scholarly journals Integrin adhesion response to chemical and mechanical stimulation

2017 ◽  
Author(s):  
◽  
Huang Huang
Keyword(s):  
Mechanical Properties ◽  
Cell Adhesion ◽  
Mechanical Stimulation ◽  
Focal Adhesions ◽  
Mechanical Stretch ◽  
Cell Stiffness ◽  
Integrin Activation ◽  
Adhesion Behavior ◽  
Failure Point ◽  
Inside Out

It is well recognized that arterial stiffness increases with aging and aging-related diseases, such as hypertension. The mechanisms for the increase in stiffness have been largely thought to be the result of changes in the composition and structure of the extracellular matrix (ECM). However, recent evidence suggests that intrinsic mechanical properties of vascular smooth muscle cells (VSMCs) may also play an important role. The changes noted in VSMCs include an increase in cell stiffness and enhanced cell adhesion to the ECM protein fibronectin (FN). The stimuli that provoke these changes are not well known, nor are the underlying causes of these changes. In addition, previous work from our laboratory revealed that there is coordination between cell stiffness and cell adhesion to FN of VSMCs treated with vasoactive agents. VSMCs adhesion to ECM is largely mediated by the transmembrane receptors, integrins, which provide a physical connection between the cytoskeleton and ECM proteins. This unique molecular axis allows integrins to act as an ideal transducer for initiating signaling from both outside-in and inside-out signaling pathways. Integrin-mediated cell adhesion is known to play an important role in VSMCs normal function and it is also involved in various pathological conditions. Despite the growing body of evidence for the importance of integrins in vascular function and dysfunction, there are gaps in our knowledge concerning how integrin adhesion is linked to changes in VSMC mechanical properties and how integrin adhesions respond to dynamic mechanical stimulation. Therefore, my overall research goal was to better understand integrin adhesion behavior in VSMCs response to cellular and mechanical stimuli. Atomic force microscopy (AFM) was used to measure VSMC mechanical properties and adhesion to ECM as well as to provide a tool for applying mechanical stimulation to the VSMC. The first part of this research focuses on clarifying the mechanism of coordination between VSMC stiffness and adhesion to FN. We hypothesized that enhanced cell adhesion to FN is mediated by changes in the level of intracellular calcium ([Ca2+]i). To test this hypothesis, confocal imaging of fluo-4, a fluorescent calcium indicator, combined with AFM force spectroscopy were used simultaneously to record levels of[Ca2+]i and force-distance curves to measure VSMC mechanical properties and adhesion. The cell mechanical properties and adhesion to FN were correlated with levels of [Ca2+]i. KCl and BAPTA-AM were used to modulate the level of [Ca2+]i. KCl-treated VSMCs showed a rapid transient increase in cell stiffness as well as cell adhesion to FN, and these two events were synchronized with the superimposed transient increase in the level of [Ca2+]i. In contrast, VSMCs incubated with an intracellular calcium chelator, BAPTAAM, exhibited decrease in stiffness and cell adhesion to FN as well as reduced levels of [Ca2+]i. These findings suggest that in VSMCs integrin activation is linked to the level of [Ca2+]i. Further studies with ML-7pretreated cells to inhibit myosin light chain kinase showed KCl induced changes were not abolished, suggesting that calcium-induced integrin activation is not dependent on mechanical events associated with contraction or signaling events downstream of contraction. In the second part of my research integrin adhesion behavior was studied in VSMC focal adhesions subjected to oscillating mechanical stimulation. VSMCs from the aorta, a large elastic conduit artery, exposed to cyclic strain stress induced by heart rate-associated changes in pulse pressure, were selected for study. We worked together with applied mathematician scientists from the University of Nottingham. Through collaborative discussions, they developed a mathematical model to predict interactions between integrins and ECM during dynamic changes in mechanical stretch. In this study, my goal was to provide biological data to test and inform the model. We used the AFM with FN-coated probes and measured VSMC adhesion to the FN by applying vertically oscillating stretch to integrin focal adhesions. Our experimental data provided evidence to support model predictions that changes in the degree of mechanical stretch applied to an integrin adhesion would behave in a bistable manner. The bistability was manifest as a breakpoint or failure point at which integrin adhesions rupture and reform. The simulation model and experimental data indicate that the bistable behavior occurs during intermediate amplitude stretches between full detachment and no detachment. The data also indicated that the failure point for adhesion was dependent on the initial conditions of the adhesion and influenced by whether the adhesion was pre-existing or newly formed. These data suggest this bistability behavior could be an indication of a unique switch point in the nature of integrin signaling. In conclusion, this research has provided new information on integrin adhesion in response to inside-out cellular stimulation and outside-in dynamic mechanical stimulation. These data indicate the involvement of a calcium-related signaling pathway in VSMC integrin activation. In addition, these data show unique integrin adhesion behavior in response to a dynamic vs static physical environment. It is clear from this work that further studies will be needed to develop a whole picture and to understand the functional and pathological implications of mechanisms coordinating integrin adhesion with cell mechanical properties and the dynamic behavior of integrins.

Long Range Conformational Transmission Of Integrin Activation and Signaling By α1/α1’-Helix Unbending At The Junction

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 198-198
Author(s):  
Jieqing Zhu ◽  
Chuanmei Zhang ◽  
Jiafu Liu ◽  
Xiuli Jiang ◽  
Nada Haydar ◽  
...  
Keyword(s):  
Cell Adhesion ◽  
Ligand Binding ◽  
Focal Adhesion ◽  
Helical Conformation ◽  
Integrin Activation ◽  
High Affinity ◽  
Affinity Ligand ◽  
Inside Out ◽  
High Affinity Ligand ◽  
Αiibβ3 Integrin

Abstract Platelet specific integrin αIIbβ3 plays an essential role in hemostasis and thrombosis. It has been used as a prototype for understanding integrin activation and conformational regulation. Crystal structures of αIIbβ3 headpiece composed of the αIIb β-propeller and β3 βI, hybrid, and PSI domains in the absence or presence of RGD-mimetic drugs revealed the headpiece changing from a closed to an open conformation upon ligand binding. A striking change is the swing-out motion of the β3 hybrid domain away from the βI and the αIIb thigh domains. This is accompanied by the changing of the α1/α1'-helix from a bent to a merged and straightened α-helical conformation. The α1/α1'-helix is bent at the α1/α1' junction (β3 Ile-131 to Gly-135) as revealed by the crystal structures of β3, β1, β2, and β7 integrins in the closed headpiece conformation. The β3 Gly-135 at the α1/α1' junction is completely conserved among all the β integrins. We propose that the conserved glycine at the α1/α1' junction is critical for maintaining the bent α1/α1'-helix conformation, and the α1/α1'-helix unbending is required for integrin activation and bidirectional signaling. To test this hypothesis, we mutated the β3 Gly-135 to alanine and showed that the β3-G135A mutation rendered αIIbβ3 integrin constitutively binding the activation-dependent mAb PAC-1. In contrast, the β3-G135P mutation had minor effect on integrin activation. This is consistent with the idea that alanine tends to stabilize a straight α-helical structure, while glycine and proline tend to introduce a bend or kink into the α-helical conformation when present at the internal positions of an α-helix. That is, the conserved β3 Gly-135 is essential for restraining the α1/α1'-helix in the bent conformation. The β3 Gly-135 is partially exposed in the bent conformation of α1/α1'-helix and buried deeply into the hydrophobic environment upon the α1/α1'-helix unbending. We rationalized that the hydrophilic substitutions will restrain, while the hydrophobic substitutions will facilitate the burying of β3 Gly-135, and thus block and induce α1/α1'-helix unbending, respectively. As expected, the β3-G135R and G135K mutations completely blocked PAC-1 binding to αIIbβ3 integrin stimulated by Mn2+ or by the αIIb-R995D mutation that mimics integrin inside-out activation. In sharp contrast, the β3-G135L and G135M mutations constitutively induced PAC-1 binding to αIIbβ3 integrin. To further confirm the α1/α1'-helix unbending is required for integrin activation and signaling, we introduced tandem double or triple glycine substitutions into the α1/α1' junction to reinforce the bent conformation of α1/α1'-helix. Remarkably, all the double or triple glycine mutations completely abolished soluble PAC-1 binding stimulated by Mn2+ from outside or by the αIIb-R995D or αIIb-F993A mutation from inside the cell. This data provide compelling evidence that the integrin α1/α1'-helix unbending is indispensible for high affinity ligand binding. Interestingly, the β3-G135R or double glycine mutant still mediated cell adhesion to immobilized PAC-1 or fibrinogen, but at a reduced level. The cell adhesion could be blocked by eptifibatide, indicating the binding ability of the mutant integrins with the high affinity small molecule ligand. However, eptifibatide failed to induce the ectodomain extension of the mutant integrins. In addition, integrin-mediated outside-in signaling, such as cell spreading, focal adhesion and F-actin stress fiber formation, and focal adhesion kinase activation was inhibited by the β3-G135R or double glycine mutations. This data demonstrated that the conformational communication initiated by ligand binding is interrupted due to the defect of α1/α1'-helix unbending. We further showed that overexpression of talin1 head domain failed to induce PAC-1 binding to the αIIbβ3 integrin with double glycine mutations at the α1/α1' junction, but still induced integrin ectodomain extension. That is, in the inside-out integrin activation, the ectodomain extension alone does not result in high affinity ligand binding. The conformational signal has to be relayed to the ligand binding site through α1/α1'-helix unbending. In conclusion, our data established the structural role of the α1/α1' junction that allows relaxation of the α1/α1'-helix in the resting state and transmission of bidirectional conformational signals by helix unbending upon integrin activation. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Sticking around: Optimal cell adhesion patterning for energy minimization and substrate mechanosensing

2020 ◽  
Author(s):  
Josephine Solowiej-Wedderburn ◽  
Carina M. Dunlop
Keyword(s):  
Mechanical Properties ◽  
Focal Adhesion ◽  
Cell Model ◽  
Stress Component ◽  
Focal Adhesions ◽  
Cell Stiffness ◽  
Adhesion Sites ◽  
Cell Shapes ◽  
A Cell ◽  
Sense And Respond

AbstractCell mechanotransduction, in which cells sense and respond to the physical properties of their micro-environments, is proving fundamental to understanding cellular behaviours across biology. Tissue stiffness (Young’s modulus) is typically regarded as the key control parameter and bioengineered gels with defined mechanical properties have become an essential part of the toolkit for interrogating mechanotransduction. We here, however, show using a mechanical cell model that the effective substrate stiffness experienced by a cell depends not just on the engineered mechanical properties of the substrate but critically also on the particular arrangement of adhesions between cell and substrate. In particular, we find that cells with different adhesion patterns can experience two different gel stiffnesses as equivalent and will generate the same mean cell deformations. For small adhesive patches, which mimic experimentally observed focal adhesions, we demonstrate that the observed dynamics of adhesion growth and elongation can be explained by energy considerations. Significantly we show different focal adhesions dynamics for soft and stiff substrates with focal adhesion growth not preferred on soft substrates consistent with reported dynamics. Equally, fewer and larger adhesions are predicted to be preferred over more and smaller, an effect enhanced by random spot placing with the simulations predicting qualitatively realistic cell shapes in this case. The model is based on a continuum elasticity description of the cell and substrate system, with an active stress component capturing cellular contractility. This work demonstrates the necessity of considering the whole cell-substrate system, including the patterning of adhesion, when investigating cell stiffness sensing, with implications for mechanotransductive control in biophysics and tissue engineering.Author summaryCells are now known to sense the mechanical properties of their tissue micro-environments and use this as a signal to control a range of behaviours. Experimentally, such cell mechanotransduction is mostly investigated using carefully engineered gel substrates with defined stiffness. Here we show, using a model integrating active cellular contractility with continuum mechanics, that the way in which a cell senses its environment depends critically not just on the stiffness of the gel but also on the spatial patterning of adhesion sites. In this way, two gels of substantially different stiffnesses can be experienced by the cell as similar, if the adhesions are located differently. Exploiting this insight, we demonstrate that it is energetically favourable for small adhesions to grow and elongate on stiff substrates but that this is not the case on soft substrates. This is consistent with experimental observations that nascent adhesions only mature to stable focal adhesion (FA) sites on stiff substrates where they also grow and elongate. These focal adhesions (FAs) have been the focus of work on mechanotransduction. However, our paper demonstrates that there is a fundamental need to consider the combined cell and micro-environment system moving beyond a focus on individual FAs.

scholarly journals Nascent Adhesion Clustering: Integrin-Integrin and Integrin-Substrate Interactions

Biophysica ◽  
2022 ◽  
Vol 2 (1) ◽  
pp. 34-58
Author(s):  
Kuanpo Lin ◽  
Robert J. Asaro
Keyword(s):  
Cell Proliferation ◽  
Cell Adhesion ◽  
Cell Membrane ◽  
Focal Adhesions ◽  
Interaction Force ◽  
Integrin Activation ◽  
Substrate Rigidity ◽  
Clear Explanation ◽  
Membrane Cell ◽  
Fundamental Mechanism

Nascent adhesions (NAs) are a general precursor to the formation of focal adhesions (FAs) that provide a fundamental mechanism for cell adhesion that is, in turn, involved in cell proliferation, migration, and mechanotransduction. Nascent adhesions form when cells come into contact with substrates at all rigidities and generally involve the clustering of ligated integrins that may recruit un-ligated integrins. Nascent adhesions tend to take on characteristic sizes in the range of O(100nm–150nm) in diameter and tend to contain integrin numbers of O(20–60). The flexible, adaptable model we present provides and clear explanation of how these conserved cluster features come about. Our model is based on the interaction among ligated and un-ligated integrins that arise due to deformations that are induced in the cell membrane-cell glycocalyx and substrate system due to integrin activation and ligation. This model produces a clearly based interaction potential, and from it an explicit interaction force among integrins, that our stochastic diffusion-interaction simulations then show will produce nascent clusters with experimentally observed characteristics. Our simulations reveal effects of various key parameters related to integrin activation and ligation as well as some unexpected and previously unappreciated effects of parameters including integrin mobility and substrate rigidity. Moreover, the model’s structure is such that refinements are readily incorporated and specific suggestions are made as to what is required for further progress in understanding nascent clustering and the development of mature focal adhesions in a truly predictive manner.

scholarly journals Superior Alignment of Human iPSC-Osteoblasts Associated with Focal Adhesion Formation Stimulated by Oriented Collagen Scaffold

2021 ◽  
Vol 22 (12) ◽  
pp. 6232
Author(s):  
Ryosuke Ozasa ◽  
Aira Matsugaki ◽  
Tadaaki Matsuzaka ◽  
Takuya Ishimoto ◽  
Hui-Suk Yun ◽  
...  
Keyword(s):  
Bone Tissue ◽  
Structural Organization ◽  
Critical Role ◽  
Bone Matrix ◽  
Adhesion Formation ◽  
Focal Adhesions ◽  
Collagen Scaffold ◽  
Patient Specific ◽  
Specific Cell ◽  
Adhesion Behavior

Human-induced pluripotent stem cells (hiPSCs) can be applied in patient-specific cell therapy to regenerate lost tissue or organ function. Anisotropic control of the structural organization in the newly generated bone matrix is pivotal for functional reconstruction during bone tissue regeneration. Recently, we revealed that hiPSC-derived osteoblasts (hiPSC-Obs) exhibit preferential alignment and organize in highly ordered bone matrices along a bone-mimetic collagen scaffold, indicating their critical role in regulating the unidirectional cellular arrangement, as well as the structural organization of regenerated bone tissue. However, it remains unclear how hiPSCs exhibit the cell properties required for oriented tissue construction. The present study aimed to characterize the properties of hiPSCs-Obs and those of their focal adhesions (FAs), which mediate the structural relationship between cells and the matrix. Our in vitro anisotropic cell culture system revealed the superior adhesion behavior of hiPSC-Obs, which exhibited accelerated cell proliferation and better cell alignment along the collagen axis compared to normal human osteoblasts. Notably, the oriented collagen scaffold stimulated FA formation along the scaffold collagen orientation. This is the first report of the superior cell adhesion behavior of hiPSC-Obs associated with the promotion of FA assembly along an anisotropic scaffold. These findings suggest a promising role for hiPSCs in enabling anisotropic bone microstructural regeneration.

Influence of the mechanical stretch property of fabrics on the wear comfort of men’s suit pants

2021 ◽  
pp. 004051752110191
Author(s):  
Hiroyuki Kanai ◽  
Kentaro Ogawa ◽  
Tetsu Sasagawa ◽  
Kiyohiro Shibata
Keyword(s):  
Mechanical Properties ◽  
Pressure Measurement ◽  
Mechanical Stretch ◽  
The Body ◽  
Weft Yarn ◽  
Elongation Ratio ◽  
Weft Direction ◽  
Wear Comfort ◽  
Finishing Process ◽  
Elastic Thread

The stretch property of fabrics is one of the most important factors that provide comfort to wearers. It is expected that tension building up in the fabric can be relaxed and the garment pressure on the body can be reduced by appropriately exploiting its stretch property. Currently, the stretch property is predominantly realized using spandex. However, weaving or knitting elastic threads cannot be employed for the worsted fabric used to design men’s suits because of their effects on the mechanical properties of the fabric (e.g., embrittlement), which deteriorate with time. In this study, worsted fabric with a graded mechanical stretch property was produced, and the effect of the mechanical stretch property on comfort was verified. The mechanical stretch property is developed from the tension relaxation and fabric shrinkage along the weft yarn during the crabbing, scouring, and drying stages of the finishing process. Then, the form of the fabric is set by heating. In this study, the worsted fabric had an elongation ratio varying from 5.9% to 16.1% along the weft direction that was produced without using elastic thread. Furthermore, men’s suit pants were made from the fabrics. The effect of the stretch property on the garment comfort was verified through sensory evaluation and garment pressure measurement. The contribution of the mechanical stretch property in improving the garment comfort of men’s suit pants is discussed.

scholarly journals Serine and Threonine Phosphorylation of the Paxillin LIM Domains Regulates Paxillin Focal Adhesion Localization and Cell Adhesion to Fibronectin

1998 ◽  
Vol 9 (7) ◽  
pp. 1803-1816 ◽  
Author(s):  
Michael C. Brown ◽  
Joseph A. Perrotta ◽  
Christopher E. Turner
Keyword(s):  
Cell Adhesion ◽  
Focal Adhesion ◽  
Binding Sites ◽  
Protein Localization ◽  
Model System ◽  
Lim Domain ◽  
Amino Terminal ◽  
Lim Domains ◽  
Inside Out

We have previously shown that the LIM domains of paxillin operate as the focal adhesion (FA)-targeting motif of this protein. In the current study, we have identified the capacity of paxillin LIM2 and LIM3 to serve as binding sites for, and substrates of serine/threonine kinases. The activities of the LIM2- and LIM3-associated kinases were stimulated after adhesion of CHO.K1 cells to fibronectin; consequently, a role for LIM domain phosphorylation in regulating the subcellular localization of paxillin after adhesion to fibronectin was investigated. An avian paxillin-CHO.K1 model system was used to explore the role of paxillin phosphorylation in paxillin localization to FAs. We found that mutations of paxillin that mimicked LIM domain phosphorylation accelerated fibronectin-induced localization of paxillin to focal contacts. Further, blocking phosphorylation of the LIM domains reduced cell adhesion to fibronectin, whereas constitutive LIM domain phosphorylation significantly increased the capacity of cells to adhere to fibronectin. The potentiation of FA targeting and cell adhesion to fibronectin was specific to LIM domain phosphorylation as mutation of the amino-terminal tyrosine and serine residues of paxillin that are phosphorylated in response to fibronectin adhesion had no effect on the rate of FA localization or cell adhesion. This represents the first demonstration of the regulation of protein localization through LIM domain phosphorylation and suggests a novel mechanism of regulating LIM domain function. Additionally, these results provide the first evidence that paxillin contributes to “inside-out” integrin-mediated signal transduction.

scholarly journals The Small GTPase Rap1b: A Bidirectional Regulator of Platelet Adhesion Receptors

2012 ◽  
Vol 2012 ◽  
pp. 1-9 ◽  
Author(s):  
Gianni Francesco Guidetti ◽  
Mauro Torti
Keyword(s):  
Cell Adhesion ◽  
Signaling Pathways ◽  
Platelet Adhesion ◽  
Thrombus Formation ◽  
Small Gtpases ◽  
Small Gtpase ◽  
Complex Pattern ◽  
Adhesive Properties ◽  
Adhesion Receptors ◽  
Inside Out

Integrins and other families of cell adhesion receptors are responsible for platelet adhesion and aggregation, which are essential steps for physiological haemostasis, as well as for the development of thrombosis. The modulation of platelet adhesive properties is the result of a complex pattern of inside-out and outside-in signaling pathways, in which the members of the Rap family of small GTPases are bidirectionally involved. This paper focuses on the regulation of the main Rap GTPase expressed in circulating platelets, Rap1b, downstream of adhesion receptors, and summarizes the most recent achievements in the investigation of the function of this protein as regulator of platelet adhesion and thrombus formation.

Cell adhesion behavior of poly(ε-caprolactone)/poly( L -lactic acid) nanofibers scaffold

2016 ◽  
Vol 171 ◽  
pp. 178-181 ◽  
Author(s):  
Zeeshan Khatri ◽  
Abdul Wahab Jatoi ◽  
Farooq Ahmed ◽  
Ick-Soo Kim
Keyword(s):  
Lactic Acid ◽  
Cell Adhesion ◽  
Adhesion Behavior
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