DOCSLIB.ORG

Morphological Study of Cell Protrusions During Redirected Migration in Human Fibroblast Cells

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text

Load more

MORPHOLOGICAL STUDY OF CELL PROTRUSIONS DURING REDIRECTED MIGRATION IN HUMAN FIBROBLAST CELLS

Congyingzi Zhang
A Thesis

Submitted to the Graduate College of Bowling Green
State University in partial fulfillment of the requirements for the degree of

MASTER OF SCIENCE
August 2013
Committee: Dr. Carol Heckman, Advisor Dr. Roudabeh Jamasbi Dr. Peter Gorsevski ii
ABSTRACT

Carol A. Heckman, Advisor
From the perspective of cell motility mechanisms, migration patterns arise from two opposing sources which can be viewed as forces. One, called intrinsic, maintains the cell persistence. The extrinsic arises from signals (repulsive or attractive) exerted by an external stimulus. The extrinsic force is stronger than the intrinsic, since it can overcome the intrinsic force and cause the cell to change direction. The current studies were designed to determine whether these forces were associated with different protrusions. I studied human fibroblast cells that collide with a haptotactic boundary between an adhesive substrate (germanium) and a nonadhesive substrate (plastic) in a chemokinesis system. The morphologies of cells migrating on the two substrates reflected the cells’ preference for the adhesive substrate. I measured the prevalence of various protrusions during the process of cells turning away from the boundary and reorienting their direction of travel. Classes that corresponded to protrusive features were identified by extracting latent factors from a number of primary, geometric variables, and included factor 4 (filopodia), factor 5 (cell mass displacement), and factor 7 (nascent neurites). The data showed that as cells moved further and further from the boundary, they had progressively lower values of factor 5. The correlation coefficient between the values is -0.4924. Factor 4 appeared to decrease near the boundary and recover as cells migrated onto the adhesive substrate. The results suggested that reorientation caused by the extrinsic force occurs by reducing filopodia and increasing cell mass displacement. It can be suggested that the simplest iii explanation for turning would be that the cell takes directional cues from the remaining filopodia

and obtains redirecting force from displacements of cell mass (factor 5). iv
DEDICATION

To my parents, Mr. Yuwei Zhang and Mrs. Qi Zhang for their love, support, and encouragement.

v
ACKNOWLEDGMENTS

I want to express my sincere gratitude to my advisor Dr. Carol Heckman for her continuous support of my project. She is a great advisor who always helpful and patient to me, passionate and devoted to the research, cheerful and encouraging to everyone around her. She guided me through all the stages in my research and helped me writing this report for my project. I am lucky to have her as my advisor and enjoyed all the fun from my study.
I sincerely thank Dr. Roudabeh Jamasbi from the Department of Biology and Dr. Peter
Gorsevski from the Department of Geology for their tremendous help in developing research methods and suggestions in writing this report. My sincere thanks to Dr. Marilyn Cayer for her help in my experiments.
I also would like to thank my lab mates Dr. Mita Varghese, Dr. Surya Amarachintha, and Francis Bugyei for their cooperation in the lab.
Most importantly, I want to thank my family members for their love, support, and encouragement. vi
TABLE OF CONTENTS

Page
1. INTRODUCTION .............................................................................................................
1.1 Roles of cell migration......................................................................................... 1.2 Mechanisms of cell migration and the related membrane structures................... 1.3 Studies on cell redirection....................................................................................
1.3.1 Haptotaxis ............................................................................................. 1.3.2 Chemokinesis and chemotaxis..............................................................
1.3.2.1 Chemokinesis vs. Chemotaxis ...............................................
1124799

  • 9
  • 1.3.2.2 Studies on chemotactic attractants.........................................

1.3.3 Durotaxis............................................................................................... 11
1.4 Factors and shape analysis................................................................................... 11 1.5 Haptotactic cell migration re-orientation model.................................................. 15
2. MATERIALS AND METHODS....................................................................................... 17
2.1 Cells and cell culture............................................................................................ 17 2.2 Creating the boundary between two different substrate ...................................... 18 2.3 Making cell migration tracking system................................................................ 18 2.4 Additives to DMEM for chemokinesis................................................................ 19 2.5 Microscopy for track and trace ............................................................................ 19 2.6 Shape analysis of track and trace......................................................................... 20 2.7 Statistical analysis................................................................................................ 21
3. RESULTS .......................................................................................................................... 22
3.1 Cell morphology on preferred and non-preferred substrate................................. 22 vii
3.2 Cell migration on preferred and non-preferred substrate..................................... 24

3.3 Cell migration redirected at haptotactic boundary............................................... 25 3.4 Protrusions and their dependence on the distance migrated after turning ........... 29 3.5 Protrusions and their dependence on the net distance from the boundary........... 32
4. DISCUSSION.................................................................................................................... 35
4.1 Re-orientation is characterized by cell mass displacement.................................. 35 4.2 Cell does not generate extra filopodia during re-orientation ............................... 36 4.3 Further study of filopodia sensitivity distribution from angular data.................. 38
REFERENCES ...................................................................................................................... 40 viii
LIST OF FIGURES

  • Figure
  • Page

123456789
Cell migration pattern ................................................................................................ 3T3 cells crowding in the gold area of a dual substratum system .............................
58
Substrate with certain elasticity mathematically modeled as a spring....................... 13 Micrographs of 1000w cells with high and low factor values................................... 15 Haptotactic model on a culture plate ......................................................................... 16 SEM micrographs of fibroblasts migrating on germanium substrate........................ 23 SEM micrograph of fibroblasts on plastic substrate. ............................................... 24 SEM micrograph of fibroblasts growing in culture dish after 48 hours .................... 25 Fibroblasts from the germanium side reoriented at the haptotactic boundary........... 27
10 Fibroblasts from the plastic side migrated across the haptotactic boundary. .......... 28 11 Schematic diagram of track length after turning and net distance from the haptotactic boundary. ....................................................... 30

  • 12 Four factor values at cells front when the cells migrated away from the boundary
  • 31

13 Four factor values at cells rear when the cells migrated away from the boundary.... 32 14 Four factor values at cells front with net distance between cell and the haptotactic boundary ..................................................................................... 33
15 Four factor values at cells rear with net distance between cell and the haptotactic boundary ..................................................................................... 33 Fibroblasts from the germanium side reoriented at the haptotactic boundary........... 27
16 A model showing the most possible turning angles for a cell colliding with a haptotactic boundary........................................................................ 37 ix
17 A fibroblast hit the haptotactic boundary with nearly 90 degree angle ..................... 38 x
LIST OF TABLES

Table
1
Page
Definition of factors representing cell protrusions .................................................... 14

1

1. INTRODUCTION

1.1 Roles of cell migration
Cell migration is an important cell behavior orchestrating various developmental, physiological, and pathological processes. For example, during gastrulation in embryogenesis, cells migrate as sheet forming new layers in embryo so as to form specialized tissue in the new sites. Additionally, neurogenesis and neuron development are closely linked with cell migration. Neuron cells have to migrate from their birth site hundreds of cell-body distances to reach the final position within the correct cortical layers which is facilitated by continual reconstruction of the cytoskeleton network [1]. Cell migration in adults is integral to tissue repairing, wound healing, immune response, tissue differentiation. In disease processes, cell migration is playing crucial role regardless of whether the mechanisms of migration are normal or abnormal [2].

In any multicellular organism, the sustenance of system integrity is important, and a good example is found in wound healing. Once a wound is created on a surface, which is ordinarily covered by an epithelial layer, the cells will have the potential to close the gap through cell migration and cell proliferation. This process is meaningful to the maintenance of any living system, while also being exploited for research on cell migration and biomaterials applications in tissue engineering [3]. However, over-active cell migration could also be dangerous. Drivers of cell migration help to convert cancer cells to metastasizing cells [4]. In tumor metastasis, invasive tumor cells migrate through the extracellular matrix (ECM) and move into the blood vessels for proliferation and translocation, which is termed intravasation. Translocated cancer cells can again escape the blood vessels and penetrate the surrounding tissue, termed extravasation. Two proteases secreting adhesive structures, podosomes and invadopodia, are

2

related to the penetration process, while the other two actin filament structures, filopodia and lamellipodia, and actin-rich adhesive structures attaching to the extracellular matrix are required for cell migration [5].

In early studies in cell migration, various cell lines and single cell microorganisms were studied by a typical locomotion parameter, speed. For example, neutrophils migrated at 10 µm per minute, polymorphnuclear leukocytes migrated at 3 – 30 µm per minute, macrophages at 2 µm per minute, astrocytes at 0.1 – 0.2 µm per minute, and fibroblasts at 0.08 – 1 µm per minute [6-10].

To further understand the cell migration processes and study its mechanisms, systems with various environmental cues were frequently used. These cues are categorized by their physical and chemical properties: haptotaxis, chemotaxis and chemokinesis, durotaxis, and so on. Haptotaxis refers to cells’ relative preference for different substrates. Chemotaxis and chemokinesis are both related to motogenic factors but differ by the presence or absence of a concentration gradient. Durotaxis focuses on the substrate rigidity (Young’s modulus) and its effect on cell migration.

1.2 Mechanisms of cell migration and the related membrane structures
The initial requirement for the migration process is a spatial asymmetry which could be induced by applying microscopic nonuniformities including in material composition of the substratum, temporal stimulus gradients, or directional signals [11]. Distinct front and rear regions could be identified on polarized cells during migration because of the shape of the cells. Two of the protrusions at the cell leading edge, filopodia and lamellipodia, are the results of

3

dynamic actin assemblies underneath the cell membrane although they differ morphologically in actin assembly mechanisms [12].

The thin, needle-like filopodia are filled with tight, parallel bundles of filamentous (F)- actin, which could be identified in cell types such as fibroblasts, keratinocytes, and migrating neuron growth cones. The dynamics of filopodia growth are a balance between the actin extension at the fast-growing barbed end and the retraction by the retrograde flow of the actin bundle [13, 14]. The polymerization of actin bundle in filopodia is regulated by Cdc42, a Rho subfamily GTPase, which activates IRSp53 (Insulin Receptor Substrate p53) causing it to bind Mena (mammalian Ena/VASP (Enabled/Vasodilator-Stimulated Phophoprotein)) [15]. As a key regulator of filopodia induction, directional migration and cell cycle progress, Cdc42 loss causes defects in filopodia formation, adhesion to fibronectin, wound-healing, polarization, and response to stimulus gradients[16]. The most acknowledged function of filopodia is its sensory role working as antennae to explore the ECM or other environment cues [17]. In addition, it has been known to explore for the target cell to fuse the epithelial sheets during dorsal closure and wound repair [17-19].

The other protruding membrane structure on the cell leading edge is the sheet-like lamellipodia regulated by Rac1, also a Rho subfamily GTPase, which stimulates actin network polymerization forming membrane ruffles [20]. The lamellipodium actin network disassembles about 1-3 um behind the leading edge, and the lamella provides the scaffold for actomyosin integrating with substrate adhesion producing the force for migration [21].

Another key structure regulating the cell’s morphology, focal adhesions (FA) are the adhesive interactions between a cell and the ECM. On the outside of the cell, the transmembrane

4

receptors for ECM (integrins) interact with ECM ligands for stability. This is called “ligation.” On the inside of the cell, it forms clusters which integrate with the actin filaments to form bundles called stress fibers. These fibers or cables provide the contractile force for migration and anchorage to the substrate [22, 23]. Signaled by Rho, also a Rho subfamily GTPase, the assembly, maintenance, and dissolution of FAs is mediated through the phosphorylation of the myosin II light chain facilitated by Rho kinase p160 Rho-associated kinase (ROCK). The dominant mutant of ROCK has been found to cause defects in the formation of FAs, and an selective inhibitor of the kinase suppressed FA activity in smooth-muscle cells [24, 25]. Notably, the fine, linear contact sites associated with filopodia and the small, punctate ones with the lamellipodia are the precursors of the prominent FAs at the lamellipodium-lamellum interface [22]. It is widely accepted that activated Rac, often associated with ruffling and lamellipodia, initiates new punctate contacts at the cell front while Rho matures and transits the nascent contacts into elongated cell-matrix adhesions [26-28]. In what is known as the basic cell motility cycle, protrusions formed at the leading edge set up the orientation of the polarity, and then, the contact sites are laid down on the substrate underneath. The contraction and detachment orchestrated in specific regions by the actomyosin and the integrin dynamics cause a cell to move without directional bias or, if chemotactic factors are present, up the stimulus gradient. However, the machinery of how environmental factors reorient or regulate the cell migration is not fully understood [29].

1.3 Studies on cell redirection
In the studies of mammalian cell migration behavior, cell trajectories have been segmented and categorized into two alternative modes. In mode I, the directional-mode, cells

5

migrate with more directional persistency at a higher speed. In comparison, in mode II, the reorientation mode, cells migrate with a change in directions at a lower speed. This bimodal paradigm provided a great entry point for our understanding of the different behaviors in cell migration and the study of the correlative activities (Figure 1) [30].

Figure 1. Cell migration pattern falls between two extreme particle migration modes, ballistic and diffusive. Experimental mean squared displacement versus time for the MCF-10A human mammary epithelial cells expressing the plasmid vector alone (pbabe), the normal (neuN) or transforming (neuT) versions of the rat Her 2/Neu oncogene [30]. The neuN gene is commonly amplified in human breast cancer. Reproduced from [30] by permission of Springer Press.

Among all the studies on aspects of cell reorientation or redirection, researchers have focused on indexes to describe cell behavior during migration. Linear cell locomotion velocity and directional persistence were frequently used in such studies [31, 32]. For example, in studies of embryonic chick dorsal root ganglion growth cone turning, the growth cones were reorientated by the localized treatment of myosin 1c which caused lamellipodial expansion in a certain range. The instantaneous velocity (as mean speed) and time between significant direction changes (as directional persistence parameter) were measured and fit into the Persistence

6

Random Walk (PRW) model [33]. One of the problems of these models may be distinguishing between the velocity-dependence and pattern-dependent variations in motility.

In addition to mean speed V and persistence time P, the cell polarization is another essential parameter revealing the instantaneous orientation which is highly related to the directional persistency. For example, the motility of mouse 3T3 and sarcoma cells was observed and compared on substrates of different rigidity and topography. In this study, the cell polarization was defined by the ratio of short to long cell axes. A threshold value (0.75) for the ratio was defined to categorize the polarized and non-polarized cells. The durotactic effect (see Durotaxis) from different substrates was evaluated and compared by the counts of the polarized and the non-polarized cells showing increasing substrate rigidity influenced 3T3 cell polarization and it exhibited a persistent type of migration. However, sarcoma cell spreading was more modified by the substrate topography and it migration seemed to escape from ECM cues [34].

In the studies of migration persistence, turning angle is another parameter relied upon in models of random walk migration and biased random walk migration. In Wang’s study, growth cone movement in random directions has an average turning angle near zero, which shows the conservation in turning [33]. Arrieumerlou and Meyer observed primary dendritic cells, fibroblasts, and neutrophils changed direction by turning their leading edge. In a biased random walk triggered by C5a, global cell orientation was shown as a result of small discrete turns towards the chemoattractant led by local extension of a lamellipodium in the left and right leading edge [35]. In the case with no external cue, Li et al. found that amoeboid cells from Dictyostelium discoideum migrated in a zig-zag manner and turned every 1 - 2 minutes on average. This is a result of the reorientation between characteristic ranges of –π/4 to π/4 within 2

7

- 3 minutes. The behavior of cell remembering its last turn and consistently turning away from it was considered as a strategy seeking for potential guidance cue or target [36].

Within all these quantitative cell migration models, researchers examine how cells respond to the various stimulants being applied to manipulate or bias their pattern of migration. In the following section, I review several types of models and the conclusions from those studies that inspired me. This is where I began my research on the topic of cell migration and graphical and statistical methods to understand how cells use protrusions and how they contribute to the process of migration.

1.3.1 Haptotaxis

In previous work of S. B. Carter [37], a gradient of palladium coating on a cellulose acetate surface was applied to determine if it had an effect on cell motility. Compared with the evenly coated surface, fibroblasts on the gradient showed a more uniform migration with longer tracks, and they moved towards the upper end of the gradient. Thus, the cells became dense at the upper gradient end. Carter used ‘haptotaxis’ to describe this phenomenon and suggested that the relative strength difference among peripheral adhesions contributed to directed migration and arrangement of cell shape. Later, Carter separately studied different sites on leading and trailing margins of the cell and pointed out the passive nature of cell mass, i.e. migration is not initiated by any force inside the cell but depends on the substrate on which cell is sitting [38]. In Letourneau’s study on sensory ganglia from chicken embryo, the axonal growth cone also showed relative preference for various substrata including petri dishes, palladium, dishes coated for tissue culture, and polyornithine dishes. The author showed that the direction of the elongating growth cones could be guided by the patterning of the culturing surface [39]. Similar

8

dual substratum systems were made to guide cell motility using electron microscope grids as stencils and evaporating metal (palladium or gold) evenly on other substrata, i.e. agarose and cellulose acetate [40, 41]. Making observations on 3T3 cells (Figure 2) [42], Albrecht-Buehler found that filopodia served as a rapid scanner over the environment and led the lamellipodia to spread behind it on its preferred substratum. The author’s explanation of this phenomenon was that the firmness of attachment between filopodia and the substratum dictated the direction. After filopodia retraction, only the filopodia still attached could lead the spreading of the lamellipodium. Directed cell migration introduced by a haptotactic system was considered as a result of uneven competition among different parts of lamellipodia in the cell periphery [43] which was mainly associated with the filopodia activities. Clearly, filopodia can sense the more adhesive substrate and set the direction to the favorable substrate. However the role of lamellipodium is not clear. In the history of the field, the lamellipodium and especially the ruffling membrane is the main motility organ [44].

Recommended publications
  • Development of Microfluidic Devices to Study Algal Chemotaxis and Long-Term Growth Dynamics" (2016)

    Development of Microfluidic Devices to Study Algal Chemotaxis and Long-Term Growth Dynamics" (2016)

    Louisiana State University LSU Digital Commons LSU Master's Theses Graduate School 2016 Development of Microfluidic evD ices to Study Algal Chemotaxis and Long-Term Growth Dynamics Benjamin Seth Roberts Louisiana State University and Agricultural and Mechanical College, [email protected] Follow this and additional works at: https://digitalcommons.lsu.edu/gradschool_theses Part of the Chemical Engineering Commons Recommended Citation Roberts, Benjamin Seth, "Development of Microfluidic Devices to Study Algal Chemotaxis and Long-Term Growth Dynamics" (2016). LSU Master's Theses. 4496. https://digitalcommons.lsu.edu/gradschool_theses/4496 This Thesis is brought to you for free and open access by the Graduate School at LSU Digital Commons. It has been accepted for inclusion in LSU Master's Theses by an authorized graduate school editor of LSU Digital Commons. For more information, please contact [email protected]. DEVELOPMENT OF MICROFLUIDIC DEVICES TO STUDY ALGAL CHEMOTAXIS AND LONG-TERM GROWTH DYNAMICS A Thesis Submitted to the Graduate Faculty of the Louisiana State University and Agricultural and Mechanical College in partial fulfillment of the requirements for the degree of Master of Science in The Cain Department of Chemical Engineering by Benjamin S. Roberts B.S., Mississippi State University, 2014 December 2016 TABLE OF CONTENTS ABSTRACT ................................................................................................................................... iii CHAPTER 1. INTRODUCTION ....................................................................................................1
  • Negative Durotaxis: Cell Movement Toward Softer Environments

    Negative Durotaxis: Cell Movement Toward Softer Environments

    bioRxiv preprint doi: https://doi.org/10.1101/2020.10.27.357178; this version posted October 27, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. Negative durotaxis: cell movement toward softer environments Aleksi Isomursu1,*, Keun-Young Park2,*, Jay Hou3,*, Bo Cheng4,5,*, Ghaidan Shamsan3, Benjamin Fuller3, Jesse Kasim3, M. Mohsen Mahmoodi2, Tian Jian Lu6,7, Guy M. Genin4,5,8, Feng Xu4,5, Min Lin4,5,#, Mark Distefano2,#, Johanna Ivaska1,9,#, and David J. Odde3,# 1Turku Bioscience Centre, University of Turku and Åbo Akademi University, 20520 Turku, Finland 2Department of Chemistry, University of Minnesota, Minneapolis 55455, MN, USA 3Department of Biomedical Engineering, University of Minnesota, Minneapolis 55455, MN, USA 4The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an 710049, P.R. China 5Bioinspired Engineering and Biomechanics Center (BEBC), Xi’an Jiaotong University, Xi’an 710049, P.R. China 6State Key Laboratory of Mechanics and Control of Mechanical Structures, Nanjing University of Aeronautics and Astronautics, NanjinG 210016, P.R. China 7MOE Key Laboratory of Multifunctional Materials and Structures, Xi’an JiaotonG University, Xi’an 710049, P.R. China 8NSF Science and TechnoloGy Center for EnGineerinG MechanobioloGy, WashinGton University in St. Louis, St. Louis 63130, MO, USA 9Department of Biochemistry, University of Turku, 20520 Turku, Finland *Equal contribution #Corresponding authors Email: [email protected] (D.J.O.); [email protected] (J.I.); [email protected] (M.D.); [email protected] (M.L.) Abstract: Durotaxis – the ability of cells to sense and migrate along stiffness gradients – is important for embryonic development and has been implicated in pathologies including fibrosis and cancer.
  • Bimodal Rheotactic Behavior Reflects Flagellar Beat Asymmetry in Human Sperm Cells

    Bimodal Rheotactic Behavior Reflects Flagellar Beat Asymmetry in Human Sperm Cells

    Bimodal rheotactic behavior reflects flagellar beat asymmetry in human sperm cells Anton Bukatina,b,1, Igor Kukhtevichb,c,1, Norbert Stoopd,1, Jörn Dunkeld,2, and Vasily Kantslere aSt. Petersburg Academic University, St. Petersburg 194021, Russia; bInstitute for Analytical Instrumentation of the Russian Academy of Sciences, St. Petersburg 198095, Russia; cITMO University, St. Petersburg 197101, Russia; dDepartment of Mathematics, Massachusetts Institute of Technology, Cambridge, MA 02139-4307; and eDepartment of Physics, University of Warwick, Coventry CV4 7AL, United Kingdom Edited by Charles S. Peskin, New York University, New York, NY, and approved November 9, 2015 (received for review July 30, 2015) Rheotaxis, the directed response to fluid velocity gradients, has whether this effect is of mechanical (20) or hydrodynamic (21, been shown to facilitate stable upstream swimming of mamma- 22) origin. Experiments (23) show that the alga’s reorientation lian sperm cells along solid surfaces, suggesting a robust physical dynamics can lead to localization in shear flow (24, 25), with mechanism for long-distance navigation during fertilization. How- potentially profound implications in marine ecology. In contrast ever, the dynamics by which a human sperm orients itself relative to taxis in multiflagellate organisms (2, 5, 18, 26, 27), the navi- to an ambient flow is poorly understood. Here, we combine micro- gation strategies of uniflagellate cells are less well understood. fluidic experiments with mathematical modeling and 3D flagellar beat For instance, it was discovered only recently that uniflagellate reconstruction to quantify the response of individual sperm cells in marine bacteria, such as Vibrio alginolyticus and Pseudoalteromonas time-varying flow fields. Single-cell tracking reveals two kinematically haloplanktis, use a buckling instability in their lone flagellum to distinct swimming states that entail opposite turning behaviors under change their swimming direction (28).
  • A Hybrid Computational Model for Collective Cell Durotaxis

    A Hybrid Computational Model for Collective Cell Durotaxis

    Biomechanics and Modeling in Mechanobiology https://doi.org/10.1007/s10237-018-1010-2 ORIGINAL PAPER A hybrid computational model for collective cell durotaxis Jorge Escribano1 · Raimon Sunyer2,5 · María Teresa Sánchez3 · Xavier Trepat2,4,5,6 · Pere Roca-Cusachs2,4 · José Manuel García-Aznar1 Received: 13 September 2017 / Accepted: 17 February 2018 © Springer-Verlag GmbH Germany, part of Springer Nature 2018 Abstract Collective cell migration is regulated by a complex set of mechanical interactions and cellular mechanisms. Collective migration emerges from mechanisms occurring at single cell level, involving processes like contraction, polymerization and depolymerization, of cell–cell interactions and of cell–substrate adhesion. Here, we present a computational framework which simulates the dynamics of this emergent behavior conditioned by substrates with stiffness gradients. The computational model reproduces the cell’s ability to move toward the stiffer part of the substrate, process known as durotaxis. It combines the continuous formulation of truss elements and a particle-based approach to simulate the dynamics of cell–matrix adhesions and cell–cell interactions. Using this hybrid approach, researchers can quickly create a quantitative model to understand the regulatory role of different mechanical conditions on the dynamics of collective cell migration. Our model shows that durotaxis occurs due to the ability of cells to deform the substrate more in the part of lower stiffness than in the stiffer part. This effect explains why cell collective movement is more effective than single cell movement in stiffness gradient conditions. In addition, we numerically evaluate how gradient stiffness properties, cell monolayer size and force transmission between cells and extracellular matrix are crucial in regulating durotaxis.
  • Engineering 3D Systems with Tunable Mechanical Properties to Mimic the Tumor Microenvironment Shalini Raj Unnikandam Veettil Iowa State University

    Engineering 3D Systems with Tunable Mechanical Properties to Mimic the Tumor Microenvironment Shalini Raj Unnikandam Veettil Iowa State University

    Iowa State University Capstones, Theses and Graduate Theses and Dissertations Dissertations 2018 Engineering 3D systems with tunable mechanical properties to mimic the tumor microenvironment Shalini Raj Unnikandam Veettil Iowa State University Follow this and additional works at: https://lib.dr.iastate.edu/etd Part of the Chemical Engineering Commons Recommended Citation Unnikandam Veettil, Shalini Raj, "Engineering 3D systems with tunable mechanical properties to mimic the tumor microenvironment" (2018). Graduate Theses and Dissertations. 17339. https://lib.dr.iastate.edu/etd/17339 This Thesis is brought to you for free and open access by the Iowa State University Capstones, Theses and Dissertations at Iowa State University Digital Repository. It has been accepted for inclusion in Graduate Theses and Dissertations by an authorized administrator of Iowa State University Digital Repository. For more information, please contact [email protected]. Engineering 3D systems with tunable mechanical properties to mimic the tumor microenvironment by Shalini Raj Unnikandam Veettil A thesis submitted to the graduate faculty in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Major: Chemical Engineering Program of Study Committee: Ian C Schneider, Major Professor Kaitlin Bratlie Michael Bartlett The student author, whose presentation of the scholarship herein was approved by the program of study committee, is solely responsible for the content of this thesis. The Graduate College will ensure this thesis is globally accessible and will not permit alterations after a degree is conferred. Iowa State University Ames, Iowa 2018 Copyright © Shalini Raj Unnikandam Veettil, 2018. All rights reserved. ii DEDICATION This thesis is dedicated to my family and friends who have been a great source of support.
  • Phenomenlogical Modeling of Durotaxis

    Phenomenlogical Modeling of Durotaxis

    Phenomenlogical Modeling of Durotaxis Guangyuan Yu1;2, Jingchen Feng2, Haoran Man1,Herbert Levine2;3 1 Physics and Astronomy Department, Rice University, Houston, USA 2 Center for Theoretical Biological Physics, Rice University, Houston, USA 3 Bioengineering Department, Rice University, Houston, USA (Dated: February 23, 2017) Cells exhibit qualitatively different behaviors on substrates with different rigidities. The fact that cells are more polarized on the stiffer substrate motivates us to construct a two-dimensional cell with the distribution of focal adhesions dependent on substrate rigidities. This distribution affects the forces exerted by the cell and thereby determines its motion. Our model reproduces the experimental observation that the persistence time is higher on the stiffer substrate. This stiffness dependent persistence will lead to durotaxis, the preference in moving towards stiffer substrates. This propensity is characterized by the durotaxis index first defined in experiments. We derive and validate a 2D corresponding Fokker-Planck equation associated with our model. Our approach highlights the possible role of the focal adhesion arrangement in durotaxis. Cells are capable of sensing and responding to the me- sense a stiffer substrate, they take on a more elongated chanical properties of their external environment. For shape [25, 26] as a response. Now, cells move by protru- example, cytoskeletal stiffness [1], cellular differentia- sion which occurs with the help of focal adhesions which tion [2–5] and cell morphology and motility [6–8] are all allow force transmission to the substrate. We will assume strongly influenced by ECM stiffness. In particular, it that the change in shape to being more polarized implies has been shown experimentally that cells prefer crawl- that focal adhesions (FA) are formed within a narrower ing towards the stiffer parts on substrates with spatially wedge on the cell front.
  • Hipocrate 460BC–370 BC

    Hipocrate 460BC–370 BC

    L31 - Motility or Morbidity in Neurosurgery Motto: „I move, therefore I am” (Haruki Murakami, Japanese Writer) Alexandru Vlad Ciurea1,2 MD., PhD., MSc., Dr. h.c. Mult. Prof. Alexandru Vlad Ciurea – e-mail: [email protected] Balneo Research Journal DOI: http://dx.doi.org/10.12680/balneo.2019.276 R Vol.10, No.3, September 2019 p: 353-354 1: Department of Neurosurgery, Carol Davila University School of Medicine, Bucharest, Romania. 2: Department of Neurosurgery, Sanador Medical Center Hospital, Bucharest, Romania. Abstract Background : „The lack of motion is a basic process in the birth of diseases”. (Hipocrate 460BC–370 BC) Biological motility is the capacity of an organism to move spontaneously and voluntary at the expense of energy and represents one of the crucial traits (if not the most important trait) of all living matter. Motility is a key process present in unicellular organisms as well as multicellular organisms and is the basis of embryonic development, would healing, immune response, tumor formation, metastasis, cell migration, digestion, circulation, respiration and reasoning and various other critical processes including biological evolution itself. The constant motility of developing organisms (from trilobites to human beings) has undoubtedly shaped evolution by constantly improving the functions of the skeletal system and other internal organs such as the digestive tract, liver, kidneys etc. The evolution from prehistoric human beings to modern men went through several steps during which human motility represented once more a crucial element. Once industrial food production appeared – with the overproduction we are familiarized with, human motility became a protection factor against obesity which is one of the most problematic medical conditions of our time.
  • Vascular Smooth Muscle Cell Durotaxis Depends on Extracellular Matrix Composition

    Vascular Smooth Muscle Cell Durotaxis Depends on Extracellular Matrix Composition

    Vascular smooth muscle cell durotaxis depends on extracellular matrix composition Christopher D. Hartmana, Brett C. Isenberga, Samantha G. Chuaa, and Joyce Y. Wonga,1 aDepartment of Biomedical Engineering, Boston University, Boston, MA 02215 Edited by Robert Langer, Massachusetts Institute of Technology, Cambridge, MA, and approved August 9, 2016 (received for review July 14, 2016) Mechanical compliance has been demonstrated to be a key de- by an increase in collagen I concentration (21), and in breast cancer terminant of cell behavior, directing processes such as spreading, an increase in stiffness from the tumor core to the periphery is migration, and differentiation. Durotaxis, directional migration from associated with increased levels of collagen I and laminin (12). In softer to more stiff regions of a substrate, has been observed for a atherosclerosis, a disease characterized by the thickening of the variety of cell types. Recent stiffness mapping experiments have intimal region of the arterial wall, changes in the mechanics and shown that local changes in tissue stiffness in disease are often composition of the intimal matrix occur in conjunction with the accompanied by an altered ECM composition in vivo. However, accumulation of smooth muscle and inflammatory cells (22–24). the importance of ECM composition in durotaxis has not yet been Stiffness mapping experiments have shown that plaque stiffness is explored. To address this question, we have developed and charac- spatially heterogeneous, and that these changes can be histo- terized a polyacrylamide hydrogel culture platform featuring highly logically related to the ECM composition of the plaque (20, 25). tunable gradients in mechanical stiffness. This feature, together with Given the increasing number of examples for which changes in the ability to control ECM composition, allows us to isolate the effects ECM composition in disease are coupled to changes in me- of mechanical and biological signals on cell migratory behavior.
  • The Role of Cell-Substrate Interactions in ECM Remodeling, Migration, and The

    The Role of Cell-Substrate Interactions in ECM Remodeling, Migration, and The

    The Role of Cell-Substrate Interactions in ECM Remodeling, Migration, and the Formation of Multicellular Structures Dissertation Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By James William Reinhardt B.E. Graduate Program in Biomedical Engineering The Ohio State University 2014 Dissertation Committee: Dr. Keith Gooch, Advisor Dr. Richard Hart Dr. Samir Ghadiali Dr. Peter Anderson ©Copyright by James William Reinhardt 2014 Abstract Active, mechanical interactions between cells and their extracellular matrix (ECM) are essential for ECM remodeling and cell migration, two behaviors that support diverse biological processes including embryonic development, wound healing, fibrosis, and cancer progression. Cell-populated reconstituted type I collagen hydrogels are often used as a model system in which to study ECM remodeling and cell migration in vitro. Unfortunately, this system has limitations since it is not possible to independently control individual microstructural properties. This limitation has inspired theoretical models as an alternative way to study ECM remodeling and migration. However, so far no single approach has been able to capture both fibril-level detail and dynamic cell traction force. With the goal of creating a model that can capture both fibril-level detail and dynamic cell traction force, we developed an agent-based model of cell-mediated collagen compaction and migration. With this model we observed behaviors that were not programmed, but emerged from simple rules for cell-fibril interactions. Among these, our model qualitatively reproduced remodeling commonly seen in cell-populated collagen gels: macroscopic, pericellular, and intercellular compaction. Similar to experimental observations, matrical tracks formed between pairs of cells before directional migration ii of nearby cells toward on another.
  • Topotaxis of Active Brownian Particles

    Topotaxis of Active Brownian Particles

    Topotaxis of active Brownian particles Koen Schakenraad,1, 2 Linda Ravazzano,1, 3 Niladri Sarkar,1 Joeri A.J. Wondergem,4 Roeland M.H. Merks,2, 5 and Luca Giomi1, ∗ 1Instituut-Lorentz, Leiden University, P.O. Box 9506, 2300 RA Leiden, The Netherlands 2Mathematical Institute, Leiden University, P.O. Box 9512, 2300 RA Leiden, The Netherlands 3Center for Complexity and Biosystems, Department of Physics, University of Milan, Via Celoria 16, 20133, Milano, Italy 4Kamerlingh Onnes-Huygens Laboratory, Leiden University, P.O. Box 9504, 2300 RA Leiden, The Netherlands 5Institute of Biology, Leiden University, P.O. Box 9505, 2300 RA Leiden, The Netherlands Recent experimental studies have demonstrated that cellular motion can be directed by topograph- ical gradients, such as those resulting from spatial variations in the features of a micropatterned substrate. This phenomenon, known as topotaxis, is especially prominent among cells persistently crawling within a spatially varying distribution of cell-sized obstacles. In this article we introduce a toy model of topotaxis based on active Brownian particles constrained to move in a lattice of obsta- cles, with space-dependent lattice spacing. Using numerical simulations and analytical arguments, we demonstrate that topographical gradients introduce a spatial modulation of the particles' per- sistence, leading to directed motion toward regions of higher persistence. Our results demonstrate that persistent motion alone is sufficient to drive topotaxis and could serve as a starting point for more detailed studies on self-propelled particles and cells. I. INTRODUCTION quently force the cells to move around them. If the ob- stacles' density smoothly varies across the substrate, the Whether in vitro or in vivo, cellular motion is of- cells have been shown to perform topotaxis: the topo- ten biased by directional cues from the cell's micro- graphical gradient serves as a directional cue for the cells environment.
  • Signaling Networks That Regulate Cell Migration

    Signaling Networks That Regulate Cell Migration

    Downloaded from http://cshperspectives.cshlp.org/ on September 29, 2021 - Published by Cold Spring Harbor Laboratory Press Signaling Networks that Regulate Cell Migration Peter Devreotes1 and Alan Rick Horwitz2 1Department of Cell Biology, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205 2Department of Cell Biology, University of Virginia School of Medicine, Charlottesville, Virginia 22908 Correspondence: [email protected] a005959 SUMMARY Stimuli that promote cell migration, such as chemokines, cytokines, and growth factors in metazoans and cyclic AMP in Dictyostelium, activate signaling pathways that control organi- zation of the actin cytoskeleton and adhesion complexes. The Rho-family GTPases are a key convergence point of these pathways. Their effectors include actin regulators such as formins, members of the WASP/WAVEfamilyand the Arp2/3 complex, and the myosin II motor protein. Pathways that link to the Rho GTPases include Ras GTPases, TorC2, and PI3K. Many of the molecules involved form gradients within cells, which define the front and rear of migrating cells, and are also established in related cellular behaviors such as neuronal growth cone extension and cytokinesis. The signaling molecules that regulate migration can be integrated to provide a model of network function. The network displays biochemical excitability seen as spontaneous waves of activation that propagate along the cell cortex. These events coordinate cell movement and can be biased by external cues to bring about directed migration. Outline 1
  • Single Cells of Pseudomonas Aeruginosa Exhibit Electrotaxis And

    Single Cells of Pseudomonas Aeruginosa Exhibit Electrotaxis And

    bioRxiv preprint doi: https://doi.org/10.1101/020511; this version posted June 5, 2015. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 1 Single Cells of Pseudomonas aeruginosa 2 Exhibit Electrotaxis and Durotaxis Behaviours 3 4 5 6 7 Lee Preiss1 8 Suresh Neethirajan1* 9 10 11 1BioNano Laboratory, School of Engineering, University of Guelph, Guelph, ON Canada N1G 12 2W1 13 *Corresponding author: Email: [email protected] 14 1 bioRxiv preprint doi: https://doi.org/10.1101/020511; this version posted June 5, 2015. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-NC-ND 4.0 International license. 15 Abstract 16 Pseudomonas aeruginosa is frequently associated with nosocomial infections, including 17 polymicrobial wound infections and the complex biofilm communities that reside within the 18 cystic fibrosis lung. P. aeruginosa utilizes flagellum-mediated motility to approach, attach to, 19 and spread across a surface using a combination of swimming, swarming and twitching (type IV 20 pili extension and retraction) motility. We report an innovative approach to study the motility of 21 single P. aeruginosa cells in microfluidic channels possessing different structural geometry, all 22 with the flexibility of being able to manipulate stiffness gradients and electric fields to 23 investigate changes in motility in response to specific stimuli.