Computational model to evaluate modulation of the acoustic field in an ultrasonic bioreactor by incorporation of a water layer bounded by an acoustic absorbent boundary layer.

Ultrasonic bioreactors have been used for in vitro experimentation to study cellular responses to low-intensity pulsed ultrasound. The presence of an air interface in these bioreactors contributes to variability in the acoustic pressure field, reducing experimental reproducibility. A multiphysics finite element model was developed to simulate the acoustic field in an in-dish ultrasonic bioreactor, where the transducer is immersed in culture medium above the dish surface, and the effects of replacing air below the dish in the bioreactor with a water layer bounded by an acoustic absorbent layer were evaluated. Frequency domain simulations showed that the spatially-averaged pressure at the dish surface alternated between a minimum and maximum level as the distance between the dish and transducer increased. The ratio of the maximum to minimum level was 6.5-fold when the air interface was present, and this ratio dropped to 1.8-fold with replacement of the air interface. However, radial pressure variability was present with or without the air interface in the bioreactor model. Time-dependent simulations showed that the increase in acoustic pressure to a maximum level after US signal activation and the pressure drop after signal cessation were faster when the water-coupled non-reflective layer was used to replace the air layer below the dish, generating a pressure pattern that more closely followed the applied pulsed ultrasound signal due to reduced wave reflection and interference. Overall, this work showed that having water rather than air in contact with the lower dish surface when paired with an acoustic absorbent layer resulted in a less variable pressure field, providing an improved bioreactor design for in vitro experiments.

Electrospun core-sheath poly(vinyl alcohol)/silk fibroin nanofibers with Rosuvastatin release functionality for enhancing osteogenesis of human adipose-derived stem cells.

The aim of this study is to evaluate a core-shell nanofiber as a useful matrix for tuning Rosuvastatin (RSV) release and osteogenic differentiation in vitro. Polyvinyl alcohol (PVA) and silk fibroin were used as the shell and the core, respectively. To obtain a linear and beadless core-shell structure and an optimal release profile, the shell/core flow rate ratio was varied (0, 0.4, 0.6, 0.8, and 1). Formation of continuous nanofibers with an obvious core/sheath structure was proved using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Differential scanning calorimetry indicated the presence of two distinct phase structures in the nanofibers. Also, RSV molecules were dispersed in an amorphous state in the nanofibers. The in vitro release profile of the core-shell structure exhibited a biphasic release profile and the amount of released RSV was controlled by adjusting the shell flow rate. Human adipose-derived stem cells cultured on the RSV loaded nanofibers were found to improve cell proliferation and assist osteogenic differentiation as revealed by Alizarin red staining and real-time RT-PCR.

Interrelation of Hydration, Collagen Cross-Linking Treatment, and Biomechanical Properties of the Cornea.

PURPOSE: The present study was designed to investigate the effects of hydration and collagen cross-linking treatment on biomechanical properties of the cornea. METHODS: The original corneal collagen cross-linking protocol was used to induce cross-links in bovine corneas. The thickness of samples was used as a measure of their hydration and five different thickness groups (n = 5 each) were considered. The cross-linked corneal strips were allowed to hydrate/dehydrate until their thickness reached 500, 700, 900, 1100, and 1500 µm. The tensile behavior of specimens in each thickness group was characterized by conducting uniaxial tensile experiments. The experiments were done in mineral oil in order to keep the thickness of samples constant and minimize hydration changes. RESULTS: It was observed that collagen cross-linking treatment significantly increased both the maximum tensile stress and the equilibrium (relaxed) stress of the bovine cornea (p < 0.01). Furthermore, with increasing the thickness (hydration) of the collagen cross-linked samples, their tensile stiffness significantly decreased (p < 0.01). An exponential relation and a logarithmic expression successfully represented experimentally measured stress-strain behavior and relaxation response of all groups (r(2 )> 0.99), respectively. CONCLUSION: Hydration and collagen cross-linking treatment concomitantly affect biomechanical properties of the cornea. Therefore, an accurate estimate of stiffening effects of collagen cross-linking treatment option using uniaxial tensile experiments is only possible if the hydration of specimens is fully controlled.

Collagen cross-linking treatment effects on corneal dynamic biomechanical properties.

Cornea is a soft tissue with the principal function of transmitting and refracting light rays. The objective of the current study was to characterize possible effects of the riboflavin/UVA collagen cross-linking on corneal dynamic properties. The original corneal cross-linking protocol was used to induce cross-links in the anterior portion of the bovine cornea. A DMA machine was used to conduct mechanical tensile experiments at different levels of tensile strains. The samples were divided into a control group (n = 5) and a treated group (n = 5). All specimens were first stretched to a strain of 5% and allowed to relax for twenty minutes. After completion of the stress-relaxation experiment, a frequency sweep test with oscillations ranging from 0.01 to 10 Hz was performed. The same procedure was repeated to obtain the stress-relaxation and dynamic properties at 10% strain. It was observed that the collagen cross-linking therapy significantly increased the immediate and equilibrium tensile behavior of the bovine cornea (P < 0.05). Furthermore, for all samples in control and treated groups and throughout the whole range of frequencies, a significantly larger tensile storage modulus was measured at an axial strain of 10% compared to what was obtained at a tensile strain of 5%. Finally, it was noted that although this treatment procedure resulted in a significant increase in the storage and loss modulus at any axial strain and frequency (P < 0.05), it significantly reduced the ratio of the dissipated and stored energy during a single cycle of deformation. Therefore, it was concluded that while the riboflavin/UVA collagen cross-linking increased significantly corneal stiffness, it decreased significantly its damping capability and deformability. This reduced damping ability might adversely interfere with corneal mechanical performance.

Evaluation of hydration effects on tensile properties of bovine corneas.

PURPOSE: To determine effects of hydration on tensile stress-relaxation behavior of the bovine cornea. SETTING: Computational Biomechanics Laboratory, School of Mechanical and Aerospace Engineering, Oklahoma State University, Stillwater, Oklahoma, USA. DESIGN: Experimental study. METHODS: Bovine corneal strips were obtained from a local slaughterhouse and divided into 6 groups based on their thickness. The samples were first air dried and then rehydrated to reach the predetermined thickness of each group as follows: 0.4 mm (Group 1), 0.5 mm (Group 2), 0.7 mm (Group 3), 0.9 mm (Group 4), 1.1 mm (Group 5), and 1.5 mm (Group 6). A custom-built tensile machine was used to characterize the stress-relaxation properties. Mineral oil was used as the bathing fluid to prevent hydration variation during the experiments. RESULTS: Hydration significantly affected the immediate and transient tensile behavior of the cornea. With increasing hydration, there was a significant decrease in peak and equilibrium stress (P < .01). At all hydration levels, the tensile stress-strain curves and relaxation behavior were numerically represented with exponential and logarithmic mathematical expressions (R(2) > 0.99), respectively. An exponential relationship was found between the tangent modulus and hydration. CONCLUSIONS: Hydration of the corneal strips significantly affected their tensile and viscoelastic mechanical behavior. Therefore, while careful attention must be taken in interpreting the results in previous studies in which hydration of specimens was not fully controlled, it is important to monitor and report hydration of corneal samples in future studies. FINANCIAL DISCLOSURE: Neither author has a financial or proprietary interest in any material or method mentioned.

The relation between hydration and mechanical behavior of bovine cornea in tension.

The cornea is a transparent soft tissue covering the front of the eye. The biomechanical properties of the cornea have been commonly investigated by uniaxial tensile and inflation testing methods. The cornea like many other hydrated tissue swells when immersed in an ionic solution. Previous studies on hydrated tissues have shown that mechanical properties and hydration are closely related. The present study was designed to investigate the effects of thickness (hydration) variation due to swelling/dehydration on non-linear stress-strain response of the bovine cornea. Corneal strips were first air-dried and then soaked in a bathing solution until they reached an average thickness ranging from 0.3mm to 1.1mm. Based on their thickness, the samples were divided into different groups and uniaxial tests were performed to measure tensile properties. All experiments were done in mineral oil to prevent any hydration gain or loss during the tests. It was observed that swollen corneas had softer tensile properties in comparison with dehydrated ones. In particular, there was a significant difference between elastic tangent modulus of different groups (P<0.05). It was also shown that tensile behavior of bovine strips at any thickness within the range of 0.4-1.1mm can be obtained from a single experiment conducted on samples with known thickness (hydration). The findings of the present study confirm that mechanical properties obtained from uniaxial tensile experiments are strongly dependent on thickness (water amount) of samples; therefore, careful attention must be taken in interpreting previous studies which did not fully control the thickness of specimens.

Effects of bathing solution on tensile properties of the cornea.

The cornea is a transparent tissue with the major functions of protecting the inner contents of the eye and refracting incoming light. The biomechanical properties of the cornea strongly depend on the microstructure and composition of the stromal layer, a hydrated bio-gel. The uniaxial strip testing is a convenient and well-accepted experimental technique for characterizing corneal material parameters. It is known that the water content of specimens in this method depends on the osmolality of the bathing solution. The present study was designed to investigate the effects of different bathing solutions on uniaxial tensile material properties of the cornea. The tensile behavior of bovine corneal samples was measured in six different bathing solutions, i.e., hypertonic solution (12% NaCl solution), common preserving isotonic solutions (e.g., phosphate buffer saline, ophthalmic balanced salt solution, and 0.9% NaCl solution), hypotonic solution (distilled water), and neutral solution (mineral oil). It was observed that the bathing solution had significant influence on the tensile behavior of the corneal samples. In particular, the specimens tested in bathing solutions causing less swelling had significantly stiffer tensile properties. Furthermore, a simple mathematical model based on Voigt composite material model was developed to represent the measured solution-dependent tensile properties. The present study suggests that extra attention should be paid to corneal thickness (hydration) in uniaxial tensile experiments. It also provides important data on tensile properties of the cornea; such information could significantly contribute to improving the accuracy of numerical predictions of corneal biomechanics.

Swelling pressure and hydration behavior of porcine corneal stroma.

PURPOSE: The aim of this study was to characterize swelling pressure-thickness, swelling pressure-hydration and hydration-thickness relations of porcine cornea. METHODS: Mechanical compression tests and free swelling experiments were performed on porcine cornea. A rheometer (DHR-2, TA Instruments) with a thermally controlled fluid chamber filled with 0.9% NaCl solution was used to measure the equilibrium swelling pressure of (n = 17) corneal stromal specimens. The samples were compressed incrementally and their swelling pressure-thickness relations were obtained. In parallel to this investigation, a transient digital imaging microscope (H800-CL, American Scope Inc.), a USB autofocus camera (UM05, ViTiny), and a precision weighing scale (AGZN100, Torbal) were simultaneously used to measure the weight-thickness relation of (n = 8) corneal specimens. This experimental study gave the thickness-hydration relationship required for expressing swelling pressure measurements as a function of hydration. RESULTS: At the in vivo 666 ± 68 µm central corneal thickness, an average swelling pressure of 52 ± 13 mmHg and hydration of 3.36 ± 0.25 mg H2O/mg dry tissue were found. The swelling pressure was reported as functions of both tissue thickness and hydration. The average fixed charge density of ρF/F ~ 42.8 mM and dry density of 1.47±0.15 g/cm3 were found. The thickness-hydration relationship was only linear when the tissue thickness was within the range of physiological thickness. CONCLUSION: Overall, the physiological hydration and swelling pressure of the porcine cornea were within the same range of those reported previously for other mammalian corneas such as steers, rabbits and humans. Nevertheless, the thickness-hydration behavior of the porcine cornea was only similar to that of the human cornea.