Seiji Omata

Effects of both vitamin C and mechanical stimulation on improving the mechanical characteristics of regenerated cartilage

The present work describes the influence of both vitamin C (VC) and mechanical stimulation on development of the extracellular matrix (ECM) and improvement in mechanical properties of a chondrocyte-agarose construct in a regenerating tissue disease model of hyaline cartilage. We used primary bovine chondrocytes and two types of VC, ascorbic acid (AsA) as an acidic form and ascorbic acid 2-phosphate (A2P) as a non-acidic form, and applied uniaxial compressive strain to the tissue model using a purpose-built bioreactor. When added to the medium in free-swelling culture conditions, A2P downregulated development of ECM and suppressed improvement of the tangent modulus more than AsA. By contrast, application of mechanical stimulation to the construct both increased the tangent modulus more than the free-swelling group containing A2P and enhanced the ECM network of inner tissue to levels nearly as high as the free-swelling group containing AsA. Thus, mechanical stimulation and strain appears to enhance the supply of nutrients and improve the synthesis of ECM via mechanotransduction pathways of chondrocytes. Therefore, we suggest that mechanical stimulation is necessary for homogenous development of ECM in a cell-associated construct with a view to implantation of a large-sized articular cartilage defect.

Training system using Bionic-eye for internal limiting membrane peeling

This paper reports on a training system for internal limiting membrane (ILM) peeling. ILM peeling task is difficult even for skilled surgeons, because ILM is a quite thin film on retina and there is a risk to damage the retina at peeling. In order to obtain the skill of ILM peeling and even evaluate newly developed medical devices, the surgical model that can be used for quantitative evaluation is highly demanded. Therefore, here we propose a surgical model for eye surgery named “Bionic-eye”. Fundus of Bionic-eye has artificial ILM and retina. In addition, we assembled Bionic-eye on a whole body surgical model of human named “Bionic-humanoid”.

Effect of Vitamin C on Mechanical Property of the Regenerated-Cartilage Tissue

Tissue engineering has become a promising option toward the treatment of osteoarthritis, allowing researchers to produce functional replacements for diseased cartilage. The mechanical behavior of articular cartilage depends on an extracellular matrix (ECM), i.e., type II collagen and proteoglycans. The collagen fibers play an important role to establish the mechanical strength in the cartilage tissue. Thus we focused the influence of L-ascorbic acid (AsA, as an acidic form) and ascorbic acid 2-phosphate (A2P, as a non-acidic form), as different concentrations in culture medium on mechanical property of the regenerated cartilage tissue because the collagen synthesis needs AsA. By increasing AsA concentration in culture medium, the result indicated that the tangent modulus of the cultured constructs was significantly increased and the synthesized collagen fibrils were spread out spatially in the immunofluorescence images. This study strongly suggests, therefore, that mechanical property of regenerated-cartilage tissue depended on interconnecting among cells by the ECMs, mainly by the collagen network. In contrast, significant damage by cellular toxicity was found in the high AsA concentration and the tangent modulus did not increase. Although damage of the construct was not shown by dose of A2P into culture medium as a consequence of its low reactivity, the tangent modulus did not statistically increase as collagen network had connected among chondrocytes.

Relationship between dynamic stress field and ECM production in regenerated cartilage tissue

Mechanical stresses exerted in articular cartilage during daily joint movements can stimulate the metabolism of chondrocytes in cartilage. Upregulative effects of mechanical stimuli on extracellular matrix (ECM) biosynthesis of chondrocytes have been utilized in the cartilage tissue for regenerative tissue engineering. However, dynamic functionalities of these regenerated cartilages were lacked compared with the natural arthrodial cartilage. Natural synovial joint surfaces are contacting and sliding each other. As a result, the loading condition applied to the cartilage tissue is not simple. Under the contact between surfaces, non-uniform stress distribution is exerted in the cartilage tissue. The shear stress at contact area is highly depended on the distance from the surface. Moreover, the surface region is exposed to larger shear stress compared with the deep region. We hypothesized that high stress in surface region may stimulate chondrocytes selectively and establish anisotropic structure in elaborated tissue. In this study, the relative motion between cartilage surfaces in a synovial joint is simulated by the rolling-sliding motion of the roller on the cultured chondrocyte-agarose construct. Then, we investigated effects on the ECM distribution and morphology of regenerated cartilage tissue.

Tissue Development and Mechanical Property in the Regenerated-Cartilage Tissue

Vertebrates have multiple synovial joints, and these joints in humans enable a number of activities of daily life. In particular, the hip, knee, shoulder, elbow, and ankle are all synovial joints with synovium inwhich the secretes synovial fluid. The articular surface is coveredwith smooth hyaline cartilage, which forms the part of the skeletal system that is notably different from mineralized bone in both function and histological composition [1]. The formation of animal cartilage from mesenchyme occurs in numerous areas of the embryo, such as the skull, limbs, and spine. This tissue contains chondrocytes which synthesize and maintain extracellular matrix (ECM), which is composed of a dense network of collagen molecules and proteoglycans [2]. There are three types of cartilage; fibrocartilage, elastic cartilage, and hyaline cartilage. Fibrocartilage contains regions of organized fibrous tissue, containing type I collagen in addition to the normal type II collagen. It is found in the annulus fibrosus of the intervertebral discs, meniscus and temporomandibular joints. Elastic cartilage contains additional elastin fibers and is a type of cartilage present in the ear, larynx, and epiglottis. Finally, hyaline cartilage coats the articular surfaces of bone epiphyses and is composed of individual chondrocytes bound together by the ECM. The major constituent of the hyaline cartilage is held in place by proteoglycan (about 10 % of wet weight) and type II collagen (10–20 % of wet weight), which forms a meshwork with high tensile strength [3–5].

A surgical simulator for peeling the inner limiting membrane during wet conditions

The present study was performed to establish a novel ocular surgery simulator for training in peeling of the inner limited membrane (ILM). This simulator included a next-generation artificial ILM with mechanical properties similar to the natural ILM that could be peeled underwater in the same manner as in actual surgery. An artificial eye consisting of a fundus and eyeball parts was fabricated. The artificial eye was installed in the eye surgery simulator. The fundus part was mounted in the eyeball, which consisted of an artificial sclera, retina, and ILM. To measure the thickness of the fabricated ILM on the artificial retina, we calculated the distance of the step height as the thickness of the artificial ILM. Two experienced ophthalmologists then assessed the fabricated ILM by sensory evaluation. The minimum thickness of the artificial ILM was 1.9 ± 0.3 μm (n = 3). We were able to perform the peeling task with the ILM in water. Based on the sensory evaluation, an ILM with a minimum thickness and 1000 degrees of polymerization was suitable for training. We installed the eye model on an ocular surgery simulator, which allowed for the performance of a sequence of operations similar to ILM peeling. In conclusion, we developed a novel ocular surgery simulator for ILM peeling. The artificial ILM was peeled underwater in the same manner as in an actual operation.

Fabrication of 3D Capillary Vessel Models with Circulatory Connection Ports

Bionic microscopic vessel models can contribute to the development of vascular treatment skills and techniques for clinical training. Most microscopic vessel models are limited to two dimensions, but three-dimensional (3D) models are important for surgery, such as on retina microscopic vessels, for the observation of colon microvessels, for measuring the deformability of red blood cell (RBC), and so on. Therefore, bionic 3D blood vessel models are increasingly in demand. For this reason, it is necessary to establish 3D fabrication techniques for microchannels. In this study, we established two fabrication methods for 3D microfluidic devices for the development of microscopic vessel models. First, we employed an exposure method using photolithographic technology. Second, we employed a 3D method using femtosecond laser and mask hybrid exposure (FMEx). Both methods made it possible to fabricate a millimeter-scale 3D structure with a submicrometer resolution and achieve an easy injection of solution. This is because it was possible to fabricate typical microfluidic channels used for model inlet and outlet ports. Furthermore, in the FMEx method, we employed an acid-diffusion effect using a chemically amplified resist to form a circular channel cross-section. The acid-diffusion effect made it realizable to fabricate a smooth surface independent of the laser scanning line width. Thus, we succeeded in establishing two methods for the fabrication of bionic 3D microfluidic devices with microfluidic channels having diameters of 15–16 µm for mimicking capillary vessels.

Automatic segmentation of eyeball structures from micro-CT images based on sparse annotation

A surgical simulator with elaborate artificial eyeball models has been developed for ophthalmic surgeries, in which sophisticated skills are required. To create the elaborate eyeball models with microstructures included in an eyeball, a database of eyeball models should be compiled by segmenting eye structures based on high-resolution medical images. Therefore, this paper presents an automated segmentation of eye structures from micro-CT images by using Fully Convolutional Networks (FCNs). In particular, we aim to construct a method for accurately segmenting eye structures from sparse annotation data. This method performs end-to-end segmentation of eye structures, including a workflow from training the FCN based on sparse annotation to obtaining the segmentation of the entire eyeball. We use the FCN trained on the slices sparsely annotated in a micro-CT volume to segment the remaining slices in the same volume. To achieve accurate segmentation from less annotated images, the multi-class segmentation is performed by using the network trained on the preprocessed and augmented micro-CT images; in the preprocessing, we apply filters for removing ring artifacts and random noises to the images, while in the data augmentation process, rotation and elastic deformation operations are performed on the sparsely-annotated training data. From the results of experiments for evaluating segmentation performances based on sparse annotation, we found that the FCN trained with data augmentation could achieve high segmentation accuracy of more than 90% even from a sparse training subset of only 2.5% of all slices.

Effect of Gamma Ray Irradiation on Friction Property of Poly(vinyl alcohol) Cast-Drying on Freeze-Thawed Hybrid Gel

Poly(vinyl alcohol) (PVA) is a biocompatible polymer with low toxicity. It is possible to prepare physically cross-linked PVA gels having hydrogen bonds without using a cross-linking agent. The newly reported physically cross-linked PVA cast-drying (CD) on freeze-thawed (FT) hybrid gel has an excellent friction property, which is expected to be applied as a candidate material for artificial cartilage. Gamma ray sterilization for clinical applications usually causes additional chemical cross-linking and changes physical properties of gels. In this study, CD on FT hybrid gels were irradiated using gamma rays at a different dose rate and irradiance. The results showed the optimized irradiation conditions for gamma irradiated gels to retain excellent friction characteristics.

Fabrication of retina model having photoelastic pressure sensor for vitreoretinal surgery simulator

We proposed sensor-integrated retina model with similar Youngs modulus to human retina for training of ophthalmologist. Mimic of mechanical properties of human retina is important issue for surgical simulator. Moreover, integration of sensor into surgical model is useful for evaluation of surgeon skill. First, we fabricate eye ball model having retina and scleral with similar Youngs modulus to human by adjusting the concentration of catalyst. Then, photoelastic constant of retinal model is increased by adding the carbon nanotube. As a demonstration, pressure distribution on retinal model was measured using the assembled retina and sclera model in skull model using photoelastic pressure measurement.