The ins and outs of engineering functional tissues and organs:evaluating the in-vitro and in-situ processes

Purpose of review For many disorders that result in loss of organ function, the only curative treatment is organ transplantation. However, this approach is severely limited by the shortage of donor organs. Tissue engineering has emerged as an alternative solution to this issue. This review discusses the concept of tissue engineering from a technical viewpoint and summarizes the state of the art as well as the current shortcomings, with the aim of identifying the key lessons that we can learn to further advance the engineering of functional tissues and organs.\u3cbr/\u3e\u3cbr/\u3eRecent findings A plethora of tissue-engineering strategies have been recently developed. Notably, these strategies put different emphases on the in-vitro and in-situ processes (i.e. preimplantation and postimplantation) that take place during tissue formation. Biophysical and biomechanical interactions between the cells and the scaffold/biomaterial play a crucial role in all steps and have started to be exploited to steer tissue regeneration.\u3cbr/\u3e\u3cbr/\u3eSummary Recent works have demonstrated the need to better understand the in-vitro and in-situ processes during tissue formation, in order to regenerate complex, functional organs with desired cellular organization and tissue architecture. A concerted effort from both fundamental and tissue-specific research has the potential to accelerate progress in the field

Mechanobiology of the cell–matrix interplay: catching a glimpse of complexity via minimalistic models

Biological tissues present a grand challenge for mechanicians. Not only are their mechanical properties complex, they show a strong spatiotemporal heterogeneity due to ongoing and active remodeling of the living matter they are composed of: cells and extracellular matrix (ECM). The main sensors and effectors in this process are the cells, which direct tissue structure and mechanics by changing their own behavior and by producing and reorganizing the ECM. Vice versa, the ECM exhibits unique mechanical signatures associated with its fibrous polymer network that can modulate cell behavior. Grasping the full complexity of this reciprocal mechanical interaction between cells and the ECM is key for understanding physiological tissue function and maladaptation. An emerging approach is to explain the role of tissue biomechanics one component at a time and gradually (re)build tissue complexity. Here we highlight how this approach has been valuable in providing new insights in the relative and combined roles of cells and matrix, and in raising new questions into the origins of cellular and tissue responses. The answers may offer new approaches for mechanically driven tissue regeneration and biomaterial design

Cellular geometry sensing at different length scales and its implications for scaffold design

\u3cp\u3eGeometrical cues provided by the intrinsic architecture of tissues and implanted biomaterials have a high relevance in controlling cellular behavior. Knowledge of how cells sense and subsequently respond to complex geometrical cues of various sizes and origins is needed to understand the role of the architecture of the extracellular environment as a cell-instructive parameter. This is of particular interest in the field of tissue engineering, where the success of scaffold-guided tissue regeneration largely depends on the formation of new tissue in a native-like organization in order to ensure proper tissue function. A well-considered internal scaffold design (i.e., the inner architecture of the porous structure) can largely contribute to the desired cell and tissue organization. Advances in scaffold production techniques for tissue engineering purposes in the last years have provided the possibility to accurately create scaffolds with defined macroscale external and microscale internal architectures. Using the knowledge of how cells sense geometrical cues of different size ranges can drive the rational design of scaffolds that control cellular and tissue architecture. This concise review addresses the recently gained knowledge of the sensory mechanisms of cells towards geometrical cues of different sizes (from the nanometer to millimeter scale) and points out how this insight can contribute to informed architectural scaffold designs.\u3c/p\u3

An automated quantitative analysis of cell, nucleus and focal adhesion morphology

\u3cp\u3eAdherent cells sense the physical properties of their environment via focal adhesions. Improved understanding of how cells sense and response to their physical surroundings is aided by quantitative evaluation of focal adhesion size, number, orientation, and distribution in conjunction with the morphology of single cells and the corresponding nuclei. We developed a fast, user-friendly and automated image analysis algorithm capable of capturing and characterizing these individual components with a high level of accuracy. We demonstrate the robustness and applicability of the algorithm by quantifying morphological changes in response to a variety of environmental changes as well as manipulations of cellular components of mechanotransductions. Finally, as a proof-of-concept we use our algorithm to quantify the effect of Rho-associated kinase inhibitor Y-27632 on focal adhesion maturation. We show that a decrease in cell contractility leads to a decrease in focal adhesion size and aspect ratio.\u3c/p\u3

Factor XIII stiffens fibrin clots by causing fiber compaction

\u3cbr/\u3eBackground\u3cbr/\u3eFactor XIII-induced cross-linking has long been associated with the ability of fibrin blood clots to resist mechanical deformation, but how FXIII can directly modulate clot stiffness is unknown.\u3cbr/\u3eObjectives and Methods\u3cbr/\u3eWe hypothesized that FXIII affects the self-assembly of fibrin fibers by altering the lateral association between protofibrils. To test this hypothesis, we studied the cross-linking kinetics and the structural evolution of the fibers and clots during the formation of plasma-derived and recombinant fibrins by using light scattering, and the response of the clots to mechanical stresses by using rheology.\u3cbr/\u3eResults\u3cbr/\u3eWe show that the lateral aggregation of fibrin protofibrils initially results in the formation of floppy fibril bundles, which then compact to form tight and more rigid fibers. The first stage is reflected in a fast (10 min) increase in clot stiffness, whereas the compaction phase is characterized by a slow (hours) development of clot stiffness. Inhibition of FXIII completely abrogates the slow compaction. FXIII strongly increases the linear elastic modulus of the clots, but does not affect the non-linear response at large deformations.\u3cbr/\u3eConclusions\u3cbr/\u3eWe propose a multiscale structural model whereby FXIII-mediated cross-linking tightens the coupling between the protofibrils within a fibrin fiber, thus making the fiber stiffer and less porous. At small strains, fiber stiffening enhances clot stiffness, because the clot response is governed by the entropic elasticity of the fibers, but once the clot is sufficiently stressed, the modulus is independent of protofibril coupling, because clot stiffness is governed by individual protofibril stretching.\u3cbr/\u3

Mechanical adaptability of cell migration in 3D collagen gels

Migration of cells across tissues with diverse biophysical environments plays a crucial role in a wide variety of physiological functions and pathological processes, such as in embryonic development, wound healing, haemostasis, tumor and cancer progression. Indeed, one of the most devastating features of cancer is metastasis_the ability of cancer cells to escape from the primary tumor and invade and colonize a distant tissue. Understanding the biophysical and biochemical mechanisms underlying cell migration remains a challenge, however, partly because it has been only recently realized that cells employ different strategies and molecular mechanisms in three-dimensional (3D) environments, compared to on traditional 2D glass surfaces.\u3cbr/\u3e\u3cbr/\u3eIn this work, we examined cell migration, simultaneously at the individual cell and cell population levels, in a 3D collagen hydrogel model mimicking the connective tissue topology confronted by malignant breast cancer cells. Our findings revealed two distinct migration patterns that depend specifically on the location of the individual cells within the population: a rapid and directionally persistent migration of the “leader cells” and a more randomized migration of the “follower cells”. This disparity, strikingly, occurred with minimal cell-cell contacts. Rather, this heterogeneity is associated with local remodeling of the pericellular matrix and results in an apparent independence of the inherent migration on matrix condition. Despite such robustness, effects of anti-migratory drugs were interestingly observed to vary strongly with matrix stiffness and architecture. Specifically, cytoskeletal contractility-targeting drugs reduced migration speed in sparse gels, whereas migration in dense gels was retarded effectively by inhibiting proteolysis. Our results therefore corroborate a mechanistic plasticity that allows cells to actively adapt their invasion machinery depending on the local biophysical microenvironment.\u3cbr/\u3

Mechanobiology of cell migration in the context of dynamic two-way cell–matrix interactions

Migration of cells is integral in various physiological processes in all facets of life. These range from embryonic development, morphogenesis, and wound healing, to disease pathology such as cancer metastasis. While cell migratory behavior has been traditionally studied using simple assays on culture dishes, in recent years it has been increasingly realized that the physical, mechanical, and chemical aspects of the matrix are key determinants of the migration mechanism. In this paper, we will describe the mechanobiological changes that accompany the dynamic cell–matrix interactions during cell migration. Furthermore, we will review what is to date known about how these changes feed back to the dynamics and biomechanical properties of the cell and the matrix. Elucidating the role of these intimate cell–matrix interactions will provide not only a better multi-scale understanding of cell motility in its physiological context, but also a more holistic perspective for designing approaches to regulate cell behavior

Concentric gel system to study the biophysical role of matrix microenvironment on 3D cell migration

The ability of cells to migrate is crucial in a wide variety of cell functions throughout life from embryonic development and wound healing to tumor and cancer metastasis. Despite intense research efforts, the basic biochemical and biophysical principles of cell migration are still not fully understood, especially in the physiologically relevant three-dimensional (3D) microenvironments. Here, we describe an in vitro assay designed to allow quantitative examination of 3D cell migration behaviors. The method exploits the cell’s mechanosensing ability and propensity to migrate into previously unoccupied extracellular matrix (ECM). We use the invasion of highly invasive breast cancer cells, MDA-MB-231, in collagen gels as a model system. The spread of cell population and the migration dynamics of individual cells over weeks of culture can be monitored using live-cell imaging and analyzed to extract spatiotemporally-resolved data. Furthermore, the method is easily adaptable for diverse extracellular matrices, thus offering a simple yet powerful way to investigate the role of biophysical factors in the microenvironment on cell migration

Cell-perceived substrate curvature dynamically coordinates the direction, speed, and persistence of stromal cell migration

Adherent cells residing within tissues or biomaterials are presented with 3D geometrical cues from their environment, often in the form of local surface curvatures. While there is growing evidence that cellular decision‐making is influenced by substrate curvature, the effect of physiologically relevant, cell‐scale anisotropic curvatures remains poorly understood. This study systematically explores the migration behavior of human bone marrow stromal cells (hBMSCs) on a library of anisotropic curved structures. Analysis of cell trajectories reveals that, on convex cylindrical structures, hBMSC migration speed and persistence are strongly governed by the cellular orientation on the curved structure, while migration on concave cylindrical structures is characterized by fast but non‐aligned and non‐persistent migration. Concurrent presentation of concave and convex substrates on toroidal structures induces migration in the direction where hBMSCs can most effectively avoid cell bending. These distinct migration behaviors are found to be universally explained by the cell‐perceived substrate curvature, which on anisotropic curved structures is dependent on both the temporally varying cell orientation and the 3D cellular morphology. This work demonstrates that cell migration is dynamically guided by the perceived curvature of the underlying substrate, providing an important biomaterial design parameter for instructing cell migration in tissue engineering and regenerative medicine