Cyclic Mechanical Stimulus Research Articles

Combined Effects of Cyclic Hypoxic and Mechanical Stimuli on Human Bone Marrow Mesenchymal Stem Cell Differentiation: A New Approach to the Treatment of Bone Loss.

Background: The prevention and treatment of bone loss and osteoporotic fractures is a public health challenge. Combined with normobaric hypoxia, whole-body vibration has a high clinic potential in bone health and body composition. The effect of this therapy may be mediated by its action on bone marrow mesenchymal stem cells (MSCs). Objectives: Evaluate the effects of cyclic low-vibration stimuli and/or hypoxia on bone marrow-derived human MSC differentiation. Methods: MSCs were exposed four days per week, two hours/day, to hypoxia (3% O2) and/or vibration before they were induced to differentiate or during differentiation into osteoblasts or adipocytes. Gene and protein expression of osteoblastic, adipogenic, and cytoskeletal markers were studied, as well as extracellular matrix mineralization and lipid accumulation. Results: early osteoblastic markers increased in undifferentiated MSCs, pretreated in hypoxia and vibration. This pretreatment also increased mRNA levels of osteoblastic genes and beta-catenin protein in the early stages of differentiation into osteoblasts without increasing mineralization. When MSCs were exposed to vibration under hypoxia or normoxia during osteoblastic differentiation, mineralization increased with respect to cultures without vibrational stimuli. In MSCs differentiated into adipocytes, both in those pretreated as well as exposed to different conditions during differentiation, lipid formation decreased. Changes in adipogenic gene expression and increased beta-catenin protein were observed in cultures treated during differentiation. Conclusions: Exposure to cyclic hypoxia in combination with low-intensity vibratory stimuli had positive effects on osteoblastic differentiation and negative ones on adipogenesis of bone marrow-derived MSCs. These results suggest that in elderly or frail people with difficulty performing physical activity, exposure to normobaric cyclic hypoxia and low-density vibratory stimuli could improve bone metabolism and health.

Low-Hysteresis and Tough Ionogels via Low-Energy-Dissipating Cross-Linking.

Low-hysteresis merits can help polymeric gel materials survive from consecutive loading cycles and promote life span in many burgeoning areas. However, it is a big challenge to design low-hysteresis and tough polymeric gel materials, especially for ionogels. This can be attributed to the fact that higher viscosities of ionic liquids (ILs) would increase chain friction of polymeric gels and eventually dissipate large amounts of energy under deformation. Herein, a chemical design of ionogels is proposed to achieve low-hysteresis characteristics in both mechanical and electric aspects via hierarchical aggregates formed by supramolecular self-assembly of quadruple H-bonds in a soft IL-rich polymeric matrix. These self-assembled nanoaggregates not only can greatly reinforce the polymeric matrix and enhance resilience, but also exhibit low-energy-dissipating features under stress conditions, simultaneously benefiting for low-hysteresis properties. These aggregates can also promote toughness and subsequent anti-fatigue properties in response to external cyclic mechanical stimuli. More importantly, these ionogels are presented as a model system to elucidate the underlying mechanism of the low hysteresis and fatigue resistance. Based on these findings, it is further demonstrated that the supramolecular low-hysteresis strategy is universal.

Earthworm-Inspired Ultra-Durable Sliding Triboelectric Nanogenerator with Bionic Self-Replenishing Lubricating Property for Wind Energy Harvesting and Self-Powered Intelligent Sports Monitoring.

Triboelectric nanogenerators (TENGs), a promising strategy for harvesting distributed low-quality power sources, face inevitable bottlenecks regarding long-term abrasion and poor durability. Herein, both issues are addressed by selecting an earthworm-inspired self-replenishing bionic film (ERB) as the tribo-material of sliding-freestanding TENGs (SF-TENGs), it consists of an interconnected 3D porous network structure capable of storing and releasing lubricant under cyclic mechanical stimuli. Thanks to the superiority of self-replenishing property, there is no need for periodic replenishment and accurate content control of lubricant over the interfacial-lubricating SF-TENGs based on dense tribo-layers. Additionally, an SF-TENG based on ERB film (ERB-TENG) demonstrates remarkable output stability with only a slight attenuation of 1% after continuous operation for 100000 cycles. Moreover, the ERB-TENG displays a distinguished anti-wear property, exhibiting no distinct abrasion with an ultra-low coefficient of friction (0.077) and maintaining output stability over a prolonged period of 35 days. Furthermore, integration with an energy management circuit enables the ERB-TENG to achieve a 39-fold boost in charging speed. This work proposes a creative approach to enhance the durability and extend the lifespan of TENG devices, which is also successfully applied to wind energy harvesting and intelligent sports monitoring.

Extracellular Mechanical Stimuli Alters the Metastatic Progression of Prostate Cancer Cells within 3D Tissue Matrix.

This study aimed to understand extracellular mechanical stimuli's effect on prostate cancer cells' metastatic progression within a three-dimensional (3D) bone-like microenvironment. In this study, a mechanical loading platform, EQUicycler, has been employed to create physiologically relevant static and cyclic mechanical stimuli to a prostate cancer cell (PC-3)-embedded 3D tissue matrix. Three mechanical stimuli conditions were applied: control (no loading), cyclic (1% strain at 1 Hz), and static mechanical stimuli (1% strain). The changes in prostate cancer cells' cytoskeletal reorganization, polarity (elongation index), proliferation, expression level of N-Cadherin (metastasis-associated gene), and migratory potential within the 3D collagen structures were assessed upon mechanical stimuli. The results have shown that static mechanical stimuli increased the metastasis progression factors, including cell elongation (p < 0.001), cellular F-actin accumulation (p < 0.001), actin polymerization (p < 0.001), N-Cadherin gene expression, and invasion capacity of PC-3 cells within a bone-like microenvironment compared to its cyclic and control loading counterparts. This study established a novel system for studying metastatic cancer cells within bone and enables the creation of biomimetic in vitro models for cancer research and mechanobiology.

Three-dimensional graphene foam based triboelectric nanogenerators for energy systems and autonomous sensors

In this work we investigate the potential of three-dimensional graphene (3DG) foam as an active layer in triboelectric nanogenerators (TENGs) and as an energy harvesting power source for autonomous sensors. A series of comprehensive measurements have been carried out to test the output characteristics of 3DG-TENG under cyclic mechanical stimulus, capable of operating TENG in contact-separation mode at different frequencies, gap distances between electrodes, and applied pressures. The triboelectric response of 3DG-TENG (with an effective surface of 16 cm2) showed maximum open-circuit voltage (Voc) and short-circuit current (Isc) of 400 V and 105.7 μA respectively when stimulated at 3 Hz (contact-separation frequency) and 70 mm (optimum gap distance). Under the same conditions, a maximum output power (Pout) of around 10.37 W/m2 is produced using an external load resistance of 40 MΩ; this is an order of magnitude lower resistance than that needed with other graphene based TENG variants. 3DG-TENG exhibited great stability in the output characteristics with 15,000 cyclic mechanical stimuli and a retention percentage in Pout above 95%. This is a significant improvement with respect to other carbon based TENG`s, which show enhanced deterioration of TENG performance due to material transfer between electrodes and plastic deformation of triboelectric materials. Simulations of TENG Voc using distance dependent model determined high triboelectric charge densities in the range of mC/m2. Here, we also demonstrate the potential of 3DG-TENG as an energy supply for energy storage devices, and as an active layer in an autonomous pressure sensing platform for anonymous room occupancy monitoring in smart buildings.

Location-Dependent Human Osteoarthritis Cartilage Response to Realistic Cyclic Loading: Ex-Vivo Analysis on Different Knee Compartments.

Objective: Osteoarthritis (OA) is a multifactorial musculoskeletal disorder affecting mostly weight-bearing joints. Chondrocyte response to load is modulated by inflammatory mediators and factors involved in extracellular cartilage matrix (ECM) maintenance, but regulatory mechanisms are not fully clarified yet. By using a recently proposed experimental model combining biomechanical data with cartilage molecular information, basally and following ex-vivo load application, we aimed at improving the understanding of human cartilage response to cyclic mechanical compressive stimuli by including cartilage original anatomical position and OA degree as independent factors. Methods: 19 mono-compartmental Knee OA patients undergoing total knee replacement were recruited. Cartilage explants from four different femoral condyles zones and with different degeneration levels were collected. The response of cartilage samples, pooled according to OA score and anatomical position was tested ex-vivo in a bioreactor. Mechanical stimulation was obtained via a 3-MPa 1-Hz sinusoidal compressive load for 45-min to replicate average knee loading during normal walking. Samples were analysed for chondrocyte gene expression and ECM factor release. Results: Non parametric univariate and multivariate (generalized linear mixed model) analysis was performed to evaluate the effect of compression and IL-1β stimulation in relationship to the anatomical position, local disease severity and clinical parameters with a level of significance set at 0.05. We observed an anti-inflammatory effect of compression inducing a significant downmodulation of IL-6 and IL-8 levels correlated to the anatomical regions, but not to OA score. Moreover, ADAMTS5, PIICP, COMP and CS were upregulated by compression, whereas COL-2CAV was downmodulated, all in relationship to the anatomical position and to the OA degree. Conclusion: While unconfined compression testing may not be fully representative of the in-vivo biomechanical situation, this study demonstrates the importance to consider the original cartilage anatomical position for a reliable biomolecular analysis of knee OA metabolism following mechanical stimulation.

Local mechanical stimuli correlate with tissue growth in axolotl salamander joint morphogenesis.

Movement-induced forces are critical to correct joint formation, but it is unclear how cells sense and respond to these mechanical cues. To study the role of mechanical stimuli in the shaping of the joint, we combined experiments on regenerating axolotl (Ambystoma mexicanum) forelimbs with a poroelastic model of bone rudiment growth. Animals either regrew forelimbs normally (control) or were injected with a transient receptor potential vanilloid 4 (TRPV4) agonist during joint morphogenesis. We quantified growth and shape in regrown humeri from whole-mount light sheet fluorescence images of the regenerated limbs. Results revealed significant differences in morphology and cell proliferation between groups, indicating that TRPV4 desensitization has an effect on joint shape. To link TRPV4 desensitization with impaired mechanosensitivity, we developed a finite element model of a regenerating humerus. Local tissue growth was the sum of a biological contribution proportional to chondrocyte density, which was constant, and a mechanical contribution proportional to fluid pressure. Computational predictions of growth agreed with experimental outcomes of joint shape, suggesting that interstitial pressure driven from cyclic mechanical stimuli promotes local tissue growth. Predictive computational models informed by experimental findings allow us to explore potential physical mechanisms involved in tissue growth to advance our understanding of the mechanobiology of joint morphogenesis.

3D Bioprinting of an In Vitro Model of a Biomimetic Urinary Bladder with a Contract-Release System.

The development of curative therapy for bladder dysfunction is usually hampered owing to the lack of reliable ex vivo human models that can mimic the complexity of the human bladder. To overcome this issue, 3D in vitro model systems offering unique opportunities to engineer realistic human tissues/organs have been developed. However, existing in vitro models still cannot entirely reflect the key structural and physiological characteristics of the native human bladder. In this study, we propose an in vitro model of the urinary bladder that can create 3D biomimetic tissue structures and dynamic microenvironments to replicate the smooth muscle functions of an actual human urinary bladder. In other words, the proposed biomimetic model system, developed using a 3D bioprinting approach, can recreate the physiological motion of the urinary bladder by incorporating decellularized extracellular matrix from the bladder tissue and introducing cyclic mechanical stimuli. The results showed that the developed bladder tissue models exhibited high cell viability and proliferation rate and promoted myogenic differentiation potential given dynamic mechanical cues. We envision the developed in vitro bladder mimicry model can serve as a research platform for fundamental studies on human disease modeling and pharmaceutical testing.

Spatiotemporal model of cellular mechanotransduction via Rho and YAP.

How cells sense and respond to mechanical stimuli remains an open question. Recent advances have identified the translocation of Yes-associated protein (YAP) between nucleus and cytoplasm as a central mechanism for sensing mechanical forces and regulating mechanotransduction. We formulate a spatiotemporal model of the mechanotransduction signalling pathway that includes coupling of YAP with the cell force-generation machinery through the Rho family of GTPases. Considering the active and inactive forms of a single Rho protein (GTP/GDP-bound) and of YAP (non-phosphorylated/phosphorylated), we study the cross-talk between cell polarization due to active Rho and YAP activation through its nuclear localization. For fixed mechanical stimuli, our model predicts stationary nuclear-to-cytoplasmic YAP ratios consistent with experimental data at varying adhesive cell area. We further predict damped and even sustained oscillations in the YAP nuclear-to-cytoplasmic ratio by accounting for recently reported positive and negative YAP-Rho feedback. Extending the framework to time-varying mechanical stimuli that simulate cyclic stretching and compression, we show that the YAP nuclear-to-cytoplasmic ratio's time dependence follows that of the cyclic mechanical stimulus. The model presents one of the first frameworks for understanding spatiotemporal YAP mechanotransduction, providing several predictions of possible YAP localization dynamics, and suggesting new directions for experimental and theoretical studies.

Glass Microchannel Formation by Mycelium

We propose a new method to fabricate complicated 3-dimensional glass microchannels. We employed mycelium for this purpose. Mycelium possesses a complicated, fine and three-dimensional network structure. We cultivated mycelium in silica compounds, and subsequently silica compounds were heated to be sintered. During this heating process, all the mycelium was burned off and remained a fine network channel structure in a transparent glass chip. We also tried to control of the growth of this mycelium. The growth could be changed by growth conditions. In this work, we used cyclic mechanical stimuli for this purpose. We set cyclic tensile strain to the sample under growing mycelium. This cyclic strain caused anisotropic growth of the mycelium in some condition.

Heterogeneous role of integrins in fibroblast response to small cyclic mechanical stimulus generated by a nanoporous gold actuator

It is important to understand the effects of mechanical stimulation on cell behaviors for homeostasis. Many studies have been performed on cell responses to mechanical stimuli, but the mechanosensing mechanism is still under debate. In the present study, experiments employing molecular dynamics (MD) simulations concerning the effects of cyclic mechanical stimulus on cell proliferation were performed based on the hypothesis that mechanosensing depends on integrin types. We used a nanoporous gold (NPG) actuator to prevent transfer of a mechanical stimulus via molecules other than integrins. Surprisingly, a small cyclic strain of only 0.5% enhanced the proliferation of fibroblasts. α5β1 and αvβ3 integrins showed high sensitivity to the mechanical stimulus, whereas α1β1 and α2β1 integrins exhibited low mechanosensitivity. The MD simulations showed that different conformational changes of the integrin headpiece induced by binding to the ECM led to a difference in mechanosensitivity between αI and αI-less integrin types. Thus, the present study provides evidence to support the hypothesis and suggests the mechanism for the heterogeneous roles of integrins in mechanosensing.

Combination of inductive effect of lipopolysaccharide and in situ mechanical conditioning for forming an autologous vascular graft in vivo

Autologous vascular grafts have the advantages of better biocompatibility and prognosis. However, previous studies that implanted bare polymer tubes in animals to grow autologous tubular tissues were limited by their poor yield rates and stability. To enhance the yield rate of the tubular tissue, we employed a design with the addition of overlaid autologous whole blood scaffold containing lipopolysaccharides (LPS). Furthermore, we applied in vivo dynamic mechanical stimuli through cyclically inflatable silicone tube to improve the mechanical properties of the harvested tissues. The effectiveness of the modification was examined by implanting the tubes in the peritoneal cavity of rats. A group without mechanical stimuli served as the controls. After 24 days of culture including 16 days of cyclic mechanical stimuli, we harvested the tubular tissue forming on the silicone tube for analysis or further autologous interposition vascular grafting. In comparison with those without cyclic dynamic stimuli, tubular tissues with this treatment during in vivo culture had stronger mechanical properties, better smooth muscle differentiation, and more collagen and elastin expression by the end of incubation period in the peritoneal cavity. The grafts remained patent after 4 months of implantation and showed the presence of endothelial and smooth muscle cells. This model shows a new prospect for vascular tissue engineering.

Incrementing the Frequency of Dynamic Strain on SMC-Cellularised Collagen-Based Scaffolds Affects Extracellular Matrix Remodeling and Mechanical Properties.

Notwithstanding the efforts injected in vascular tissue engineering in the past 30 years, the clinical translation of engineered artery constructs is far from being successful. One common approach to improve artery regeneration is the use of cyclic mechanical stimuli to guide cellular remodeling. However, there is a lack of information on the effect of cyclic strain on cells within a 3D environment. To this end, this work explored the effect of gradual increase in stimulation frequency on the response of human umbilical artery smooth muscle cells (HUASMCs) embedded in a 3D collagen matrix. The results demonstrate that, with an applied strain of 5%, the gradual increase of frequency from 0.1 to 1 Hz improved collagen remodeling by HUASMCs compared to samples constantly stimulated at 1 Hz. The expression of collagen, elastin and matrix metalloproteinase-2 (MMP-2) genes was similar at 7 days for gradual and 1 Hz samples which showed lower amounts than static counterparts. Interestingly the mechanical properties of the constructs, specifically the amplitude of the time constants and the elastic equilibrium modulus, were enhanced by gradual increase of frequency. Taken together, these results show an increase in collagen remodeling by the HUASMCs overtime under incremental cyclic mechanical strain. This work suggests that only the in-depth investigation of the effects of stimulation parameters on the behavior of vSMC under cyclic strain in a 3D environment could lead to the design of optimized control strategies for enhanced vascular tissue generation and maturation in bioreactors.

Three-Dimensional Mechanical Loading Modulates the Osteogenic Response of Mesenchymal Stem Cells to Tumor-Derived Soluble Signals.

Dynamic mechanical loading is a strong anabolic signal in the skeleton, increasing osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BM-MSCs) and increasing the bone-forming activity of osteoblasts, but its role in bone metastatic cancer is relatively unknown. In this study, we integrated a hydroxyapatite-containing three-dimensional (3D) scaffold platform with controlled mechanical stimulation to investigate the effects of cyclic compression on the interplay between breast cancer cells and BM-MSCs as it pertains to bone metastasis. BM-MSCs cultured within mineral-containing 3D poly(lactide-co-glycolide) (PLG) scaffolds differentiated into mature osteoblasts, and exposure to tumor-derived soluble factors promoted this process. When BM-MSCs undergoing osteogenic differentiation were exposed to conditioned media collected from mechanically loaded breast cancer cells, their gene expression of osteopontin was increased. This was further enhanced when mechanical compression was simultaneously applied to BM-MSCs, leading to more uniformly deposited osteopontin within scaffold pores. These results suggest that mechanical loading of 3D scaffold-based culture models may be utilized to evaluate the role of physiologically relevant physical cues on bone metastatic breast cancer. Furthermore, our data imply that cyclic mechanical stimuli within the bone microenvironment modulate interactions between tumor cells and BM-MSCs that are relevant to bone metastasis.

Controlling dynamic mechanical properties and degradation of composites for bone regeneration by means of filler content

Bone tissue is a dynamic composite system that adapts itself, in response to the surrounding daily (cyclic) mechanical stimuli, through an equilibrium between growth and resorption processes. When there is need of synthetic bone grafts, the biggest issue is to support bone regeneration without causing mechanically-induced bone resorption. Apart from biological properties, such degradable materials should initially support and later leave room to bone formation. Further, dynamic mechanical properties comparable to those of bone are required. In this study we prepared composites comprising calcium phosphate and L-lactide/D-lactide copolymer in various content ratios using the extrusion method. We evaluated the effect of the inorganic filler amount on the polymer phase (i.e. on the post-extrusion intrinsic viscosity). We then studied their in vitro degradation and dynamic mechanical properties (in dry and humid conditions). By increasing the filler content, we observed significant decrease of the intrinsic viscosity of the polymer phase during the extrusion process. Composites containing higher amounts of apatite had faster degradation, and were also mechanically stiffer. But, due to the lower intrinsic viscosity of their polymer phase, they had larger damping properties. Besides this, higher amounts of apatite also rendered the composites more hydrophilic letting them absorb more water and causing them the largest decrease in stiffness. These results show the importance of filler content in controlling the properties of such composites. Further, in this study we observed that the viscoelastic properties of the composite containing 50wt% apatite were comparable to those of dry human cortical bone.

Synergistic effects of cyclic tension and transforming growth factor-β1 on the aortic valve myofibroblast

Phenotypically, aortic valve interstitial cells are dynamic myofibroblasts, appearing contractile and activated in times of development, disease, and remodeling. The precise mechanism of phenotypic modulation is unclear, but it is speculated that both biomechanical and biochemical factors are influential. Therefore, we hypothesized that isolated and combined treatments of cyclic tension and transforming growth factor-beta1 would alter the phenotype and subsequent collagen biosynthesis of aortic valve interstitial cells in situ. Porcine aortic valve leaflets received 7- and 14-day treatments of 15% cyclic stretch (Tension); 0.5 ng/ml transforming growth factor-beta1 (TGF); 15% cyclic stretch and 0.5 ng/ml transforming growth factor-beta1 (Tension+TGF); or neither mechanical nor cytokine stimuli (Null). Tissues were homogenized and assayed for aortic valve interstitial cell phenotype (smooth muscle alpha-actin) and collagen biosynthesis (via heat shock protein 47, which was further verified with type I collagen C-terminal propeptide). At both 7 and 14 days, smooth muscle alpha-actin, heat shock protein 47, and type I collagen C-terminal propeptide quantities were significantly greater (P<.001) in the Tension+TGF group than in all other groups. Additionally, Tension alone appeared to maintain smooth muscle alpha-actin and heat shock protein 47 levels that were measured on Day 0, while TGF alone elicited an increase in smooth muscle alpha-actin and heat shock protein 47 compared to Day 0 levels. Null treatment revealed diminished proteins at both time points. Elevated transforming growth factor-beta1 levels, in the presence of cyclic mechanical tension, resulted in synergistic increases in contractile and biosynthetic proteins in aortic valve interstitial cells. Since cyclic mechanical stimuli can never be relieved in vivo, the presence of transforming growth factor-beta1 (possibly from infiltrating macrophages) may result in overly biosynthetic aortic valve interstitial cells, leading to altered extracellular matrix architecture, compromised valve function, and, ultimately, degenerative valvular disease.

Identification of Pip4k2β as a mechanical stimulus responsive gene and its expression during musculoskeletal tissue healing

To investigate the mechano-transduction system of cells, we identified genes responsive to a cyclic mechanical stimulus. MC3T3.E1 cells were cultured on a computer-controlled vacuum-pump-operated device designed to provide a cyclic mechanical stimulus. A maximum elongation of 15% of membrane at 10 cycles/min (3 s extension followed by 3 s relax per cycle) was repeated for 48 h. By means of a differential display, the gene expression pattern of cells exposed to the stimulus was compared with that of unexposed cells. As a result, a gene fragment that was exclusively expressed in mechanically stressed cells was identified. By using expressed sequence tag walking together with the oligo-capping method, this gene was identified as phosphatidylinositol 4-phosphate 5-kinase type II beta (initially known as Pip5k2beta but now reclassified as Pip4k2beta). The specific up-regulation of Pip4k2beta upon mechanical stimulus was also confirmed by using another apparatus, viz. a computer-controlled linearized-stepping motor system. To examine the involvement of the cyclic mechanical stimulus in the regulation of Pip4k2beta expression in musculoskeletal tissue, we created an Achilles tendon transection model in rabbits. The temporal expression of Pip4k2beta was assessed by means of a quantitative reverse-transcribed polymerase chain reaction. In the gastrocnemius muscle, expression of Pip4k2beta rapidly decreased 1 week after transection but was restored to normal levels at 4 weeks. In the Achilles tendon, however, expression remained decreased until 4 weeks after transection. We suggest that the expression of Pip4k2beta can be used as a marker for cells receiving a suitable mechanical stimulus.

Physical aging time scales and rates for poly(vinyl acetate) stimulated mechanically in the Tg‐region

AbstractThe physical aging behavior of poly(vinyl acetate), PVAc, in the Tg‐region has been investigated using dynamic mechanical analysis (DMA). Tests in shear and flexure modes were performed for samples quenched from a stabilization temperature of 40°C to aging temperatures 35.0°C and 32.5°C (down‐jumps), and from equilibrium at a stabilization temperature of 32.5°C to aging temperature 36.2°C (up‐jump). Volume recovery was investigated for the same thermal histories using mercury‐in‐glass dilatometry. The time scales for the evolution of the storage and loss moduli during physical aging have been compared with the volumetric time scales. Aging rates, µ, have been calculated in the frequency and time domains. The evolution of dynamic mechanical properties during aging is found to be different from the volumetric behavior. In the case of the temperature down‐jump, the evolution of the volume ceases earlier than that for the storage and loss moduli. In the up‐jump experiment, the opposite is found. Aging rates for the storage and loss moduli are analyzed. A suggestion is offered that the evolution of dynamic mechanical properties during physical aging is retarded by the cyclic mechanical stimulus, and that this effect (rejuvenation) is more noticeable the higher is the damping; consequently rejuvenation in the Tg‐region can be caused by small strains in dynamic mechanical analysis.

Orientation of cultured arterial smooth muscle cells growing on cyclically stretched substrates.

Arterial smooth muscle cells from rabbit aortic media were grown in first subcultures on hydrophilized and collagen-coated silicone membranes which were then subjected to directional cyclic stretches and relaxations at a frequency of 50 times/min. The membranes were stretched 2, 5 and 10% beyond their resting length. Cells on unstretched and stationary membranes in the same chamber served as controls. The cells which were stretched with an amplitude of 2% remained in random orientation after 14 days of continuously performed cyclic stretching. The cells which were stretched 5% for 12 days orientated at an angle of 61 +/- 9 degrees to the direction of stretching, while the cells which were stretched with an amplitude of 10% for 6 days orientated at an angle of 76 +/- 5 degrees. The cells on the stationary and unstretched membranes remained in random orientation. We were able to confirm that the angle of orientation is reversible, i.e. preorientated cells changed their orientation during application of another stretching amplitude. The results suggest that stretching of the artery wall by blood pulsation may be a factor influencing the orientation of smooth muscle cells within the media of the artery wall and of those smooth muscle cells which proliferate into the subendothelial space after mechanical injury of the endothelium or electrical stimulation of the artery wall. An apparatus is presented which produces cyclic and directional mechanical stimuli similar to those which may occur in the artery wall.

A new in vitro system for studying cell response to mechanical stimulation: Different effects of cyclic stretching and agitation on smooth muscle cell biosynthesis

An experimental model has been devised to permit morphologic and metabolic characterization of cells subjected to a range of cyclic mechanical stimuli similar to those which may prevail in blood vessel walls. A unique feature is the use of purified elastin membranes prepared from bovine aortas as extensible substrates for cell growth. Cells attached firmly to such membranes which could then be subjected to continuous stretching and relaxation or displacement without stretching by a motor coupled to a movable supporting frame. Various combinations of frequencies, amplitudes and rates of deformation have been used over extended periods with minimal fatigue or disruption of the elastin substrate. The effects of cyclic stretching on [ 14C]proline incorporation into protein and collagen and [ 3H]thymidine incorporation into DNA by rabbit aortic smooth muscle cells were distinct from those attributable to agitation without stretching, indicating that cells responded differently to these modes of stimulation. Increases in rate of protein or DNA synthesis induced by stretching were just as marked after 48 h of stimulation as they were at the outset of an experimental period. Since the system permits observations of cell response to independently variable components of pulsatile stress over extended periods and under a variety of culture conditions, it may be expected to provide new data concerning the interaction of mechanical with hormonal and genetic factors in the elaboration of connective tissue components.