, there must be the correct balance between architectural features notably, porosity and chemical, physical and biological properties. This paper reviews the main achievements in biomaterials design and the future challenges. r
... J. Opt. A: Pure Appl. Opt. 11 (2009) 034016 (11pp) doi:10.1088/1464-4258/11/3/034016 Microrhe... more ... J. Opt. A: Pure Appl. Opt. 11 (2009) 034016 (11pp) doi:10.1088/1464-4258/11/3/034016 Microrheology of complex fluids using optical tweezers: a comparison with macrorheological measurements G Pesce1, AC De Luca1, G Rusciano1, PA Netti2,3, S Fusco2,3 and A Sasso1 ...
ABSTRACT Cancer is a complex pathology and 2D cell model are not able to reproduce tumoral morpho... more ABSTRACT Cancer is a complex pathology and 2D cell model are not able to reproduce tumoral morphology, cell-cell and cell matrix interactions. Moreover animal models do not adequately reproduce features of human tumors, drug therapeutic responses and stem cell differentiation. In vitro 3D tissue models bridges the gap between traditional cell culture and animal models. The aim of this project is the realization of tumoral heterotypic microtissue, by seeding both stromal and tumoral cells on biodegradable microscaffold. In particular dynamic co-cultures of cancer associated fibroblasts (CAF) or their control (CAF522) with pancreatic cancer cell (PT45) on gelatin microcarriers are performed in spinner flasks for 12 days during which the cells develop a tissue structure. At the same time also CAF, CAF522 and PT45 single culture are grown to investigate the differences. Cells were counted and stained according to hematoxylin/eosin and trichromic Masson procedure. Microtissues are characterized by IF and confocal microscopy. After 12 days exogenous scaffold disappeared and new collagen has synthesized. Histological analysis showed similar features between 3D tissue and human cancer tissue. Stroma can play important roles in supporting or even inducing tumorigenesis, and 3D heterotypic tumor model realized is a good candidate to study the specific roles of stroma in tumor development, progression, local invasion. Moreover our model may help to make drug development safer and more efficient and reduce research and development expenses. Additionally, they represent promising models for individualized oncologic therapy by revealing new insights into mechanisms of organogenesis and expression of malignancy.
Liquid-liquid interfaces are highly dynamic and characterized by an elevated interfacial tension ... more Liquid-liquid interfaces are highly dynamic and characterized by an elevated interfacial tension as compared to solid-liquid interfaces. Therefore, they are gaining an increasing interest as viable templates for ordered assembly of molecules and nanoparticles. However, liquid-liquid interfaces are more difficult to handle compared to solid-liquid interfaces; their intrinsic instability may affect the assembly process, especially in the case of multiple deposition. Indeed, some attempts have been made in the deposition of polymer multilayers at liquid-liquid interfaces, but with limited control over size and stability. This study reports on the preparation of an ultrastable liquid-liquid interface based on an O/W secondary miniemulsion and its possible use as a template for the self-assembly of polymeric multilayer nanocapsules. Such polymer nanocapsules are made of entirely biodegradable materials, with highly controlled size-well under 200 nm-and multi-compartment and multifunctional features enriching their field of application in drug delivery, as well as in other bionanotechnology fields.
Cells activate signalling through ligand-receptor bonds by sensing the mechanical properties of t... more Cells activate signalling through ligand-receptor bonds by sensing the mechanical properties of the surrounding extracellular matrix (ECM). Ligands, indeed, have to withstand the pulling force elicited by cell receptors through focal adhesions (FAs). On this basis, we developed functional ligands to be simply adsorbed on surfaces and constituted by a two-domain peptide: one derived from ECM proteins and available to receptors to offer biochemical cues, and another adsorbed on material to withstand the tension upon receptor engagement. Tuneable compliance of the anchoring domain of the peptide ligand was verified by single peptide analysis through molecular dynamics and adsorption measurements. We showed that the highest adsorbed peptides combined with integrin cell-binding motifs allow for the cell recognition and polarization with larger mature FA areas. On the contrary, the lowest adsorbed se- quences did not provide mechanical resistance to the integrin pulling action, leading to more rounded cells with smaller FA areas. This evidence demonstrates that cell mechanosensory can discriminate li- gands on surfaces and should be considered as a criterion in ligand design for material bioactivation.
There is a growing interest in identifying biomacromolecules such as proteins and peptides to fun... more There is a growing interest in identifying biomacromolecules such as proteins and peptides to functionalize metallic surfaces through noncovalent binding. One method for functionalizing materials without fundamentally changing their inherent structure is using biorecognition moieties. Here, we proved a general route to select a biomolecule adhesive motif for surface functionalization by comprehensively screening phage displayed peptides. In particular, we selected a genetically engineered M13 bacteriophage and a linear dodecapeptide derived from its pIII domain for recognizing gold surfaces in a specific and selective manner. In the phage context, we demonstrated the adhesive motif was capable to adsorb on gold in a preferential way with a morphological and viscoelastic signature of the adsorbed layer as evidenced by QCM-D and AFM investigations. Out of the phage context, the linear dodecapeptide is reproducibly found to adhere to the gold surface, and by quantitative SPR measurements, high affinity constants (K eq $ 10 6 M À1 , binding energy $À8 kcal/mol) were determined.
There is a growing interest in identifying biomacromolecules such as proteins and peptides to fun... more There is a growing interest in identifying biomacromolecules such as proteins and peptides to functionalize metallic surfaces through noncovalent binding. One method for functionalizing materials without fundamentally changing their inherent structure is using biorecognition moieties. Here, we proved a general route to select a biomolecule adhesive motif for surface functionalization by comprehensively screening phage displayed peptides. In particular, we selected a genetically engineered M13 bacteriophage and a linear dodecapeptide derived from its pIII domain for recognizing gold surfaces in a specific and selective manner. In the phage context, we demonstrated the adhesive motif was capable to adsorb on gold in a preferential way with a morphological and viscoelastic signature of the adsorbed layer as evidenced by QCM-D and AFM investigations. Out of the phage context, the linear dodecapeptide is reproducibly found to adhere to the gold surface, and by quantitative SPR measurements, high affinity constants (K eq $ 10 6 M À1 , binding energy $À8 kcal/mol) were determined.
Targeted therapies represent a challenge in modern medicine. In this contest, we propose a rapid ... more Targeted therapies represent a challenge in modern medicine. In this contest, we propose a rapid and reliable methodology based on Isothermal Titration Calorimetry (ITC) coupled with confluent cell layers cultured around biocompatible templating microparticles to quantify the number of overexpressing receptors on cell membrane and study the energetics of receptor-ligand binding in near-physiological conditions. In the in vitro model here proposed we used the bEnd3 cell line as brain endothelial cells to mimic the blood brain barrier (BBB) cultured on dextran microbeads ranging from 67μm to 80μm in size (Cytodex) and the primary human umbilical vein cells (HUVEC) for comparison. The revealed affinity between transferrin (Tf) and transferrin receptor (TfR) in both systems is very high, Kd values are in the order of nM. Conversely, the value of TfRs/cell reveals a 100-fold increase in the number of TfRs per bEnd3 cells compared to HUVEC cells. The presented methodology can represent a novel and helpful strategy to identify targets, to address drug design and selectively deliver therapeutics that can cross biological barriers such as the blood brain barrier.
Imaging through turbid media is a challenging topic. A liquid is considered turbid when dispersed... more Imaging through turbid media is a challenging topic. A liquid is considered turbid when dispersed particles provoke strong light scattering, thus destroying the image formation by any standard optical system. Generally, colloidal solutions belong to the class of turbid media since dispersed particles have dimensions ranging between 0.2 μm and 2 μm. However, in microfluidics, another relevant issue has to be considered in the case of flowing liquid made of a multitude of occluding objects, e.g. red blood cells (RBCs) flowing in veins. In such a case instead of severe scattering processes unpredictable phase delays occur resulting in a wavefront distortion, thus disturbing or even hindering the image formation of objects behind such obstructing layer. In fact RBCs can be considered to be thin transparent phase objects. Here we show that sharp amplitude imaging and phase-contrast mapping of cells hidden behind biological occluding objects, namely RBCs, is possible in harsh noise conditions and with a large field-of view by Multi-Look Digital Holography microscopy (ML-DH). Noteworthy, we demonstrate that ML-DH benefits from the presence of the RBCs, providing enhancement in terms of numerical resolution and noise suppression thus obtaining images whose quality is higher than the quality achievable in the case of a liquid without occlusive objects.
Ni microparticles in nonmagnetic elastic matrices have a magnetic behavior which depends on parti... more Ni microparticles in nonmagnetic elastic matrices have a magnetic behavior which depends on particles percentage, temperature, intensity of magnetizing field and the induced strain. In particular, the elastic properties of the matrix give nonconventional effects of deformation on magnetization. When a compression is applied and the magnetizing field is high, i.e., near saturation, it becomes evident the importance of magnetic particles density increment. On the other hand, if the magnetizing field is little, below 1/4 of saturation field, the effect of elastic matrix deformation on particles' orientation and the consequent change of the magnetization intensity is prominent. We named the last peculiar behavior as ``elastomagnetic'' effect and a simple model to explain it is reported. A threshold field as a function of the particles concentration is experimentally determined which gives the prominence of one or the other of the above described mechanisms. The experimental results and the theoretical model explain that even when the intrinsic magnetoelastic effect is negligible one can find magnetoelastic effects due to coexistence of phases with different geometry, elastic and/or magnetic properties.
Mastering the interaction between cells and extracellular environment is a fundamental prerequisi... more Mastering the interaction between cells and extracellular environment is a fundamental prerequisite in order to engineer functional biomaterial interfaces able to instruct cells with specific commands. Such advanced biomaterials might find relevant application in prosthesis design, tissue engineering, diagnostics and stem cell biology. Because of the highly complex, dynamic, and multifaceted context, a thorough understanding of the cell-material crosstalk has not been achieved yet; however, a variety of material features including biological cues, topography, and mechanical properties have been proved to impact the strength and the nature of the cell-material interaction, eventually affecting cell fate and functions. Although the nature of these three signals may appear very different, they are equated by their participation in the same material-cytoskeleton crosstalk pathway as they regulate cell adhesion events. In this work we present recent and relevant findings on the material-induced cell responses, with a particular emphasis on how the presentation of biochemical/biophysical signals modulates cell behavior. Finally, we summarize and discuss the literature data to draw out unifying elements concerning cell recognition of and reaction to signals displayed by material surfaces.
Hyaluronic acid (HA) is a polysaccharide widely used in biomedical applications, due to its eleva... more Hyaluronic acid (HA) is a polysaccharide widely used in biomedical applications, due to its elevated biocompatibility and the peculiar viscoelastic properties of its solutions. Although the viscoelastic behaviour of HA solutions has been extensively studied in the literature it has been often reported in the range of low frequency (1-100 Hz) and high salt concentration, whereas the main rheological peculiarities of this molecule are expected at high frequency (>100 Hz) and low salt concentration. In this work we studied the viscoelastic properties of low molecular weight HA (155 kDa) in wide range of concentrations (0.01-20 mg/ml) at low ionic strength and over an extended frequency range (0.1-1000 Hz) using both optical tweezers and conventional rheometry. Good agreement between the high frequency dynamic behaviour (optical tweezers) and the viscoelastic properties at low frequency (rheometry) was found. We also found that, in apparent contradiction with polyelectrolyte solution theory, HA solution behaves as liquid-like viscoelastic fluid (G''>G') even at concentrations higher than the entanglement concentration where a weak-gel behavior should be expected.
In designing scaffolds for tissue regeneration, the principal objective is to recapitulate extrac... more In designing scaffolds for tissue regeneration, the principal objective is to recapitulate extracellular matrix (ECM) function in a temporally coordinated and spatially organised structure. A key issue is to encode required biological signals within the scaffold so that all aspects of cell response-adhesion and migration, proliferation and phenotype choice-can be controlled. In achieving this objective nanotechnology, bottom-up design approach and solid free-form fabrication (SFF) will play key roles, along with self-assembly processes. For scaffold materials, there must be the correct balance between architectural features notably, porosity and chemical, physical and biological properties. This paper reviews the main achievements in biomaterials design and the future challenges. r
... J. Opt. A: Pure Appl. Opt. 11 (2009) 034016 (11pp) doi:10.1088/1464-4258/11/3/034016 Microrhe... more ... J. Opt. A: Pure Appl. Opt. 11 (2009) 034016 (11pp) doi:10.1088/1464-4258/11/3/034016 Microrheology of complex fluids using optical tweezers: a comparison with macrorheological measurements G Pesce1, AC De Luca1, G Rusciano1, PA Netti2,3, S Fusco2,3 and A Sasso1 ...
ABSTRACT Cancer is a complex pathology and 2D cell model are not able to reproduce tumoral morpho... more ABSTRACT Cancer is a complex pathology and 2D cell model are not able to reproduce tumoral morphology, cell-cell and cell matrix interactions. Moreover animal models do not adequately reproduce features of human tumors, drug therapeutic responses and stem cell differentiation. In vitro 3D tissue models bridges the gap between traditional cell culture and animal models. The aim of this project is the realization of tumoral heterotypic microtissue, by seeding both stromal and tumoral cells on biodegradable microscaffold. In particular dynamic co-cultures of cancer associated fibroblasts (CAF) or their control (CAF522) with pancreatic cancer cell (PT45) on gelatin microcarriers are performed in spinner flasks for 12 days during which the cells develop a tissue structure. At the same time also CAF, CAF522 and PT45 single culture are grown to investigate the differences. Cells were counted and stained according to hematoxylin/eosin and trichromic Masson procedure. Microtissues are characterized by IF and confocal microscopy. After 12 days exogenous scaffold disappeared and new collagen has synthesized. Histological analysis showed similar features between 3D tissue and human cancer tissue. Stroma can play important roles in supporting or even inducing tumorigenesis, and 3D heterotypic tumor model realized is a good candidate to study the specific roles of stroma in tumor development, progression, local invasion. Moreover our model may help to make drug development safer and more efficient and reduce research and development expenses. Additionally, they represent promising models for individualized oncologic therapy by revealing new insights into mechanisms of organogenesis and expression of malignancy.
Liquid-liquid interfaces are highly dynamic and characterized by an elevated interfacial tension ... more Liquid-liquid interfaces are highly dynamic and characterized by an elevated interfacial tension as compared to solid-liquid interfaces. Therefore, they are gaining an increasing interest as viable templates for ordered assembly of molecules and nanoparticles. However, liquid-liquid interfaces are more difficult to handle compared to solid-liquid interfaces; their intrinsic instability may affect the assembly process, especially in the case of multiple deposition. Indeed, some attempts have been made in the deposition of polymer multilayers at liquid-liquid interfaces, but with limited control over size and stability. This study reports on the preparation of an ultrastable liquid-liquid interface based on an O/W secondary miniemulsion and its possible use as a template for the self-assembly of polymeric multilayer nanocapsules. Such polymer nanocapsules are made of entirely biodegradable materials, with highly controlled size-well under 200 nm-and multi-compartment and multifunctional features enriching their field of application in drug delivery, as well as in other bionanotechnology fields.
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Papers by Paolo A Netti
surrounding extracellular matrix (ECM). Ligands, indeed, have to withstand the pulling force elicited by
cell receptors through focal adhesions (FAs). On this basis, we developed functional ligands to be simply
adsorbed on surfaces and constituted by a two-domain peptide: one derived from ECM proteins and
available to receptors to offer biochemical cues, and another adsorbed on material to withstand the
tension upon receptor engagement. Tuneable compliance of the anchoring domain of the peptide ligand
was verified by single peptide analysis through molecular dynamics and adsorption measurements. We
showed that the highest adsorbed peptides combined with integrin cell-binding motifs allow for the cell
recognition and polarization with larger mature FA areas. On the contrary, the lowest adsorbed se-
quences did not provide mechanical resistance to the integrin pulling action, leading to more rounded
cells with smaller FA areas. This evidence demonstrates that cell mechanosensory can discriminate li-
gands on surfaces and should be considered as a criterion in ligand design for material bioactivation.
surrounding extracellular matrix (ECM). Ligands, indeed, have to withstand the pulling force elicited by
cell receptors through focal adhesions (FAs). On this basis, we developed functional ligands to be simply
adsorbed on surfaces and constituted by a two-domain peptide: one derived from ECM proteins and
available to receptors to offer biochemical cues, and another adsorbed on material to withstand the
tension upon receptor engagement. Tuneable compliance of the anchoring domain of the peptide ligand
was verified by single peptide analysis through molecular dynamics and adsorption measurements. We
showed that the highest adsorbed peptides combined with integrin cell-binding motifs allow for the cell
recognition and polarization with larger mature FA areas. On the contrary, the lowest adsorbed se-
quences did not provide mechanical resistance to the integrin pulling action, leading to more rounded
cells with smaller FA areas. This evidence demonstrates that cell mechanosensory can discriminate li-
gands on surfaces and should be considered as a criterion in ligand design for material bioactivation.