Biomechanical Phenotypes Research Articles

Biomechanical defects and rescue of cardiomyocytes expressing pathologic nuclear lamins.

Given the clinical impact of LMNA cardiomyopathies, understanding lamin function will fulfill a clinical need and will lead to advancement in the treatment of heart failure. A multidisciplinary approach combining cell biology, atomic force microscopy (AFM), and molecular modeling was used to analyse the biomechanical properties of human lamin A/C gene (LMNA) mutations (E161K, D192G, N195K) using an in vitro neonatal rat ventricular myocyte model. The severity of biomechanical defects due to the three LMNA mutations correlated with the severity of the clinical phenotype. AFM and molecular modeling identified distinctive biomechanical and structural changes, with increasing severity from E161K to N195K and D192G, respectively. Additionally, the biomechanical defects were rescued with a p38 MAPK inhibitor. AFM and molecular modeling were able to quantify distinct biomechanical and structural defects in LMNA mutations E161K, D192G, and N195K and correlate the defects with clinical phenotypic severity. Improvements in cellular biomechanical phenotype was demonstrated and may represent a mechanism of action for p38 MAPK inhibition therapy that is now being used in human clinical trials to treat laminopathies.

Stiff vessels approached in a flexible way: Advancing quantification and interpretation of arterial stiffness

IntroductionAlthough pulse wave velocity (PWV), a proxy of arterial stiffness, is a strong predictor of cardiovascular complications, it is confounded by blood pressure (BP) and heart rate at the time of examination. Furthermore, establishing whether an artery behaves stiffer or less stiff does not inform a clinician on the cause of the stiffening. Quantification of arterial stiffnessThis paper focuses on BP as a confounder of PWV. We developed a method to patient-specifically determine the dependence of PWV on BP — on average 1 m/s per 10 mmHg diastolic BP — and used it to disentangle BP-dependent and -independent stiffening in hypertension and cancer patients. We furthermore showed that the so-called cardio-ankle vascular index (CAVI) — a measure deemed BP-independent — shows a residual BP-dependence that is readily correctable using a modified equation (CAVI0). Both developed methods are directly applicable to clinical measurements in individual patients. Interpretation of arterial stiffnessWe developed a computer modelling procedure to disentangle contributions of the individual wall components — collagen, elastin, and smooth muscle — to arterial stiffening as observed in patients. Our model-based approach shows that with ageing, the biomechanical phenotype shifts from elastin-dominated to collagen-dominated load bearing.Model-based assessment of arterial wall mechanics provides a promising tool to further improve interpretation of arterial stiffness measurements in patients. Further development of such methodology applied to various mouse models may improve the understanding and interpretation of arterial stiffening in ageing and disease.

Comparison of 10 murine models reveals a distinct biomechanical phenotype in thoracic aortic aneurysms

Thoracic aortic aneurysms are life-threatening lesions that afflict young and old individuals alike. They frequently associate with genetic mutations and are characterized by reduced elastic fibre integrity, dysfunctional smooth muscle cells, improperly remodelled collagen and pooled mucoid material. There is a pressing need to understand better the compromised structural integrity of the aorta that results from these genetic mutations and renders the wall vulnerable to dilatation, dissection or rupture. In this paper, we compare the biaxial mechanical properties of the ascending aorta from 10 murine models: wild-type controls, acute elastase-treated, and eight models with genetic mutations affecting extracellular matrix proteins, transmembrane receptors, cytoskeletal proteins, or intracellular signalling molecules. Collectively, our data for these diverse mouse models suggest that reduced mechanical functionality, as indicated by a decreased elastic energy storage capability or reduced distensibility, does not predispose to aneurysms. Rather, despite normal or lower than normal circumferential and axial wall stresses, it appears that intramural cells in the ascending aorta of mice prone to aneurysms are unable to maintain or restore the intrinsic circumferential material stiffness, which may render the wall biomechanically vulnerable to continued dilatation and possible rupture. This finding is consistent with an underlying dysfunctional mechanosensing or mechanoregulation of the extracellular matrix, which normally endows the wall with both appropriate compliance and sufficient strength.

Characterization of viscoelastic properties of normal and cancerous human breast cells using a confining microchannel

Biomechanical properties have been revealed as potential biomarkers for distinguishing cancer cells from normal cells. In this work, we report a novel technique using a confining microchannel for biomechanical phenotyping for floating human cells, including one normal breast cell line (MCF-10A) and two breast cancer cell lines (MCF-7 and MDA-MB-231). The floating cells move under a defined pressure profile along the microchannel, in which the cells deform dynamically under compression by the channel sidewalls. We adopt the Hertz and Tatara model to convert deformed cell shapes to cell diameters and transient stress–strain ratios. By further considering cell viscoelasticity as a standard linear solid model, we compute for whole-cell viscosity, and instantaneous and relaxed moduli. Our results show that the selected cell types have significant different viscoelastic properties. We further implement cell-type classification based on the multiple parametric biomechanical cell properties with reasonable sensitivities (>65%). Applications of the confining microchannel can be further extended for high-throughput, continuous-flow deep phenotyping of rare cells by surface functionalization for both biomechanical and biochemical biomarkers for more comprehensive and promising cell characterization.

Biomechanical Phenotyping of the Murine Aorta: What Is the Best Control?

The availability of diverse mouse models is revealing increasingly greater information on arterial mechanics, including homeostatic adaptations and pathologic maladaptations to genetic, pharmacological, and surgical manipulations. Fundamental to understanding such biomechanical changes, however, is reliable information on appropriate control vessels. In this paper, we contrast 15 different geometrical and mechanical metrics of biaxial wall mechanics for the ascending aorta across seven different types of possible control mice. We show that there is a comforting similarity across these multiple controls for most, though not all, metrics. In particular, three potential controls, namely, noninduced conditional mice, exhibit higher values of distensibility, an important clinical metric of structural stiffness, and two of these potential controls also have higher values of intrinsic circumferential material stiffness. There is motivation, therefore, to understand better the biomechanical changes that can arise with noninduced Cre-lox or similar approaches for generating mutations conditionally. In cases of germline mutations generated by breeding heterozygous +/- mice, however, the resulting homozygous +/+ mice tend to exhibit properties similar to traditional (C57BL/6) controls.

Lateral Wedge Insoles for Reducing Biomechanical Risk Factors for Medial Knee Osteoarthritis Progression: A Systematic Review and Meta-Analysis.

Lateral wedge insoles are intended to reduce biomechanical risk factors of medial knee osteoarthritis (OA) progression, such as increased knee joint load; however, there has been no definitive consensus on this topic. The aim of this systematic review and meta-analysis was to establish the within-subject effects of lateral wedge insoles on knee joint load in people with medial knee OA during walking. Six databases were searched from inception until February 13, 2015. Included studies reported on the immediate biomechanical effects of lateral wedge insoles during walking in people with medial knee OA. Primary outcomes of interest relating to the biomechanical risk of disease progression were the first and second peak external knee adduction moment (EKAM) and knee adduction angular impulse (KAAI). Eligible studies were pooled using random-effects meta-analysis. Eighteen studies were included with a total of 534 participants. Lateral wedge insoles resulted in a small but statistically significant reduction in the first peak EKAM (standardized mean difference [SMD] -0.19; 95% confidence interval [95% CI] -0.23, -0.15) and second peak EKAM (SMD -0.25; 95% CI -0.32, -0.19) with a low level of heterogeneity (I(2) = 5% and 30%, respectively). There was a favorable but small reduction in the KAAI with lateral wedge insoles (SMD -0.14; 95% CI -0.21, -0.07, I(2) = 31%). Risk of methodologic bias scores (quality index) ranged from 8 to 13 out of 16. Lateral wedge insoles cause small reductions in the EKAM and KAAI during walking in people with medial knee OA. Current evidence demonstrates that lateral wedge insoles appear ineffective at attenuating structural changes in people with medial knee OA as a whole and may be better suited to targeted use in biomechanical phenotypes associated with larger reductions in knee load.

Consistent Biomechanical Phenotyping of Common Carotid Arteries from Seven Genetic, Pharmacological, and Surgical Mouse Models

The continuing lack of longitudinal histopathological and biomechanical data for human arteries in health and disease highlights the importance of studying the many genetic, pharmacological, and surgical models that are available in mice. As a result, there has been a significant increase in the number of reports on the biomechanics of murine arteries over the past decade, particularly for the common carotid artery. Whereas most of these studies have focused on wild-type controls or comparing controls vs. a single model of altered hemodynamics or vascular disease, there is a pressing need to compare results across many different models to understand more broadly the effects of genetic mutations, pharmacological treatments, or surgical alterations on the evolving hemodynamics and the microstructure and biomechanical properties of these vessels. This paper represents a first step toward this goal, that is, a biomechanical phenotyping of common carotid arteries from control mice and seven different mouse models that represent alterations in elastic fiber integrity, collagen remodeling, and smooth muscle cell functionality.

Multiscale agent-based modelling of ovarian cancer progression under the stimulation of the STAT 3 pathway.

This research is developed to simulate ovarian cancer progression with signal transducers and activators of the transcription 3 (STAT 3) pathway. The main focus is on studying how the STAT 3 pathway affects the cancer cells' biomechanical phenotype under the stimulation of the interleukin-6 (IL-6) cytokine and various well-known microscopic factors. The simulated results agreed with recent experimental evidence that ovarian cancer cells with a stimulated STAT 3 pathway have high survival rates and drug resistance. And we discussed how the IL6 and these well-known microscopic factors impacted the cancer progression.

A microfluidic platform for profiling biomechanical properties of bacteria.

The ability to resist mechanical forces is necessary for the survival and division of bacteria and has traditionally been probed using specialized, low-throughput techniques such as atomic force microscopy and optical tweezers. Here we demonstrate a microfluidic technique to profile the stiffness of individual bacteria and populations of bacteria. The approach is similar to micropipette aspiration used to characterize the biomechanical performance of eukaryotic cells. However, the small size and greater stiffness of bacteria relative to eukaryotic cells prevents the use of micropipettes. Here we present devices with sub-micron features capable of applying loads to bacteria in a controlled fashion. Inside the device, individual bacteria are flowed and trapped in tapered channels. Less stiff bacteria undergo greater deformation and therefore travel further into the tapered channel. Hence, the distance traversed by bacteria into a tapered channel is inversely related to cell stiffness. We demonstrate the ability of the device to characterize hundreds of bacteria at a time, measuring stiffness at 12 different applied loads at a time. The device is shown to differentiate between two bacterial species, E. coli (less stiff) and B. subtilis (more stiff), and detect differences between E. coli submitted to antibiotic treatment from untreated cells of the same species/strain. The microfluidic device is advantageous in that it requires only minimal sample preparation, no permanent cell immobilization, no staining/labeling and maintains cell viability. Our device adds detection of biomechanical phenotypes of bacteria to the list of other bacterial phenotypes currently detectable using microchip-based methods and suggests the feasibility of separating/selecting bacteria based on differences in cell stiffness.

Biomechanical Phenotyping of Central Arteries in Health and Disease: Advantages of and Methods for Murine Models

The stiffness and structural integrity of the arterial wall depends primarily on the organization of the extracellular matrix and the cells that fashion and maintain this matrix. Fundamental to the latter is a delicate balance in the continuous production and removal of structural constituents and the mechanical state in which such turnover occurs. Perturbations in this balance due to genetic mutations, altered hemodynamics, or pathological processes result in diverse vascular phenotypes, many of which have yet to be well characterized biomechanically. In this paper, we emphasize the particular need to understand regional variations in the biaxial biomechanical properties of central arteries in health and disease and, in addition, the need for standardization in the associated biaxial testing and quantification. As an example of possible experimental methods, we summarize testing protocols that have evolved in our laboratory over the past 8years. Moreover, we note advantages of a four fiber family stress-stretch relation for quantifying passive biaxial behaviors, the use of stored energy as a convenient scalar metric of the associated material stiffness, and the utility of appropriate linearizations of the nonlinear, anisotropic relations both for purposes of comparison across laboratories and to inform computational fluid-solid-interaction models. We conclude that, notwithstanding prior advances, there remain many opportunities to advance our understanding of arterial mechanics and mechanobiology, particularly via the diverse genetic, pharmacological, and surgical models that are, or soon will be, available in the mouse.

Analysis of multiple physical parameters for mechanical phenotyping of living cells

Since the cytoskeleton is known to regulate many cell functions, an increasing amount of effort to characterize cells by their mechanical properties has occured. Despite the structural complexity and dynamics of the multicomponent cytoskeleton, mechanical measurements on single cells are often fit to simple models with two to three parameters, and those parameters are recorded and reported. However, different simple models are likely needed to capture the distinct mechanical cell states, and additional parameters may be needed to capture the ability of cells to actively deform. Our new approach is to capture a much larger set of possibly redundant parameters from cells' mechanical measurement using multiple rheological models as well as dynamic deformation and image data. Principal component analysis and network-based approaches are used to group parameters to reduce redundancies and develop robust biomechanical phenotyping. Network representation of parameters allows for visual exploration of cells' complex mechanical system, and highlights unexpected connections between parameters. To demonstrate that our biomechanical phenotyping approach can detect subtle mechanical differences, we used a Microfluidic Optical Cell Stretcher to mechanically stretch circulating human breast tumor cells bearing genetically-engineered alterations in c-src tyrosine kinase activation, which is known to influence reattachment and invasion during metastasis.

Endogenous contrast blood flow imaging in embryonic hearts using hemoglobin contrast subtraction angiography

The genetic basis of congenital heart disease is yet to be defined, and the interactions between the malformed heart and biomechanical cardiac performance remain poorly understood. Functional optical imaging enables detailed biomechanical phenotyping of cardiac dysfunction in small animal models, which in turn enables specific gene-phenotype relationship. We have developed a new microangiography technique based on flow imaging using endogenous hemoglobin contrast enabling in vivo assessment and biomechanical phenotyping of Xenopus tropicalis embryonic heart. We demonstrated that hemoglobin contrast angiography can be used to quantify physiological response to treatment with well-established cardioactive drugs.

Mechanical Properties of Plant Cell Walls Probed by Relaxation Spectra

Transformants and mutants with altered cell wall composition are expected to display a biomechanical phenotype due to the structural role of the cell wall. It is often quite difficult, however, to distinguish the mechanical behavior of a mutant's or transformant's cell walls from that of the wild type. This may be due to the plant's ability to compensate for the wall modification or because the biophysical method that is often employed, determination of simple elastic modulus and breakstrength, lacks the resolving power necessary for detecting subtle mechanical phenotypes. Here, we apply a method, determination of relaxation spectra, which probes, and can separate, the viscoelastic properties of different cell wall components (i.e. those properties that depend on the elastic behavior of load-bearing wall polymers combined with viscous interactions between them). A computer program, BayesRelax, that deduces relaxation spectra from appropriate rheological measurements is presented and made accessible through a Web interface. BayesRelax models the cell wall as a continuum of relaxing elements, and the ability of the method to resolve small differences in cell wall mechanical properties is demonstrated using tuber tissue from wild-type and transgenic potatoes (Solanum tuberosum) that differ in rhamnogalacturonan I side chain structure.

Genetic effects for femoral biomechanics, structure, and density in C57BL/6J and C3H/HeJ inbred mouse strains.

Genome-wide QTL analysis for bone density, structure, and biomechanical phenotypes was performed in 999 (B6xC3H)F2 mice. Multivariate phenotypes were also derived to test for pleiotropic QTL effects. Highly significant QTLs were detected with pleiotropic effects on many of these phenotypes, and QTLs with unique effects on specific phenotypes were found as well. The inbred C57BL/6J (B6) and C3H/HeJ (C3H) mouse strains were previously shown to segregate quantitative trait loci (QTLs) for femoral bone density. The 999 s filial (F2) mouse progeny were further phenotyped for measures of femoral biomechanics (load to failure, Fu; work to failure, U; stiffness, S), structure (polar moment of inertia, Ip; moment of inertia ratio, Ir), and more specific femoral midshaft bone density measures (cortical and total vBMD). Two novel multivariate phenotypes were computed using principal component analysis, thus aiding in the exploration of pleiotropic effects of the QTLs detected. Results of a genome-wide analysis provided strong evidence of pleiotropic QTL effects on chromosome 4, with six of the seven primary phenotypic measures, representing femoral biomechanics, density, and structure, producing LOD scores greater than 8. Chromosomes 1, 8, 13, and 14 were also identified as harboring QTLs that affect phenotypes in two of the three aspects of bone properties. QTLs uniquely contributing to variability in biomechanical measures were identified on chromosomes 10 and 12, whereas a QTL solely affecting structure was found on chromosome 17. Analysis of the evidence for pleiotropic effects using principal component analysis revealed pleiotropic QTLs on chromosomes 4 and 14, influencing nearly all the bone phenotypes measured and revealed QTLs on chromosomes 1, 8, 13, and 17 with pleiotropic effects restricted to either density or the structure and stiffness phenotypes. The use of multivariate phenotypes has allowed us to identify pleiotropic effects of several QTLs previously linked in studies of other mouse strains and in human studies of bone mineral density and femoral structure, which will provide important insight regarding the importance of allelic variation on the entire skeleton.