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Regenerative medicine: from engineering to clinical applications
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Journal o f Biomechanics 2006, Vol. 39 (Suppl 1)
Wednesday, August 2 7716 We, 10:00-10:30 (P29) Regenerative medicine: from engineering to clinical applications J.F. Stoltz. Department on Cell and Tissue Therapy, University Hospital and Bioengineering group, UMR CNRS 7563, LEMTA, Medical Faculty (UHP), Vandoeuvre les Nancy, France Most human tissues do not regenerate spontaneously, this is why cell therapies and clinical engineering are promising alternatives. The principle is simple: cells are collected in a patient and introduced in the damaged tissue or in a porous support and harvested in a bioreactor in which the physico-chemical and mechanical parameters are controlled. Once the tissues (or the cells) are mature they may be implanted. In parallel, the development of new biotherapies with stem cells is a recent field of research with many potential clinical applications. Embryonic stem cells are potentially more interesting (they are totipotent), but they can only be obtained at the very early stages of the embryo. The potential of adult stem cells is more limited but isolating them induces no ethical problem. The properties of blood cells from the umbilical cord are forerunners of the haematopoietic system, but their abilities to participate to the regeneration of other tissues is a new way of research. Finally, gene therapy has been nourishing high hopes but few clinical applications can be envisaged in the short term. A large number of potential methods exist for each tissue or type of therapy. In other respect, as well as biochemical parameters, mechanical forces (pressure, shear stress ... ) influence the differentiation of cells that are used. These parameters are now considered as critical not only for understanding pathological mechanisms (osteoarthritis, inflammation, atherosclerosis . . . ) and also for tissue reconstruction. Thus, the issue of in vitro tissue or cell culture is multifaceted as it involves genetics for the choice of initial cells (progenitor cells, differentiated cells or genetically modified cells), biochemistry (choice of the polymeric scaffold) and mechanics (magnitude and frequency). In summary, the 21th century begins with perspectives in clinical engineering (mainly regenerative medicine). The clinical applications open new therapeutic horizons that can be hardly imagined today. These domains of research have also provoked intrusion of ethics and religion into scientific field (i.e. therapeutic cloning). Various examples of this new therapeutic medicine base on engineering are developed in this work (cartilage engineering, cardiac insufficiency, liver insufficiency, cancer therapies . . . . ). This work was supported by Region de Lorraine (CPER) and university Henri Poincar6 (Nancy, France).
Thursday, August 3 6172 Th, 10:00-10:30 (P39) The role o f molecular mechanics in intracellular signaling: Mechanisms and models R.D. Kamm. Biological Engineering Division, Department of Mechanical Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA That cells respond to mechanical stimuli and the nature of their response in various situations has been known and intensely studied for several decades. Yet, despite rigorous investigation, we still understand little about the mechanisms by which a cell senses mechanical force. Many processes likely contribute to the observed response, and several of these have begun to be studied. Some share a common underlying mechanism, a conformational change in a force bearing protein that alters its binding affinity, enzymatic activity, or, in the case of a channel, its conductivity. To understand these processes, one must first identify how forces are transmitted throughout the cell, determine the forces experienced by individual transducing proteins, analyze the resulting force-induced conformational change, and finally, determine the change in biochemical activity or channel conductance. Consideration of these collective processes naturally leads to the notion of a "mechanical signaling pathway" that operates parallel to, and that communicates with, the better-known biochemical signaling pathways. Cross-talk between these two constitutes the domain of the emerging field of mechanobiology. Progress in unraveling the details of mechanical signaling has been limited by the slow progress in the necessary experimental and computational tools. Methods now exist, however, to both probe the response of a single molecule to force, and to computationally analyze the distribution and molecular consequences of force. In this lecture, some of the new experimental methods for
Plenary Lectures probing single molecule function will be described, as well as current state of computational approaches used to analyze force transmission across the cell membrane and through the cytoskeleton, and to study conformational change in single proteins.
Friday, August 4 6380 Fr, 10:00-10:30 (P50) Endothelial biomechanics: focusing on the dynamic behavior o f cytoskeletal structure M. Sato. Department of Bioengineering and Robotics, Graduate School of Engineering, Tohoku University, Sendai, Japan Endothelial cells respond to mechanical stimuli, demonstrating responses that include changes in cytoskeletal structure, cell shape and cell function. Cells adhere to extracellular matrix at focal adhesions (FA), which are believed to play an important role in determining cell shape. To understand the mechanism of cell remodeling by mechanical stimuli, the dynamic behavior of FA and actin filaments was observed in endothelial cells exposed to shear stress. For this study, bovine aortic endothelial cells were obtained from thoracic aortas. RFP-FAT (focal adhesion targeting) and GFP-actin were co-transfected into cells plated in glass-bottom dishes. The confluent endothelial cell monolayer was loaded into a parallel-plate flow chamber and laminar shear stress of 1.5-2 Pa was applied to the cells. Cells expressing RFP-FAT and GFP-actin were observed under fluid flow conditions using a confocal laser-scanning microscope. Actin filaments arranged orthogonal to the direction of flow initially shrunk on exposure to shear stress. FAT relocated with the actin filaments, decreasing along the orthogonal direction. After this, lamellipodia appeared at the upstream side of the cells. During this cell remodeling process, the dynamic behavior of FAT including appearance/disappearance, elongation, sliding and aggregation was observed. Within 20 rain, significant changes in the position of FAT were no longer observed. In addition, the biomechanical properties of stress fibers isolated from cultured smooth muscle cells were measured using a custom-designed micro-tensile testing apparatus. When stress fibers were elongated to greater than two-fold the initial length, the resistance to elongation increased with strain demonstrating a nonlinear stress-strain relationship. The initial elastic modulus of stress fibers, 1.45 MPa, was much lower than that of synthesized single F-actin, 1.8 GPa. The force required to stretch the isolated stress fibers back to the original lengths (i.e. intact state in the cytoplasm) was approximately 10 nN. This value is comparable in magnitude to traction forces applied by adherent cells at single focal adhesion sites, suggesting that the tensions in stress fibers are balanced with the traction force to maintain cell structure integrity.
Journal o f Biomechanics 2006, Vol. 39 (Suppl 1)
Wednesday, August 2 7716 We, 10:00-10:30 (P29) Regenerative medicine: from engineering to clinical applications J.F. Stoltz. Department on Cell and Tissue Therapy, University Hospital and Bioengineering group, UMR CNRS 7563, LEMTA, Medical Faculty (UHP), Vandoeuvre les Nancy, France Most human tissues do not regenerate spontaneously, this is why cell therapies and clinical engineering are promising alternatives. The principle is simple: cells are collected in a patient and introduced in the damaged tissue or in a porous support and harvested in a bioreactor in which the physico-chemical and mechanical parameters are controlled. Once the tissues (or the cells) are mature they may be implanted. In parallel, the development of new biotherapies with stem cells is a recent field of research with many potential clinical applications. Embryonic stem cells are potentially more interesting (they are totipotent), but they can only be obtained at the very early stages of the embryo. The potential of adult stem cells is more limited but isolating them induces no ethical problem. The properties of blood cells from the umbilical cord are forerunners of the haematopoietic system, but their abilities to participate to the regeneration of other tissues is a new way of research. Finally, gene therapy has been nourishing high hopes but few clinical applications can be envisaged in the short term. A large number of potential methods exist for each tissue or type of therapy. In other respect, as well as biochemical parameters, mechanical forces (pressure, shear stress ... ) influence the differentiation of cells that are used. These parameters are now considered as critical not only for understanding pathological mechanisms (osteoarthritis, inflammation, atherosclerosis . . . ) and also for tissue reconstruction. Thus, the issue of in vitro tissue or cell culture is multifaceted as it involves genetics for the choice of initial cells (progenitor cells, differentiated cells or genetically modified cells), biochemistry (choice of the polymeric scaffold) and mechanics (magnitude and frequency). In summary, the 21th century begins with perspectives in clinical engineering (mainly regenerative medicine). The clinical applications open new therapeutic horizons that can be hardly imagined today. These domains of research have also provoked intrusion of ethics and religion into scientific field (i.e. therapeutic cloning). Various examples of this new therapeutic medicine base on engineering are developed in this work (cartilage engineering, cardiac insufficiency, liver insufficiency, cancer therapies . . . . ). This work was supported by Region de Lorraine (CPER) and university Henri Poincar6 (Nancy, France).
Thursday, August 3 6172 Th, 10:00-10:30 (P39) The role o f molecular mechanics in intracellular signaling: Mechanisms and models R.D. Kamm. Biological Engineering Division, Department of Mechanical Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA That cells respond to mechanical stimuli and the nature of their response in various situations has been known and intensely studied for several decades. Yet, despite rigorous investigation, we still understand little about the mechanisms by which a cell senses mechanical force. Many processes likely contribute to the observed response, and several of these have begun to be studied. Some share a common underlying mechanism, a conformational change in a force bearing protein that alters its binding affinity, enzymatic activity, or, in the case of a channel, its conductivity. To understand these processes, one must first identify how forces are transmitted throughout the cell, determine the forces experienced by individual transducing proteins, analyze the resulting force-induced conformational change, and finally, determine the change in biochemical activity or channel conductance. Consideration of these collective processes naturally leads to the notion of a "mechanical signaling pathway" that operates parallel to, and that communicates with, the better-known biochemical signaling pathways. Cross-talk between these two constitutes the domain of the emerging field of mechanobiology. Progress in unraveling the details of mechanical signaling has been limited by the slow progress in the necessary experimental and computational tools. Methods now exist, however, to both probe the response of a single molecule to force, and to computationally analyze the distribution and molecular consequences of force. In this lecture, some of the new experimental methods for
Plenary Lectures probing single molecule function will be described, as well as current state of computational approaches used to analyze force transmission across the cell membrane and through the cytoskeleton, and to study conformational change in single proteins.
Friday, August 4 6380 Fr, 10:00-10:30 (P50) Endothelial biomechanics: focusing on the dynamic behavior o f cytoskeletal structure M. Sato. Department of Bioengineering and Robotics, Graduate School of Engineering, Tohoku University, Sendai, Japan Endothelial cells respond to mechanical stimuli, demonstrating responses that include changes in cytoskeletal structure, cell shape and cell function. Cells adhere to extracellular matrix at focal adhesions (FA), which are believed to play an important role in determining cell shape. To understand the mechanism of cell remodeling by mechanical stimuli, the dynamic behavior of FA and actin filaments was observed in endothelial cells exposed to shear stress. For this study, bovine aortic endothelial cells were obtained from thoracic aortas. RFP-FAT (focal adhesion targeting) and GFP-actin were co-transfected into cells plated in glass-bottom dishes. The confluent endothelial cell monolayer was loaded into a parallel-plate flow chamber and laminar shear stress of 1.5-2 Pa was applied to the cells. Cells expressing RFP-FAT and GFP-actin were observed under fluid flow conditions using a confocal laser-scanning microscope. Actin filaments arranged orthogonal to the direction of flow initially shrunk on exposure to shear stress. FAT relocated with the actin filaments, decreasing along the orthogonal direction. After this, lamellipodia appeared at the upstream side of the cells. During this cell remodeling process, the dynamic behavior of FAT including appearance/disappearance, elongation, sliding and aggregation was observed. Within 20 rain, significant changes in the position of FAT were no longer observed. In addition, the biomechanical properties of stress fibers isolated from cultured smooth muscle cells were measured using a custom-designed micro-tensile testing apparatus. When stress fibers were elongated to greater than two-fold the initial length, the resistance to elongation increased with strain demonstrating a nonlinear stress-strain relationship. The initial elastic modulus of stress fibers, 1.45 MPa, was much lower than that of synthesized single F-actin, 1.8 GPa. The force required to stretch the isolated stress fibers back to the original lengths (i.e. intact state in the cytoplasm) was approximately 10 nN. This value is comparable in magnitude to traction forces applied by adherent cells at single focal adhesion sites, suggesting that the tensions in stress fibers are balanced with the traction force to maintain cell structure integrity.