Biaxial mechanical testing of human sclera

The biomechanical environment of the optic nerve head (ONH), of interest in glaucoma, is strongly affected by the biomechanical properties of sclera. However, there is a paucity of information about the variation of scleral mechanical properties within eyes and between individuals. We thus used biaxial testing to measure scleral stiffness in human eyes. Ten eyes from 5 human donors (age 55.4±3.5 years; mean±SD) were obtained within 24 h of death. Square scleral samples (6 mm on a side) were cut from each ocular quadrant 3–9 mm from the ONH centre and were mechanically tested using a biaxial extensional tissue tester (BioTester 5000, CellScale Biomaterials Testing, Waterloo). Stress–strain data in the latitudinal (toward the poles) and longitudinal (circumferential) directions, here referred to as directions 1 and 2, were fit to the four-parameter Fung constitutive equation W=c(e^Q?1), where $Q=c_1{E}_{11}^2+c_2{E}_{22}^2+2c_3{E}_{11}{E}_{22}$ and W, c’s and E_ij are the strain energy function, material parameters and Green strains, respectively. Fitted material parameters were compared between samples. The parameter c₃ ranged from 10^?7 to 10^?8, but did not contribute significantly to the accuracy of the fitting and was thus fixed at 10^?7. The products c?c₁ and c?c₂, measures of stiffness in the 1 and 2 directions, were 2.9±2.0 and 2.8±1.9 MPa, respectively, and were not significantly different (two-sided t-test; p=0.795). The level of anisotropy (ratio of stiffness in orthogonal directions) was 1.065±0.33. No statistically significant correlations between sample thickness and stiffness were found (correlation coefficients=?0.026 and ?0.058 in directions 1 and 2, respectively). Human sclera showed heterogeneous, near-isotropic, nonlinear mechanical properties over the scale of our samples.     

Biaxial mechanical response of bioprosthetic heart valve biomaterials to high in-plane shear

Utilization of novel biologically-derived biomaterials in bioprosthetic heart valves (BHV) requires robust constitutive models to predict the mechanical behavior under generalized loading states. Thus, it is necessary to perform rigorous experimentation involving all functional deformations to obtain both the form and material constants of a strain-energy density function. In this study, we generated a comprehensive experimental biaxial mechanical dataset that included high in-plane shear stresses using glutaraldehyde treated bovine pericardium (GLBP) as the representative BHV biomaterial. Compared to our previous study (Sacks, JBME, v.121, pp. 551-555, 1999), GLBP demonstrated a substantially different response under high shear strains. This finding was underscored by the inability of the standard Fung model, applied successfully in our previous GLBP study, to fit the high-shear data. To develop an appropriate constitutive model, we utilized an interpolation technique for the pseudo-elastic response to guide modification of the final model form. An eight parameter modified Fung model utilizing additional quartic terms was developed, which fitted the complete dataset well. Model parameters were also constrained to satisfy physical plausibility of the strain energy function. The results of this study underscore the limited predictive ability of current soft tissue models, and the need to collect experimental data for soft tissue simulations over the complete functional range.     

Model for immunologic testing of biomaterials]

Corrosion products and electric fields are capable of changing proteins to antigens, thus permitting the immunological system to identify the biomaterial as foreign. The reaction between corrosion products and a macro-molecule also leads to an antigen (carrier antigen), such as conformational changes of a macro-molecule, e.g. a protein, caused by the electric field at the implant surface (modified macro-molecule antigen). While the sensitivity to corrosion and the effectiveness of galvanic elements is measurable by electrochemical methods, suitable methods of determining the field strength in the vicinity of biomaterial surfaces are still unavailable. The influence of the double layer of uncoated and coated titanium surfaces on the conformation of proteins and their conversion to antigens are investigated with polyclonal antibodies capable of identifying the unchanged protein despite adsorption to the surface. 14C-marked Bovine Serum Albumin serves as a model protein. Determination of the total number of protein molecules adsorbed is effected via the detection of the emitted electrons. The quotient of the concentration of natural proteins to the concentration of adsorbed molecules gives the biocompatibility index, which is independent of the surface area, and gives an indication of the expected biocompatibility of the material. The results of the biological tests of titanium and two coating materials on titanium were confirmed in an animal experiment. It is possible that in the future immunological tests may replace experiments in animals.     

Biaxial tensile testing and constitutive modeling of human supraspinatus tendon

The heterogeneous composition and mechanical properties of the supraspinatus tendon offer an opportunity for studying the structure-function relationships of fibrous musculoskeletal connective tissues. Previous uniaxial testing has demonstrated a correlation between the collagen fiber angle distribution and tendon mechanics in response to tensile loading both parallel and transverse to the tendon longitudinal axis. However, the planar mechanics of the supraspinatus tendon may be more appropriately characterized through biaxial tensile testing, which avoids the limitation of nonphysiologic traction-free boundary conditions present during uniaxial testing. Combined with a structural constitutive model, biaxial testing can help identify the specific structural mechanisms underlying the tendon's two-dimensional mechanical behavior. Therefore, the objective of this study was to evaluate the contribution of collagen fiber organization to the planar tensile mechanics of the human supraspinatus tendon by fitting biaxial tensile data with a structural constitutive model that incorporates a sample-specific angular distribution of nonlinear fibers. Regional samples were tested under several biaxial boundary conditions while simultaneously measuring the collagen fiber orientations via polarized light imaging. The histograms of fiber angles were fit with a von Mises probability distribution and input into a hyperelastic constitutive model incorporating the contributions of the uncrimped fibers. Samples with a wide fiber angle distribution produced greater transverse stresses than more highly aligned samples. The structural model fit the longitudinal stresses well (median R(2) ≥ 0.96) and was validated by successfully predicting the stress response to a mechanical protocol not used for parameter estimation. The transverse stresses were fit less well with greater errors observed for less aligned samples. Sensitivity analyses and relatively affine fiber kinematics suggest that these errors are not due to inaccuracies in measuring the collagen fiber organization. More likely, additional strain energy terms representing fiber-fiber interactions are necessary to provide a closer approximation of the transverse stresses. Nevertheless, this approach demonstrated that the longitudinal tensile mechanics of the supraspinatus tendon are primarily dependent on the moduli, crimp, and angular distribution of its collagen fibers. These results add to the existing knowledge of structure-function relationships in fibrous musculoskeletal tissue, which is valuable for understanding the etiology of degenerative disease, developing effective tissue engineering design strategies, and predicting outcomes of tissue repair.     

Progeny testing for recessive genes: procedures and interpretations

Summary Methods for calculating the probability of detecting a carrier of a recessive gene by utilizing matings among related individuals are presented for single and litter bearing species. The confidence level for detection of heterozygosity depends upon: (1) the genetic relationship between mates, (2) the number of mates per male and the number of offspring per mate, (3) whether an estimate of recessive gene frequency before selection is available and (4) the magnitude of that frequency. Methods of computing probability of heterozygosity vs homozygosity utilizing Bayes theorem also are presented. In the conventional progeny test method, a sire initially is assumed heterozygous before calculations are made, but no prior information concerning his probable genotype is utilized. In the method using Bayes theorem, prior sources of information from relatives or from estimates of population allele frequency are utilized. This method gives the exact probability that a sire is not a carrier, given prior information and that he produces all normal offspring. These methods could be used in any sexually reproducing species to identify not only detrimental genes but beneficial genes as well.     

Standardization of automated cell-based protocols for toxicity testing of biomaterials

Advances in high-throughput screening (HTS) instrumentation have led to enormous reduction of costs (e.g., of pipetting stations) and to the development of smaller instruments for automation of day-to-day routines in small research laboratories. In the biomaterials community, there has been an increasing interest for standardized screening protocols to identify cell type-specific cytocompatible biomaterials suitable for tissue engineering (TE) applications. In this study, the authors established a multiplexed assay protocol for toxicity screening of biomaterials using a low- to medium-throughput robotic liquid handling station (LHS). The protocol contains analysis of viability, cytotoxicity, and apoptosis combined in one assay. This study includes performance results of a side-by-side comparison of the EpMotion 5070 LHS and conventional pipetting/dispensing systems. Critical parameters were optimized each for a given platform. Higher accuracy and reproducibility were achieved for LHS compared to manually treated samples. The practicability and accuracy of the method in a typical small laboratory setting were tested by running daily routine tasks by trained and untrained laboratory staff. In addition, advantages and disadvantages as well as the step-by-step application protocol are reported. The approach described provides a potential utility in screening biomaterials toxicity, allowing researchers to meet the needs of low- and medium-throughput laboratories.     

Novel polymer biomaterials and interfaces inspired from cell membrane functions

Background

Materials with excellent biocompatibility on interfaces between artificial system and biological system are needed to develop any equipments and devices in bioscience, bioengineering and medicinal science. Suppression of unfavorable biological response on the interface is most important for understanding real functions of biomolecules on the surface. So, we should design and prepare such biomaterials.

Scoop of review

One of the best ways to design the biomaterials is generated from mimicking a cell membrane structure. It is composed of a phospholipid bilayered membrane and embedded proteins and polysaccharides. The surface of the cell membrane-like structure is constructed artificially by molecular integration of phospholipid polymer as platform and conjugated biomolecules. Here, it is introduced as the effectiveness of biointerface with highly biological functions observed on artificial cell membrane structure.

Major conclusions

Reduction of nonspecific protein adsorption is essential for suppression of unfavorable bioresponse and achievement of versatile biomedical applications. Simultaneously, bioconjugation of biomolecules on the phospholipid polymer platform is crucial for a high-performance interface.

General significance

The biointerfaces with both biocompatibility and biofunctionality based on biomolecules must be installed on advanced devices, which are applied in the fields of nanobioscience and nanomedicine.This article is part of a Special Issue entitled Nanotechnologies - Emerging Applications in Biomedicine.     

Preoperative testing before low-risk surgical procedures

Background:There is concern about increasing utilization of low-value health care services, including preoperative testing for low-risk surgical procedures. We investigated temporal trends, explanatory factors, and institutional and regional variation in the utilization of testing before low-risk procedures.Methods:For this retrospective cohort study, we accessed linked population-based administrative databases from Ontario, Canada. A cohort of 1 546 223 patients 18 years or older underwent a total of 2 224 070 low-risk procedures, including endoscopy and ophthalmologic surgery, from Apr. 1, 2008, to Mar. 31, 2013, at 137 institutions in 14 health regions. We used hierarchical logistic regression models to assess patient- and institution-level factors associated with electrocardiography (ECG), transthoracic echocardiography, cardiac stress test or chest radiography within 60 days before the procedure.Results:Endoscopy, ophthalmologic surgery and other low-risk procedures accounted for 40.1%, 34.2% and 25.7% of procedures, respectively. ECG and chest radiography were conducted before 31.0% (95% confidence interval [CI] 30.9%–31.1%) and 10.8% (95% CI 10.8%–10.8%) of procedures, respectively, whereas the rates of preoperative echocardiography and stress testing were 2.9% (95% CI 2.9%–2.9%) and 2.1% (95% CI 2.1%–2.1%), respectively. Significant variation was present across institutions, with the frequency of preoperative ECG ranging from 3.4% to 88.8%. Receipt of preoperative ECG and radiography were associated with older age (among patients 66–75 years of age, for ECG, adjusted odds ratio [OR] 18.3, 95% CI 17.6–19.0; for radiography, adjusted OR 2.9, 95% CI 2.8–3.0), preoperative anesthesia consultation (for ECG, adjusted OR 8.7, 95% CI 8.5–8.8; for radiography, adjusted OR 2.2, 95% CI 2.1–2.2) and preoperative medical consultation (for ECG, adjusted OR 6.8, 95% CI 6.7–6.9; for radiography, adjusted OR 3.6, 95% CI 3.5–3.6). The median ORs for receipt of preoperative ECG and radiography were 2.3 and 1.6, respectively.Interpretation:Despite guideline recommendations to limit testing before low-risk surgical procedures, preoperative ECG and chest radiography were performed frequently. Significant variation across institutions remained after adjustment for patient- and institution-level factors.In response to concerns about increasing utilization of low-value health care services, the American Board of Internal Medicine Foundation launched the Choosing Wisely campaign in the United States in 2012.¹ The goal of the campaign is to encourage conversations between physicians and patients about low-value care by defining “top 5” lists of tests, treatments and procedures that may be unnecessary or unsupported by evidence.¹ Subsequent Choosing Wisely campaigns have followed in other countries, including Canada starting in April 2014.^2,3 Of interest for health policy-makers, payers and clinicians are current utilization rates for the procedures mentioned in these recommendations. Establishing baseline rates permits an understanding of the extent of the problem of low-value care, which in turn allows monitoring of the effect of initiatives such as Choosing Wisely on utilization rates over time.One Choosing Wisely item included by many specialty societies is the recommendation to avoid routinely performing preoperative testing (including chest radiography, echocardiography and cardiac stress tests) for patients undergoing low-risk surgery.^4–6 This recommendation was previously included in the 2007 American College of Cardiology/American Heart Association guidelines on perioperative cardiovascular evaluation for noncardiac surgery⁷ and was reconfirmed in a recent update.⁸ Avoiding preoperative investigations in this setting is supported by evidence that routine testing in patients undergoing low-risk surgery does not improve outcomes or change management and may lead to further unnecessary downstream testing, cancellation of surgery, and increases in patient anxiety and cost.^7,9–12 To date, neither the rate of preoperative testing across a large and diverse jurisdiction nor the degree of variation at regional and institutional levels, where data may be “actionable,” is well understood.The objectives of this study were to determine utilization rates of preoperative tests before hospital-based low-risk surgical procedures at the jurisdictional, regional and institutional level. In addition, we aimed to evaluate temporal trends of preoperative testing over a 5-year period. We hypothesized that there would be significant regional and institutional variation in preoperative cardiac testing before low-risk surgery and that patients with prior cardiac comorbidities would have a higher rate of preoperative testing than those without such comorbidities.     

A family of Bayes multiple testing procedures

Under the model of independent test statistics, we propose atwo-parameter family of Bayes multiple testing procedures. Thetwo parameters can be viewed as tuning parameters. Using theBenjamini–Hochberg step-up procedure for controlling falsediscovery rate as a baseline for conservativeness, we choosethe tuning parameters to compromise between the operating characteristicsof that procedure and a less conservative procedure that focuseson alternatives that a priori might be considered likely ormeaningful. The Bayes procedures do not have the theoreticaland practical shortcomings of the popular stepwise procedures.In terms of the number of mistakes, simulations for two examplesindicate that over a large segment of the parameter space, theBayes procedure is preferable to the step-up procedure. Anotherdesirable feature of the procedures is that they are computationallyfeasible for any number of hypotheses.     

Special stereotactic radiotherapy techniques: procedures and equipment for treatment simulation and dose delivery

Stereotactic radiotherapy (SRT ) is a multi-step procedure with each step requiring extreme accuracy. Physician-dependent accuracy includes appropriate disease staging, multi-disciplinary discussion with shared decision-making, choice of morphological and functional imaging methods to identify and delineate the tumor target and organs at risk, an image-guided patient set-up, active or passive management of intra-fraction movement, clinical and instrumental follow-up. Medical physicist-dependent accuracy includes use of advanced software for treatment planning and more advanced Quality Assurance procedures than required for conventional radiotherapy. Consequently, all the professionals require appropriate training in skills for high-quality SRT.Thanks to the technological advances, SRT has moved from a “frame-based” technique, i.e. the use of stereotactic coordinates which are identified by means of rigid localization frames, to the modern “frame-less” SRT which localizes the target volume directly, or by means of anatomical surrogates or fiducial markers that have previously been placed within or near the target. This review describes all the SRT steps in depth, from target simulation and delineation procedures to treatment delivery and image-guided radiation therapy. Target movement assessment and management are also described.     

Cardiopulmonary exercise testing in congenital heart disease: equipment and test protocols

Cardiopulmonary exercise testing (CPET) in paediatric cardiology differs in many aspects from the tests as performed in adult cardiology. Children's cardiovascular responses during exercise testing present different characteristics, particularly oxygen uptake, heart rate and blood pressure response, which are essential in interpreting haemodynamic data. Diseases that are associated with myocardial ischaemia are very rare in children. The main indications for CPET in children are evaluation of exercise capacity and the identification of exercise-induced arrhythmias. In this article we will review exercise equipment and test protocols for CPET in children with congenital heart disease. (Neth Heart J 2009;17:339–44.)     

A novel surrogate lung material for impact studies: Development and testing procedures

This work focuses on the development of a surrogate lung material (SLM) that reproduces the dynamic response of a human lung under various loading conditions and also allows for the analysis of the extent and distribution of damage. The SLM consists of polyurethane foam used to mimic the spongy lung tissue and fluid-filled gelatine microcapsules used to simulate the damage of alveoli.The bursting pressure of the microcapsules was investigated by conducting low and high rate compression tests on single microcapsules. A bursting pressure of around 5 bar was measured which is comparable to the reported lung overpressure at injury level.Low and high rate compression tests were conducted on the SLMs. From the measured mechanical properties and mass density, the stress wave speed was calculated and found to be well in the range of the reported values for human lungs (16–70 m/s).In order to study the extent and distribution of damage in the SLMs, as represented by burst microcapsules, a CT scan analysis was carried out before and after the impacts. The CT scan results clearly demonstrated the magnitude and distribution of damage within the specimen. The results are then compared to the Bowen curves, the most often used criteria for predicting blast injuries in humans. An excellent agreement was found between the observed damage in the surrogate lungs and the expected damage in real human lungs.In general, the SLM showed similar stress wave speed, bursting pressure and damage to that of the real lungs.     

Biaxial mechanical testing of posterior sclera using high-resolution ultrasound speckle tracking for strain measurements

This study aimed to characterize the mechanical responses of the sclera, the white outer coat of the eye, under equal-biaxial loading with unrestricted shear. An ultrasound speckle tracking technique was used to measure tissue deformation through sample thickness, expanding the capabilities of surface strain techniques. Eight porcine scleral samples were tested within 72 h postmortem  . High resolution ultrasound scans of scleral cross-sections along the two loading axes were acquired at 25 consecutive biaxial load levels. An additional repeat of the biaxial loading cycle was performed to measure a third normal strain emulating a strain gage rosette for calculating the in-plane shear. The repeatability of the strain measurements during identical biaxial ramps was evaluated. A correlation-based ultrasound speckle tracking algorithm was used to compute the displacement field and determine the distributive strains in the sample cross-sections. A Fung type constitutive model including a shear term was used to determine the material constants of each individual specimen by fitting the model parameters to the experimental stress–strain data. A non-linear stress–strain response was observed in all samples. The meridian direction had significantly larger strains than that of the circumferential direction during equal-biaxial loadings (P's<0.05$P\text{'}s<0.05$). The stiffness along the two directions was also significantly different (P=0.02) but highly correlated (R²=0.8). These results showed that the mechanical properties of the porcine sclera were nonlinear and anisotropic under biaxial loading. This work has also demonstrated the feasibility of using ultrasound speckle tracking for strain measurements during mechanical testing.     

Toxicity testing of polymer materials for dialysis equipment: is there any need forin vivo testing?

In an earlier work, slightly more than 650 plastic materials, intended for use in medical devices, were tested with a battery of chemical, as well asin vitro andin vivo biological tests. An analysis showed that only a limited number of the tests used were actually necessary to obtain the same pass or fail decision as that obtained using the full test battery. This prompted us to prescreen all new materials with a small test battery consisting of the two most discriminating chemical tests and anin vitro cell growth inhibition test. The present work is a report of our findings after testing another 155 materials using this prescreen system.For each single one of the 155 tested materials the same decision on whether or not to use the material in the intended medical device would have been reached without anyin vivo testing. In no single case in a total of 851in vivo tests did an eluate that had passed thein vitro cell test give rise to a reactionin vivo. Thus, among the tests on living systems the cell test alone seems to be sensitive enough to provide sufficient information. Nothing appears to be gained from thein vivo animal tests. However, some of the materials that passed the prescreening tests later failed in one or several of the chemical tests. Both nonspecific chemical tests and tests for specific molecules seem to detect undesirable levels of leachable substances not detected by the prescreening system. Therefore these tests should not be abandoned. Abandoning unnecessaryin vivo testing, on the other hand, would save considerable costs.     

In vitro hemocompatibility testing of biomaterials according to the ISO 10993-4

The development of synthetic materials, textured polymers and metals and their increasing use in medicine make research of biomaterials' hemocompatibility very relevant. Problems arise from the polymorphism and diversity of the different materials, the static and dynamic test models and the patients' individual biologic factors. First, methods, models, tests as well as preanalytical factors have to be standardized according to the current knowledge in medicine laid down in the ISO 10993 part 4. The routine controls used in clinical chemistry and hematology have to be performed. Information about normal ranges (mean value, standard deviation, 95% confidence interval) should be provided. Tests have to be performed within a minimal delay of usually 2 h since some properties of blood change rapidly following collection. Various conditions (depending on the wall shear rate) were simulated within the centrifugation system and a Chandler system. Qualities and aspects of hemocompatibility such as platelet activation, oxidative burst, hemolysis, fibrinolysis, fibrin formation, generation of thrombin, contact activation, and complement activation were analysed and the results were entered non-dimensionally into a non-dimensional score system, where 0 points stand for the best and 65 points for the worst evaluation. We found a good correlation between the total score and contact activation, thrombin generation and leukocyte activation in a low shear stress system and a good correlation between the total score and thrombin generation, hemolysis and platelet activation in the high shear stress system. Further on the effect of additives and sterilization procedures can be measured. The concepts presented underline the relevance/importance of an efficient diagnostic approach to hemocompatibility that takes account of clinical and socio-economic concerns.     

Counting by weighing and its effect on seed testing procedures

Counting by weighing is an approximate procedure for obtaining a given number of items based on an estimate of how much the items will weigh. The procedure can be considerably faster than exact counting. We show how the effects of counting by weighing can be assessed, using a particular problem in seed testing for illustration. We find that using counting by weighing in place of exact counting has little effect on the outcome of the seed test provided that the coefficient of variation of the individual seed weights is not too large, and provided a sufficiently large initial sample is used to estimate mean seed weight.     

Murine leukemic lymphoblasts: homogenization and fractionation procedures for plasma membrane vesicles isolation

Plasma membrane vesicles were isolated from murine leukemic lymphoblasts L5178Y. The isolation procedure selected involved a method of mechanical disruption in a hypoosmotic-buffered solution and the separation of plasma membrane vesicles by an adaptation of the fractionation method described by D. W. McKeel and L. Jarett for fat cells (J. Cell Biol., 44, 417, 1970). In order to select the homogenization method we took into account several parameters: the extent of cell and nuclear disruption, the integrity of the nuclear membrane, the 5′-nucleotidase activity recovered at the first step of fractionation and the mitochondrial rupture. The homogenization method finally used yielded 89% of cellular rupture with only 9% of nuclear damage. The isolation procedure showed an overall yield of 70–90%. A plasma membrane fraction was isolated with an enrichment in 5′-nucleotidase and ouabain-sensitive (Na⁺K⁺)-ATPase specific activities of 15- and 13-fold, respectively, and essentially free of mitochondrial, lysosomal, and endoplasmic reticulum contamination. The electron microscopy demonstrated that the plasma membrane fraction essentially consisted of smooth vesicles of several sizes.