@article{RoedelAbdelilahSeyfried2021, author = {R{\"o}del, Claudia Jasmin and Abdelilah-Seyfried, Salim}, title = {A zebrafish toolbox for biomechanical signaling in cardiovascular development and disease}, series = {Current opinion in hematology}, volume = {28}, journal = {Current opinion in hematology}, number = {3}, publisher = {Lippincott Williams \& Wilkins}, address = {Philadelphia}, issn = {1065-6251}, doi = {10.1097/MOH.0000000000000648}, pages = {198 -- 207}, year = {2021}, abstract = {Purpose of review The zebrafish embryo has emerged as a powerful model organism to investigate the mechanisms by which biophysical forces regulate vascular and cardiac cell biology during development and disease. A versatile arsenal of methods and tools is available to manipulate and analyze biomechanical signaling. This review aims to provide an overview of the experimental strategies and tools that have been utilized to study biomechanical signaling in cardiovascular developmental processes and different vascular disease models in the zebrafish embryo. Within the scope of this review, we focus on work published during the last two years. Recent findings Genetic and pharmacological tools for the manipulation of cardiac function allow alterations of hemodynamic flow patterns in the zebrafish embryo and various types of transgenic lines are available to report endothelial cell responses to biophysical forces. These tools have not only revealed the impact of biophysical forces on cardiovascular development but also helped to establish more accurate models for cardiovascular diseases including cerebral cavernous malformations, hereditary hemorrhagic telangiectasias, arteriovenous malformations, and lymphangiopathies. Summary The zebrafish embryo is a valuable vertebrate model in which in-vivo manipulations of biophysical forces due to cardiac contractility and blood flow can be performed. These analyses give important insights into biomechanical signaling pathways that control endothelial and endocardial cell behaviors. The technical advances using this vertebrate model will advance our understanding of the impact of biophysical forces in cardiovascular pathologies.}, language = {en} } @phdthesis{Ziege2022, author = {Ziege, Ricardo}, title = {Growth dynamics and mechanical properties of E. coli biofilms}, doi = {10.25932/publishup-55986}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-559869}, school = {Universit{\"a}t Potsdam}, pages = {xi, 123}, year = {2022}, abstract = {Biofilms are complex living materials that form as bacteria get embedded in a matrix of self-produced protein and polysaccharide fibres. The formation of a network of extracellular biopolymer fibres contributes to the cohesion of the biofilm by promoting cell-cell attachment and by mediating biofilm-substrate interactions. This sessile mode of bacteria growth has been well studied by microbiologists to prevent the detrimental effects of biofilms in medical and industrial settings. Indeed, biofilms are associated with increased antibiotic resistance in bacterial infections, and they can also cause clogging of pipelines or promote bio-corrosion. However, biofilms also gained interest from biophysics due to their ability to form complex morphological patterns during growth. Recently, the emerging field of engineered living materials investigates biofilm mechanical properties at multiple length scales and leverages the tools of synthetic biology to tune the functions of their constitutive biopolymers. This doctoral thesis aims at clarifying how the morphogenesis of Escherichia coli (E. coli) biofilms is influenced by their growth dynamics and mechanical properties. To address this question, I used methods from cell mechanics and materials science. I first studied how biological activity in biofilms gives rise to non-uniform growth patterns. In a second study, I investigated how E. coli biofilm morphogenesis and its mechanical properties adapt to an environmental stimulus, namely the water content of their substrate. Finally, I estimated how the mechanical properties of E. coli biofilms are altered when the bacteria express different extracellular biopolymers. On nutritive hydrogels, micron-sized E. coli cells can build centimetre-large biofilms. During this process, bacterial proliferation and matrix production introduce mechanical stresses in the biofilm, which release through the formation of macroscopic wrinkles and delaminated buckles. To relate these biological and mechanical phenomena, I used time-lapse fluorescence imaging to track cell and matrix surface densities through the early and late stages of E. coli biofilm growth. Colocalization of high cell and matrix densities at the periphery precede the onset of mechanical instabilities at this annular region. Early growth is detected at this outer annulus, which was analysed by adding fluorescent microspheres to the bacterial inoculum. But only when high rates of matrix production are present in the biofilm centre, does overall biofilm spreading initiate along the solid-air interface. By tracking larger fluorescent particles for a long time, I could distinguish several kinematic stages of E. coli biofilm expansion and observed a transition from non-linear to linear velocity profiles, which precedes the emergence of wrinkles at the biofilm periphery. Decomposing particle velocities to their radial and circumferential components revealed a last kinematic stage, where biofilm movement is mostly directed towards the radial delaminated buckles, which verticalize. The resulting compressive strains computed in these regions were observed to substantially deform the underlying agar substrates. The co-localization of higher cell and matrix densities towards an annular region and the succession of several kinematic stages are thus expected to promote the emergence of mechanical instabilities at the biofilm periphery. These experimental findings are predicted to advance future modelling approaches of biofilm morphogenesis. E. coli biofilm morphogenesis is further anticipated to depend on external stimuli from the environment. To clarify how the water could be used to tune biofilm material properties, we quantified E. coli biofilm growth, wrinkling dynamics and rigidity as a function of the water content of the nutritive substrates. Time-lapse microscopy and computational image analysis revealed that substrates with high water content promote biofilm spreading kinetics, while substrates with low water content promote biofilm wrinkling. The wrinkles observed on biofilm cross-sections appeared more bent on substrates with high water content, while they tended to be more vertical on substrates with low water content. Both wet and dry biomass, accumulated over 4 days of culture, were larger in biofilms cultured on substrates with high water content, despite extra porosity within the matrix layer. Finally, the micro-indentation analysis revealed that substrates with low water content supported the formation of stiffer biofilms. This study shows that E. coli biofilms respond to the water content of their substrate, which might be used for tuning their material properties in view of further applications. Biofilm material properties further depend on the composition and structure of the matrix of extracellular proteins and polysaccharides. In particular, E. coli biofilms were suggested to present tissue-like elasticity due to a dense fibre network consisting of amyloid curli and phosphoethanolamine-modified cellulose. To understand the contribution of these components to the emergent mechanical properties of E. coli biofilms, we performed micro-indentation on biofilms grown from bacteria of several strains. Besides showing higher dry masses, larger spreading diameters and slightly reduced water contents, biofilms expressing both main matrix components also presented high rigidities in the range of several hundred kPa, similar to biofilms containing only curli fibres. In contrast, a lack of amyloid curli fibres provides much higher adhesive energies and more viscoelastic fluid-like material behaviour. Therefore, the combination of amyloid curli and phosphoethanolamine-modified cellulose fibres implies the formation of a composite material whereby the amyloid curli fibres provide rigidity to E. coli biofilms, whereas the phosphoethanolamine-modified cellulose rather acts as a glue. These findings motivate further studies involving purified versions of these protein and polysaccharide components to better understand how their interactions benefit biofilm functions. All three studies depict different aspects of biofilm morphogenesis, which are interrelated. The first work reveals the correlation between non-uniform biological activities and the emergence of mechanical instabilities in the biofilm. The second work acknowledges the adaptive nature of E. coli biofilm morphogenesis and its mechanical properties to an environmental stimulus, namely water. Finally, the last study reveals the complementary role of the individual matrix components in the formation of a stable biofilm material, which not only forms complex morphologies but also functions as a protective shield for the bacteria it contains. Our experimental findings on E. coli biofilm morphogenesis and their mechanical properties can have further implications for fundamental and applied biofilm research fields.}, language = {en} } @article{BornhorstAbdelilahSeyfried2021, author = {Bornhorst, Dorothee and Abdelilah-Seyfried, Salim}, title = {Strong as a Hippo's Heart: Biomechanical Hippo Signaling During Zebrafish Cardiac Development}, series = {Frontiers in Cell and Developmental Biology}, volume = {9}, journal = {Frontiers in Cell and Developmental Biology}, publisher = {Frontiers Media}, address = {Lausanne, Schweiz}, issn = {2296-634X}, doi = {10.3389/fcell.2021.731101}, pages = {1 -- 10}, year = {2021}, abstract = {The heart is comprised of multiple tissues that contribute to its physiological functions. During development, the growth of myocardium and endocardium is coupled and morphogenetic processes within these separate tissue layers are integrated. Here, we discuss the roles of mechanosensitive Hippo signaling in growth and morphogenesis of the zebrafish heart. Hippo signaling is involved in defining numbers of cardiac progenitor cells derived from the secondary heart field, in restricting the growth of the epicardium, and in guiding trabeculation and outflow tract formation. Recent work also shows that myocardial chamber dimensions serve as a blueprint for Hippo signaling-dependent growth of the endocardium. Evidently, Hippo pathway components act at the crossroads of various signaling pathways involved in embryonic zebrafish heart development. Elucidating how biomechanical Hippo signaling guides heart morphogenesis has direct implications for our understanding of cardiac physiology and pathophysiology.}, language = {en} } @misc{BornhorstAbdelilahSeyfried2021, author = {Bornhorst, Dorothee and Abdelilah-Seyfried, Salim}, title = {Strong as a Hippo's Heart: Biomechanical Hippo Signaling During Zebrafish Cardiac Development}, series = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe}, journal = {Zweitver{\"o}ffentlichungen der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe}, publisher = {Universit{\"a}tsverlag Potsdam}, address = {Potsdam}, issn = {1866-8372}, doi = {10.25932/publishup-54873}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-548731}, pages = {1 -- 10}, year = {2021}, abstract = {The heart is comprised of multiple tissues that contribute to its physiological functions. During development, the growth of myocardium and endocardium is coupled and morphogenetic processes within these separate tissue layers are integrated. Here, we discuss the roles of mechanosensitive Hippo signaling in growth and morphogenesis of the zebrafish heart. Hippo signaling is involved in defining numbers of cardiac progenitor cells derived from the secondary heart field, in restricting the growth of the epicardium, and in guiding trabeculation and outflow tract formation. Recent work also shows that myocardial chamber dimensions serve as a blueprint for Hippo signaling-dependent growth of the endocardium. Evidently, Hippo pathway components act at the crossroads of various signaling pathways involved in embryonic zebrafish heart development. Elucidating how biomechanical Hippo signaling guides heart morphogenesis has direct implications for our understanding of cardiac physiology and pathophysiology.}, language = {en} } @misc{MuenchAbdelilahSeyfried2021, author = {M{\"u}nch, Juliane and Abdelilah-Seyfried, Salim}, title = {Sensing and Responding of Cardiomyocytes to Changes of Tissue Stiffness in the Diseased Heart}, series = {Postprints der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe}, journal = {Postprints der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe}, issn = {1866-8372}, doi = {10.25932/publishup-54580}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-545805}, pages = {15}, year = {2021}, abstract = {Cardiomyocytes are permanently exposed to mechanical stimulation due to cardiac contractility. Passive myocardial stiffness is a crucial factor, which defines the physiological ventricular compliance and volume of diastolic filling with blood. Heart diseases often present with increased myocardial stiffness, for instance when fibrotic changes modify the composition of the cardiac extracellular matrix (ECM). Consequently, the ventricle loses its compliance, and the diastolic blood volume is reduced. Recent advances in the field of cardiac mechanobiology revealed that disease-related environmental stiffness changes cause severe alterations in cardiomyocyte cellular behavior and function. Here, we review the molecular mechanotransduction pathways that enable cardiomyocytes to sense stiffness changes and translate those into an altered gene expression. We will also summarize current knowledge about when myocardial stiffness increases in the diseased heart. Sophisticated in vitro studies revealed functional changes, when cardiomyocytes faced a stiffer matrix. Finally, we will highlight recent studies that described modulations of cardiac stiffness and thus myocardial performance in vivo. Mechanobiology research is just at the cusp of systematic investigations related to mechanical changes in the diseased heart but what is known already makes way for new therapeutic approaches in regenerative biology.}, language = {en} } @article{MuenchAbdelilahSeyfried2021, author = {M{\"u}nch, Juliane and Abdelilah-Seyfried, Salim}, title = {Sensing and responding of cardiomyocytes to changes of tissue stiffness in the diseased heart}, series = {Frontiers in cell developmental biology}, volume = {9}, journal = {Frontiers in cell developmental biology}, publisher = {Frontiers Media}, address = {Lausanne}, issn = {2296-634X}, doi = {10.3389/fcell.2021.642840}, pages = {13}, year = {2021}, abstract = {Cardiomyocytes are permanently exposed to mechanical stimulation due to cardiac contractility. Passive myocardial stiffness is a crucial factor, which defines the physiological ventricular compliance and volume of diastolic filling with blood. Heart diseases often present with increased myocardial stiffness, for instance when fibrotic changes modify the composition of the cardiac extracellular matrix (ECM). Consequently, the ventricle loses its compliance, and the diastolic blood volume is reduced. Recent advances in the field of cardiac mechanobiology revealed that disease-related environmental stiffness changes cause severe alterations in cardiomyocyte cellular behavior and function. Here, we review the molecular mechanotransduction pathways that enable cardiomyocytes to sense stiffness changes and translate those into an altered gene expression. We will also summarize current knowledge about when myocardial stiffness increases in the diseased heart. Sophisticated in vitro studies revealed functional changes, when cardiomyocytes faced a stiffer matrix. Finally, we will highlight recent studies that described modulations of cardiac stiffness and thus myocardial performance in vivo. Mechanobiology research is just at the cusp of systematic investigations related to mechanical changes in the diseased heart but what is known already makes way for new therapeutic approaches in regenerative biology.}, language = {en} } @phdthesis{Landau2020, author = {Landau, Livnat}, title = {Mechanical stimulation of in-vitro tissue growth using magnetic beads}, pages = {112}, year = {2020}, abstract = {Cells and tissues are sensitive to mechanical forces applied to them. In particular, bone forming cells and connective tissues, composed of cells embedded in fibrous extracellular matrix (ECM), are continuously remodeled in response to the loads they bear. The mechanoresponses of cells embedded in tissue include proliferation, differentiation, apoptosis, internal signaling between cells, and formation and resorption of tissue. Experimental in-vitro systems of various designs have demonstrated that forces affect tissue growth, maturation and mineralization. However, the results depended on different parameters such as the type and magnitude of the force applied in each study. Some experiments demonstrated that applied forces increase cell proliferation and inhibit cell maturation rate, while other studies found the opposite effect. When the effect of different magnitudes of forces was compared, some studies showed that higher forces resulted in a cell proliferation increase or differentiation decrease, while other studies observed the opposite trend or no trend at all. In this study, MC3T3-E1 cells, a cell line of pre-osteoblasts (bone forming cells), was used. In this cell line, cell differentiation is known to accelerate after cells stop proliferating, typically at confluency. This makes this cell line an interesting subject for studying the influence of forces on the switch between the proliferation stage of the precursor cell and the differentiation to the mature osteoblasts. A new experimental system was designed to perform systematic investigations of the influence of the type and magnitude of forces on tissue growth. A single well plate contained an array of 80 rectangular pores. Each pore was seeded with MC3T3-E1 cells. The culture medium contained magnetic beads (MBs) of 4.5 μm in diameter that were incorporated into the pre-osteoblast cells. Using an N52 neodymium magnet, forces ranging over three orders of magnitude were applied to MBs incorporated in cells at 10 different distances from the magnet. The amount of formed tissue was assessed after 24 days of culture. The experimental design allowed to obtain data concerning (i) the influence of the type of the force (static, oscillating, no force) on tissue growth; (ii) the influence of the magnitude of force (pN-nN range); (iii) the effect of functionalizing the magnetic beads with the tripeptide Arg-Gly-Asp (RGD). To learn about cell differentiation state, in the final state of the tissue growth experiments, an analysis for the expression of alkaline phosphatase (ALP), a well - known marker of osteoblast differentiation, was performed. The experiments showed that the application of static magnetic forces increased tissue growth compared to control, while oscillating forces resulted in tissue growth reduction. A statistically significant positive correlation was found between the amount of tissue grown and the magnitude of the oscillating magnetic force. A positive but non-significant correlation of the amount of tissue with the magnitude of forces was obtained when static forces were applied. Functionalizing the MBs with RGD peptides and applying oscillating forces resulted in an increase of tissue growth relative to tissues incubated with "plain" epoxy MBs. ALP expression decreased as a function of the magnitude of force both when static and oscillating forces were applied. ALP stain intensity was reduced relative to control when oscillating forces were applied and was not significantly different than control for static forces. The suggested interpretation of the experimental findings is that larger mechanical forces delay cell maturation and keep the pre-osteoblasts in a more proliferative stage characterized by more tissue formed and lower expression of ALP. While the influence of the force magnitude can be well explained by an effect of the force on the switch between proliferation and differentiation, the influence of force type (static or oscillating) is less clear. In particular, it is challenging to reconcile the reduction of tissue formed under oscillating forces as compared to controls with the simultaneous reduction of ALP expression. To better understand this, it may be necessary to refine the staining protocol of the scaffolds and to include the amount and structure of ECM as well as other factors that were not monitored in the experiment and which may influence tissue growth and maturation. The developed experimental system proved well suited for a systematic and efficient study of the mechanoresponsiveness of tissue growth, it allowed a study of the dependence of tissue growth on force magnitude ranging over three orders of magnitude, and a comparison between the effect of static and oscillating forces. Future experiments can explore the multiple parameters that affect tissue growth as a function of the magnitude of the force: by applying different time-dependent forces; by extending the force range studied; or by using different cell lines and manipulating the mechanotransduction in the cells biochemically.}, language = {en} } @phdthesis{Ehrig2017, author = {Ehrig, Sebastian}, title = {3D curvature and its role on tissue organization}, school = {Universit{\"a}t Potsdam}, pages = {132}, year = {2017}, abstract = {Shape change is a fundamental process occurring in biological tissues during embryonic development and regeneration of tissues and organs. This process is regulated by cells that are constrained within a complex environment of biochemical and physical cues. The spatial constraint due to geometry has a determining role on tissue mechanics and the spatial distribution of force patterns that, in turn, influences the organization of the tissue structure. An understanding of the underlying principles of tissue organization may have wide consequences for the understanding of healing processes and the development of organs and, as such, is of fundamental interest for the tissue engineering community. This thesis aims to further our understanding of how the collective behaviour of cells is influenced by the 3D geometry of the environment. Previous research studying the role of geometry on tissue growth has mainly focused either on flat surfaces or on substrates where at least one of the principal curvatures is zero. In the present work, tissue growth from MC3T3-E1 pre-osteoblasts was investigated on surfaces of controlled mean curvature. One key aspect of this thesis was the development of substrates of controlled mean curvature and their visualization in 3D. It was demonstrated that substrates of controlled mean curvature suitable for cell culture can be fabricated using liquid polymers and surface tension effects. Using these substrates, it was shown that the mean surface curvature has a strong impact on the rate of tissue growth and on the organization of the tissue structure. It was thereby not only demonstrated that the amount of tissue produced (i.e. growth rates) by the cells depends on the mean curvature of the substrate but also that the tissue surface behaves like a viscous fluid with an equilibrium shape governed by the Laplace-Young-law. It was observed that more tissue was formed on highly concave surfaces compared to flat or convex surfaces. Motivated by these observations, an analytical model was developed, where the rate of tissue growth is a function of the mean curvature, which could successfully describe the growth kinetics. This model was also able to reproduce the growth kinetics of previous experiments where tissues have been cultured in straight-sided prismatic pores. A second part of this thesis focuses on the tissue structure, which influences the mechanical properties of the mature bone tissue. Since the extracellular matrix is produced by the cells, the cell orientation has a strong impact on the direction of the tissue fibres. In addition, it was recently shown that some cell types exhibit collective alignment similar to liquid crystals. Based on this observation, a computational model of self-propelled active particles was developed to explore in an abstract manner how the collective behaviour of cells is influenced by 3D curvature. It was demonstrated that the 3D curvature has a strong impact on the self-organization of active particles and gives, therefore, first insights into the principles of self-organization of cells on curved surfaces.}, language = {en} }