Reduction of Cardiac Fibrosis by Interference With YAP-Dependent Transactivation.

BACKGROUND: Conversion of cardiac stromal cells into myofibroblasts is typically associated with hypoxia conditions, metabolic insults, and/or inflammation, all of which are predisposing factors to cardiac fibrosis and heart failure. We hypothesized that this conversion could be also mediated by response of these cells to mechanical cues through activation of the Hippo transcriptional pathway. The objective of the present study was to assess the role of cellular/nuclear straining forces acting in myofibroblast differentiation of cardiac stromal cells under the control of YAP (yes-associated protein) transcription factor and to validate this finding using a pharmacological agent that interferes with the interactions of the YAP/TAZ (transcriptional coactivator with PDZ-binding motif) complex with their cognate transcription factors TEADs (TEA domain transcription factors), under high-strain and profibrotic stimulation. METHODS: We employed high content imaging, 2-dimensional/3-dimensional culture, atomic force microscopy mapping, and molecular methods to prove the role of cell/nuclear straining in YAP-dependent fibrotic programming in a mouse model of ischemia-dependent cardiac fibrosis and in human-derived primitive cardiac stromal cells. We also tested treatment of cells with Verteporfin, a drug known to prevent the association of the YAP/TAZ complex with their cognate transcription factors TEADs. RESULTS: Our experiments suggested that pharmacologically targeting the YAP-dependent pathway overrides the profibrotic activation of cardiac stromal cells by mechanical cues in vitro, and that this occurs even in the presence of profibrotic signaling mediated by TGF-ß1 (transforming growth factor beta-1). In vivo administration of Verteporfin in mice with permanent cardiac ischemia reduced significantly fibrosis and morphometric remodeling but did not improve cardiac performance. CONCLUSIONS: Our study indicates that preventing molecular translation of mechanical cues in cardiac stromal cells reduces the impact of cardiac maladaptive remodeling with a positive effect on fibrosis.

Mechanical properties of a new thermally deformable mitral valve annuloplasty ring and its effects on the mitral valve.

Ideally, an annuloplasty ring's shape should be changed intraoperatively if mitral valve repair is unsuccessful because of a short coaptation length or systolic anterior motion. Several post-implantation adjustable rings have been developed, but they are not freely deformable and are unsuitable for asymmetric repair of the valvular annulus. We developed a novel thermally deformable mitral annuloplasty ring to address these problems and assessed the ring's mechanical properties and its effect on the mitral valve anatomy. This ring was made of polycaprolactone. Tensile and bending tests were performed to evaluate the ring's mechanical properties. The ratio of the transverse and septal-lateral length was determined as 4:3. Using 10 pig hearts, we measured the post-deformation coaptation length and minimum distance from the coaptation to the ventricular septum, which is a factor of abnormal systolic anterior motion of the mitral valve. In the mechanical tests, the ring's yield point was greater than the deformation force of the annulus in humans. In pigs with deformation from "4:3" to "4:2", the coaptation length was significantly increased in each mitral valve part. In pigs with deformation from "4:3" to "4:4", the minimum distance from the coaptation to the ventricular septum was significantly increased. Asymmetrical ring deformation increased the coaptation length only at the deformed area. In conclusion, this new thermally deformable mitral annuloplasty ring could be "order-made" to effectively change the coaptation length in all parts of the mitral valve and the distance from the coaptation to septum post-deformation via intraoperative heating.

Temporal regulation of gap junctional communication between tenocytes subjected to static tensile strain with physiological and non-physiological amplitudes.

The present study has been performed on temporal changes in gap junctional intercellular communication (GJIC) between tenocytes under static tensile strain with the magnitude of 0% (no strain), 4% (physiological magnitude) or 8% (overloading magnitude) during a 24-h culture period. Tenocytes were isolated from rabbit Achilles tendon and seeded on a stretchable microgroove substrate. GJIC was evaluated as intercellular diffusion coefficient of calcein (DGJ) using fluorescence loss in photobleaching (FLIP) protocol accompanied with a mathematical model of molecular diffusion both within the cell and between the cells. It was exhibited that the application of 4% strain for 1 h increased DGJ significantly. The increased level was maintained for 6 h, followed by returning to the pre-strain level at 24 h. This was associated with a transient increase in connexin 43 (Cx43) gene expression and protein localisation at 1 h, suggesting the increased GJIC may have involved new synthesis of gap junctions. By contrast, the application of 8% static strain reduced DGJ to the similar or lower level from 0% strain group for 6 h, associated with inhibited Cx43 gene expression. However, Cx43 protein localisation was not changed much, and thus, there seem no direct interactions among changes in GJIC, Cx43 gene expression and Cx43 localisation. The present findings highlight the differences in mechanical regulation of GJIC between physiological and non-physiological loadings, and thus the increase or the decrease in GJIC may affect tenocyte functions in different ways.

Mechano-regulation of gap junction communications between tendon cells is dependent on the magnitude of tensile strain.

Large magnitudes of mechanical strain applied to tendon cells induce catabolic and inflammatory responses, whereas a moderate level of strain promotes anabolism. Gap junction intercellular communication (GJIC) plays an essential role in these responses, however direct regulation of GJIC by mechanical loading has not been characterised in detail. Here, we show that the GJIC between tenocytes are enhanced or inhibited depending on the magnitude of the tensile strain. The GJIC was analysed using fluorescence loss in photobleaching (FLIP), combined with a molecular diffusion model. Intercellular and intracellular transport of fluorescence tracer molecules, calcein, across multiple cells through the gap junctions was evaluated by determining the intercellular and intracellular diffusion coefficients of calcein. It was demonstrated that the intercellular diffusion coefficient was significantly higher when the cells were subjected to a physiological static tensile strain (4%) for 1 h, but significantly lower when subjected to a strain with non-physiological amplitude (8%). The intracellular diffusion coefficient was not altered by the application of static strain at any level. Connexin 43 proteins were localised within cytoplasm and at cell-cell boundaries in no strained state and were also localised near cell nuclei by the 4% strain, but the localisation was reduced by the 8% strain. The findings suggest that the increase in GJIC in response to 4% strain involves opening of gap junction pores via mechanotransduction events of tenocytes, whereas the inhibition in response to 8% strain involves mechanical disruption of the junctions.

A literature review of biomechanical studies on physiological and pathological sacroiliac joints: Articular surface structure, joint motion, dysfunction and treatments.

BACKGROUND: Sacroiliac joints are affected by mechanical environments; the joints are formed under mechanical stimulation, receive impact of walking between the upper and lower parts of the bodies and can be a cause of pain due to non-physiological loads. However, there are so far very few studies that reviewed biomechanics of physiological and pathological sacroiliac joints. This review article aims to describe the current sacroiliac joint biomechanics. METHODS: Previous original papers have been summarized based on three categories: articular surface structure, sacroiliac joint motion and sacroiliac joint dysfunction and treatments. FINDINGS: Although the articular surface morphologies vary greatly from individual to individual, many researchers have tried to classify the joints into several types. It has been suggested that the surface morphologies may not change regardless of joint dysfunction, however, the relationship between the joint structure and pain are still unclear. The range of sacroiliac joint motion is demonstrated to be less than 1 mm and there is no difference between physiological and pathological joints. The sacroiliac joint absorbs shock within the pelvis by the joint structures of pelvic morphology, ligaments and fat tissues. The morphology and motion of the sacroiliac joints may be optimized for upright bipedal walking. INTERPRETATION: There is no doubt that pelvic mechanical environments affect pain induction and treatment; however, no one has yet provided a concrete explanation. Future research could help develop treatments based on sacroiliac joint biomechanics to support joint function.

Experimental characterization of motion resistance of the sacroiliac joint.

BACKGROUND: The human sacroiliac joint (SIJ) in vivo is exposed to compressive and shearing stress environment, given the joint lines are almost parallel to the direction of gravity. The SIJ supports efficient bipedal walking. Unexpected or unphysiological, repeated impacts are believed to cause joint misalignment and result in SIJ pain. In the anterior compartment of the SIJ being synovial, the articular surface presents fine irregularities, potentially restricting the motion of the joints. OBJECTIVE: To clarify how the SIJ articular surface affects the resistance of the motion under physiological loading. METHODS: SIJ surface models were created based on computed tomography data of three patients and subsequently 3D printed. Shear resistance was measured in four directions and three combined positions using a customized setup. In addition, repositionability of SIJs was investigated by unloading a shear force. RESULTS: Shear resistance of the SIJ was the highest in the inferior direction. It changed depending on the direction of the shear and the alignment position of the articular surface. CONCLUSION: SIJ articular surface morphology is likely designed to accommodate upright bipedal walking. Joint misalignment may in consequence increase the risk of subluxation.

A new experimental system for simultaneous application of cyclic tensile strain and fluid shear stress to tenocytes in vitro.

Tenocyte mechanotransduction has been of great interest to researchers in tendon mechanobiology and biomechanics. In vivo, tenocytes are subjected to tensile strain and fluid shear stress, but most studies of tenocyte mechanobiology have been to understand how tenocytes regulate their functions in response to tensile strain. Thus, there is still much to know about tenocyte responses to fluid shear stress, partly due to the difficulty of devising a suitable experimental set-up and understanding the exact magnitude of imposed fluid shear stress. Therefore, this study was performed to test a new experimental system, which is suitable for the application of tensile strain and fluid shear stress to tenocytes in vitro. It was experimentally and numerically confirmed that tenocytes could maintain their in situ morphology within microfabricated microgrooves; also, physiological tensile strain and a wide range of fluid shear stress magnitudes can be applied to these cells. Indeed, it was demonstrated that the combined stimulation of cyclic tensile strain and oscillatory fluid shear stress induced a greater synergetic effect on tenocyte calcium response and significantly increased the percentage of tenocyte exhibiting increases in intracellular Ca(2+) concentration compared to the solo applications of these two modes of mechanical stimulation. The experimental system presented here is suitable for research of tenocyte mechanobiology, particularly mechanotransduction events, which were difficult to study using previous experimental models like explants and cell monolayers.

Numerical analysis of the effects of padded pelvic belts as a treatment for sacroiliac joint dysfunction.

BACKGROUND: Pain related to the sacroiliac joint (SIJ) accounts for low back pain in 15%-30% of patients. One of the most common treatment options is the use of pelvic belts. Various types of pelvic belts exist; however, the mechanisms underlying treatment and their effectiveness remain unclear to date. OBJECTIVE: To analyze stress distribution in the pelvis when a pelvic rubber belt or a padded pelvic belt is applied, to assess the effectiveness of treatment from a numerical biomechanical perspective. METHODS: The pressure distribution at the pelvic belts was measured using a device and subsequently modeled with the finite element method of a pelvis with soft tissues. The stress environment when wearing a pelvic belt in a double-leg stance was simulated. RESULTS: With the application of pelvic belts, the innominate bone rotated outward, which was termed an out-flare. This caused the SIJ to compress and cause reduction in sacrotuberous, sacrospinous, interosseous, and posterior sacroiliac ligament loading. Padded pelvic belts decreased the SIJ displacement to a greater extent than in pelvic rubber belts. CONCLUSION: Pelvic belts aid in compressing the SIJ and reduce its mobility.

Identification of leader cells by filopodia in collective cell migration using computer vision.

BACKGROUND: Imaging of cells and cellular organelles has been of great interest among researchers and medical staff because it can provide useful information on cell physiology and pathology. Many researches related to collective cell migration have been established and leader cells seem to be the ones that regulate the migration, however, the identification of leader cells is very time-consuming. OBJECTIVE: This study utilized computer vision with deep learning to segment cell shape and to identify leader cells through filopodia. METHODS: Healthy Madin-Darby Canine Kidney (MDCK) cells cultured in a Polydimethylsiloxane (PDMS) microchannel device allowed collective cell migration as well as the formation of leader cells. The cells were stained, and cell images were captured to train the computer using UNet++ together with their corresponding masks created using Photoshop for automated cell segmentation. Lastly, cell shape and filopodia were filtered out using Filopodyan and FiloQuant were detected. RESULTS: The segmentation of cell shape and the identification of filopodia were successful and produced accurate results in less than one second per image. CONCLUSIONS: The proposed approach of image analysis would be a great help in the field of cell science, engineering, and diagnosis.

A patient-cohort study of numerical analysis on sacroiliac joint stress distribution in pre- and post-operative hip dysplasia.

In acetabular dysplasia, the cartilaginous roof on the acetabular side does not fully cover the femoral head, which may lead to abnormal stress distribution in both the femoral head and pelvis. These stress changes may have implications to the adjacent sacroiliac joint (SIJ). The SIJ has a minimal range of motion and is closely coupled to the adjacent spine and pelvis. In consequence, the SIJ may react sensitively to changes in stress distribution at the acetabulum, with hypermobility-induced pain. The purpose of this study was to investigate the stress distribution of the SIJ in acetabular dysplasia, and to gain insight into the cause and mechanisms of hypermobility-induced pain at the SIJ. Finite element models of pre- and postoperative pelves of four patients with acetabular dysplasia were created and analyzed in double leg standing positions. The preoperative models were relatively inflare, the sacral nutation movement, SIJ cartilage equivalent stress, and the load on the surrounding ligaments decreased with increased posterior acetabular coverage. Acetabular morphology was shown to affect the SIJ, and improvement of the posterior acetabular coverage may help normalize load transmission of the pelvis and thus improve the stress environment of the SIJ in acetabular dysplasia.

Combined stimulation with cyclic stretching and hypoxia increases production of matrix metalloproteinase-9 and cytokines by macrophages.

Macrophages in the vessel wall of advanced abdominal aortic aneurysms (AAAs) are subjected to cyclic stretching and hypoxia because of pulsatile blood flow and intraluminal thrombi, respectively. It is possible that these conditions induce abnormal changes in macrophage functions, such as increased production of matrix metalloproteinase-2 (MMP-2), MMP-9, and inflammatory cytokines, leading to weakening of the aortic wall through excessive extracellular matrix disruption. Here we show the effects of cyclic stretching and hypoxia on the production of MMP-9 and inflammatory cytokines by macrophages. Gelatin zymography revealed that MMP-9 production by macrophages was significantly increased by 5% and 10% cyclic stretching under hypoxia (2.2% O(2)). Using enzyme-linked immunosorbent assay, we also evaluated the production of 12 different inflammatory cytokines and found that there was a tendency toward higher expressions of interleukin-8 and tumor necrosis factor-α by macrophages subjected to 10% cyclic stretching under normoxia and hypoxia. Next, we evaluated apoptosis of smooth muscle cells (SMCs) in medium conditioned by macrophages cultured under the 2 conditions described above. SMC apoptosis increased significantly when exposed to media harvested from macrophages subjected to 10% cyclic stretching under normoxia and hypoxia. On the basis of these results, we believe that macrophages produce cytokines that induce SMC apoptosis. Our results suggest that the combination of cyclic stretching and hypoxia stimulates MMP-9 and cytokine production in macrophages, which may result in weakening of AAA walls.

Mechanical Properties of Isolated Primary Cilia Measured by Micro-tensile Test and Atomic Force Microscopy.

Mechanotransduction is a well-known mechanism by which cells sense their surrounding mechanical environment, convert mechanical stimuli into biochemical signals, and eventually change their morphology and functions. Primary cilia are believed to be mechanosensors existing on the surface of the cell membrane and support cells to sense surrounding mechanical signals. Knowing the mechanical properties of primary cilia is essential to understand their responses, such as sensitivity to mechanical stimuli. Previous studies have so far conducted flow experiments or optical trap techniques to measure the flexural rigidity EI (E: Young's modulus, I: second moment of inertia) of primary cilia; however, the flexural rigidity is not a material property of materials and depends on mathematical models used in the determination, leading to a discrepancy between studies. For better characterization of primary cilia mechanics, Young's modulus should be directly and precisely measured. In this study, the tensile Young's modulus of isolated primary cilia is, for the first time, measured by using an in-house micro-tensile tester. The different strain rates of 0.01-0.3 s-1 were applied to isolated primary cilia, which showed a strain rate-dependent Young's modulus in the range of 69.5-240.0 kPa on average. Atomic force microscopy was also performed to measure the local Young's modulus of primary cilia, showing the Young's modulus within the order of tens to hundreds of kPa. This study could directly provide the global and local Young's moduli, which will benefit better understanding of primary cilia mechanics.

Quantitative evaluation of the sacroiliac joint fixation in stress reduction on both sacroiliac joint cartilage and ligaments: A finite element analysis.

BACKGROUND: The sacroiliac joint fixation is the last resort for patients with prolonged and severe joint pain. Although the clinical results of anterior fixations are conclusive, there exist several inevitable drawbacks with the surgical method such as the difficulty performing the surgery due to the presence of many organs. The posterior fixation technique has thus been developed to overcome those inconveniences. This study aims to assess in silico the mechanical environment following posterior and anterior fixations, focusing on stresses in both the sacroiliac cartilage and dorsal ligamentous part, as well as loads experienced by the pelvic ligaments. METHODS: Sacroiliac joint cartilage, dorsal ligamentous part stresses and pelvic ligaments loads were evaluated with three types of fixation models. A vertical load of 600 N was applied, equally distributed via both acetabula when standing and sitting. FINDINGS: Results show that the anterior sacroiliac joint fixation reduced von Mises stresses in the cartilage and dorsal ligamentous part and decreased ligaments loads more extensively than the posterior fixation when compared to the untreated model as a reference. However, the posterior fixation still remains the desirable and preferential treatment. INTERPRETATION: The anterior sacroiliac joint fixation showed better performances compared to the posterior one; however, the lower invasive aspect of the latter is a fundamental clinical advantage which also has the possibility to be improved by considering various screws and cages configurations. This study provides a beneficial suggestion to improve the current fixation technique.

Size sorting of kinesin-driven microtubules with topographical grooves on a chip.

Gliding microtubules (MTs) on a surface coated with kinesin biomolecular motors have been suggested for the development of nanoscale transport systems. In order to establish a sorting function for gliding MTs, events for MTs approaching micro-scale grooves were investigated. MTs longer than the width of grooves fabricated on a Si substrate bridged the grooves (bridging) and many MTs shorter than the groove width almost began to bridge, but returned to the surface that they approached from (guiding). Occurrence probabilities for the events were analyzed with focus on the geometric conditions, such as length of the MTs, width of the grooves, and the incident angle (alpha) of the MTs approaching the grooves. The occurrence probability for bridging increased with an increase in the incident angle (16%, alpha = 0-30 degrees; 51%, alpha = 30-60 degrees; 75%, alpha = 60-90 degrees), and the probability for guiding decreased with an increase in the incident angle (79%, alpha = 0-30 degrees; 55%, alpha = 30-60 degrees; 5%, alpha = 60-90 degrees). The results indicate that an incident angle of 30-60 degrees is an effective condition for MT sorting, because the bridging and guiding events can sort MTs that are longer and shorter than the groove widths, respectively. Furthermore, the occurrence probabilities of both bridging and guiding in a higher concentration of methylcellulose (0.5%) increased up to approximately 70% at incident angles of 30-60 degrees, indicating good feasibility for the development of devices for the sorting of MTs on surfaces with topographical grooves.

Effect of spatial gradient in fluid shear stress on morphological changes in endothelial cells in response to flow.

Arterial bifurcations are common sites for development of cerebral aneurysms. Although this localization of aneurysms suggests that high shear stress (SS) and high spatial SS gradient (SSG) occurring at the bifurcations may be crucial factors for endothelial dysfunction involved in aneurysm formation, the details of the relationship between the hemodynamic environment and endothelial cell (EC) responses remain unclear. In the present study, we sought morphological responses of ECs under high-SS and high-SSG conditions using a T-shaped flow chamber. Confluent ECs were exposed to SS of 2-10Pa with SSG of up to 34Pa/mm for 24 and 72h. ECs exposed to SS without spatial gradient elongated and oriented to the direction of flow at 72h through different processes depending on the magnitude of SS. In contrast, cells did not exhibit preferred orientation and elongation under the combination of SS and SSG. Unlike cells aligned to the flow by exposure to only SS, development of actin stress fibers was not observed in ECs exposed to SS with SSG. These results indicate that SSG suppresses morphological changes of ECs in response to flow.

Role of p120-catenin in the morphological changes of endothelial cells exposed to fluid shear stress.

p120-Catenin is known to play important roles in cell-cell adhesion stability by binding to cadherin and morphological changes of cells by regulating small RhoGTPase activities. Although the expression and binding states of p120-catenin are thought to dynamically change due to morphological adaptation of endothelial cells (ECs) to fluid shear stress, these dynamics remain to be explored. In the present study, we examined the time course of changes in p120-catenin expression and its binding to vascular endothelial (VE)-cadherin in ECs exposed to shear stress. Human umbilical vein ECs began to change their morphologies at 3-6h, and became elongated and oriented to the direction of flow at 24h after exposure to a shear stress of 1.5Pa. Binding and co-localization of p120-catenin with VE-cadherin at the foci of cell-cell adhesions were retained in ECs during exposure to shear stress, indicating that VE-cadherin was stabilized in the plasma membrane. In contrast, cytoplasmic p120-catenin that was dissociated from VE-cadherin was transiently increased at 3-6h after the flow onset. These results suggest that the transient increase of cytoplasmic p120-catenin may stimulate RhoGTPase activities and act as a switch for the morphological changes in ECs in response to shear stress.

Modification of the descending thoracic aortic anastomotic site using biodegradable felt: study in a canine model with or without basic fibroblast growth factor.

OBJECTIVES: We investigated the outcomes of reinforcing anastomotic sites using (1) nonbiodegradable polytetrafluoroethylene (PTFE) felt, (2) biodegradable polyglycolic acid (PGA) felt, and (3) PGA felt with basic fibroblast growth factor (bFGF) in a canine descending thoracic aortic replacement model. METHODS: Thirty-seven beagles underwent descending thoracic aorta replacement using a prosthetic graft with one of the above-mentioned reinforcements or no reinforcement for controls. Histologic evaluations were carried out 1 month and 3 months after surgery. The biomechanical strength of the anastomosis was assessed along the longitudinal axis of the aortic segments using a tensile tester. Local compliance at the anastomotic site was also evaluated in the circumferential direction. RESULTS: The media was significantly thinner in the PTFE group than in the control group (65.8% +/- 5.1% vs 95.0% +/- 9.3% of normal thickness; P < .05). Relative to the control group, the adventitial layer was significantly thinner in the PTFE group (42.3% +/- 8.2% of control; P < .05) but significantly thicker in the PGA and the PGA + bFGF groups (117.2% +/- 11.3% and 134.1% +/- 14.2% of control, respectively; P < .05). There were more vessels in the adventitial layer in the PGA + bFGF group than in the control, PTFE, and PGA groups (29.2 +/- 2.1/mm(2) vs 13.8 +/- 0.8, 5.4 +/- 0.7, 17.0 +/- 1.3/mm(2), respectively; P < .01). There were no significant differences between the four groups in the failure force at anastomotic sites. Local compliance at the anastomotic site was higher in the PGA group than that in the PTFE group (11.6 +/- 1.6 10(-6) m(2)/N vs 5.6 +/- 1.9 10(-6) m(2)/N; P < .05). CONCLUSION: Reinforcement of the experimental aortic wall with PTFE felt resulted in thinning of the media and adventitia and fewer vessels at the anastomotic site. These histologic changes were not observed when biodegradable felt was used. The bFGF failed to augment the modification of the aortic wall with the exception of increased adventitial vessel number. Biomechanical strength of the anastomosis along the longitudinal axis was comparable in all four groups; however, local vascular compliance was better in the biodegradable PGA felt group.

Effect of Geometric Curvature on Collective Cell Migration in Tortuous Microchannel Devices.

Collective cell migration is an essential phenomenon in many naturally occurring pathophysiological processes, as well as in tissue engineering applications. Cells in tissues and organs are known to sense chemical and mechanical signals from the microenvironment and collectively respond to these signals. For the last few decades, the effects of chemical signals such as growth factors and therapeutic agents on collective cell behaviors in the context of tissue engineering have been extensively studied, whereas those of the mechanical cues have only recently been investigated. The mechanical signals can be presented to the constituent cells in different forms, including topography, substrate stiffness, and geometrical constraint. With the recent advancement in microfabrication technology, researchers have gained the ability to manipulate the geometrical constraints by creating 3D structures to mimic the tissue microenvironment. In this study, we simulate the pore curvature as presented to the cells within 3D-engineered tissue-scaffolds by developing a device that features tortuous microchannels with geometric variations. We show that both cells at the front and rear respond to the varying radii of curvature and channel amplitude by altering the collective migratory behavior, including cell velocity, morphology, and turning angle. These findings provide insights into adaptive migration modes of collective cells to better understand the underlying mechanism of cell migration for optimization of the engineered tissue-scaffold design.

Finite element analysis of load transition on sacroiliac joint during bipedal walking.

The sacroiliac joint (SIJ) is burdened with variant loads. However, no methods have allowed to measure objectively how the SIJ deforms during bipedal walking. In this study, in-vivo walking conditions were replicated in a kinematic model combining the finite element method with 3D walking analysis data divided into five phases in order to visualize the load transition on the SIJ and clarify the role of the SIJ. Both models with and without inclusion of the SIJ were investigated. In models with bilateral SIJs, the displacement differed greatly between the sacrum and both hip bones on the SIJ as the boundary. The movements of the sacrum involved a nutation movement in the stance phase and a counter-nutation in the swing phase relative to the ilium. In models without SIJs, the displacement of the pelvis and loads of pelvic ligaments decreased, and the equivalent stress of the SIJs increased compared to the model with SIJs. The walking loads cause distortion of the entire pelvis, and stress concentration at the SIJ are seen due to the morphology of the pelvic ring. However, the SIJs help dissipate the resulting stresses, and the surrounding ligaments are likewise involved in load transmission.

A microwell array device with integrated microfluidic components for enhanced single-cell analysis.

Increasing awareness of the importance of cell heterogeneity in many biological and medical contexts is prompting increasing interest in systems that allow single-cell analysis rather than conventional bulk analysis (which provides average values for variables of interest from large numbers of cells). Recently, we presented a microwell chip for long-term, high-throughput single-cell analysis. The chip has proved to be useful for purposes such as screening individual cancer and stem cells for protein/gene markers. However, liquids in the wells can only be added or changed by manually rinsing the chip, or parts of it. This procedure has several well-known drawbacks--including risks of cross-contamination, large dead volumes and laboriousness--but there have been few previous attempts to integrate liquid rinsing/switching channels in "ready-to-use" systems for single-cell analysis. Here we present a microwell system designed (using flow simulations) for single-cell analysis with integrated microfluidic components (microchannels, magnetically driven micropumps and reservoirs) for supplying the cell culture wells with reagents, or rinsing, thus facilitating controlled, directed liquid handling. It can be used totally independently, since tubing is not essential. The practical utility of the integrated system has been demonstrated by culturing endothelial cells in the microwells, and successfully applying live-cell Calcein AM staining. Systems such as this combining high-density microwell chips with microfluidic components have great potential in numerous screening applications, such as exploring the important, but frequently undetected, heterogeneity in drug responses among individual cells.