Platelet–rich fibrin (PRF) was developed as an advanced form of platelet–rich plasma (PRP) and it is widely used as a source of growth factors for tissue regeneration. From this bio material, our laboratory made two important modifications. One was a thin PRF membrane prepared by the use of a novel PRF compression device. This method was capable of minimizing the loss of platelets and bio active growth factors and thereby enabled the PRF membrane to more effectively stimulate cell proliferation and neovascularization. To further clinically enhance the modified PRF membrane, it was exposed to mild heat compression technique. The heat compressed membrane was found to exhibit a longer period, three to four weeks, of chemical/physical stability. However, non–treated PRF membrane was resorbed within two weeks or less at implantation sites and therefore could barely maintain sufficient stability over an adequate amount of time for periodontal regeneration (bone and connective tissue). Compared with the control non–heat exposed gauze–compressed PRF, the resulting heat modified PRF membrane exhibited greater plasmin-resistant and remained stable for a significantly longer time period both in vitro and in vivo. Although additional modification may be required to further improve its clinical applicability, we suggest that this new modified membrane would be a promising bio material for guided tissue regeneration (GTR) treatment with the added advantage over currently available GTR membranes of contributing growth factors to improve regeneration potential.

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Culture of autologous cell sheets is one of the promising methods in regenerative medicine. Cultured epidermis is used for the coverage of massive burn wounds and cultured periosteum is for the treatment of infrabony periodontal defects. In cell sheet engineering cardiomyocyte sheets prepared with temperature–responsive culture dishes are layered to construct cardiac grafts. Those cultured cell sheets, however, need high techniques in culture and/or surgery. We have been developing several microporous membranes of biodegradable polyesters to handle cultured cell sheets easily. Microporous surfaces of poly(_L–lactic acid) (PLLA) membrane, which was prepared by thermally induced phase separation method, anchored periosteal tissue segments on the membranes. The cultured periosteal cell sheets on the membranes were successfully osteoinduced in an induction medium. The PLLA membrane implanted with the cell sheet was gradually degraded in a nude mouse without severe inflammation or fibrous encapsulation. Ductility of the membrane with a microporous surface was conferred by blending brittle PLLA with ductile poly(ε–caprolactone) (PCL). Established osteoblast–like (Saos–2) cells grew on a flexible microporous membrane of PCL and hydroxyapatite, the latter of which is a major mineral component of bone and used in bone defect treatment, although the cells grew in a single layer. A finger–like structure of PLLA microporous membrane prepared by nonsolvent–induced phase separation method with a surfactant enhances the cell growth in the membrane possibly by higher diffusion of oxygen, nutrients, and wastes. The biodegradable microporous sheets including those described in the review will complement the existing cultured cell sheet technology.

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As a powerful cell manipulation alternative in the recent rapid progress of cell biology, we are introducing a novel method which enables selective killing or detachment of the adherently cultured cells by micropattern projection onto a photo–responsive culture substrate functionalized with photo–acid–generating (PAG) polymers. We synthesized PAG polymers, which generate strong acid in response to visible light irradiation, and fabricated photo–responsive cell culture substrates by forming a thin layer of the PAG polymer on a polystyrene culture surface. Several anchorage dependent cell lines including CHO–K1, NIH/3T3, MDCK, HeLa and HepG2 were cultured on this surface until confluent, and test patterns were projected onto the substrates with the blue light using a PC–controlled micro–pattern irradiation system. A few minutes after irradiation, cell viability was examined by checking esterase activity and membrane permeability to find that all cells in the irradiated area were effectively killed, leaving neighboring cells perfectly intact. Further, we fabricated another type of culture substrate by introducing the PAG polymer and copolymer containing 4–vinylpyridine on the culture surface of a polystyrene dish in order to implement photo–induced cell detachment. NIH/3T3 cells were cultured on this surface until confluent, and the blue light was irradiated locally within a circular area of 5 mm diameter for a few minutes. In a few hours after irradiation, we observed that most of the cells became detached from the irradiated area while no detectable change was found for the cells outside of the area. We collected the detached cells, and examined their viability by checking membrane permeability. As a result, about 85 % of the cells were estimated to maintain their viability even after photo-induced detachment. Also the detachment of HepG2 adherently cultured on the culture substrate was induced by the light irradiation.

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Microfluidic technologies are capable of producing small materials such as particles, fibers, and sheets with sizes of several hundred micrometers. In this paper, we briefly introduce our recent progresses on the fabrication of hydrogel materials for tissue engineering using microfluidic devices. The first topic is the production of thin, stripepatterned sheets made of alginate hydrogel, having soft and solid regions. These functional hydrogel sheets were used to generate heterotypic cellular constructs composed of hepatocytes and non-parenchymal cells, for the purpose of upregulating hepatic functions. The second topic is the formation of vascular–tissue like tubular structures composed of alginate hydrogel and two types of cells, within hydrogel microchannels made of agarose hydrogel. These studies are good examples of the application of microfluidic devices to the production of thin hydrogel materials that are potentially useful for various biological research and tissue engineering.

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Various forms of topical hemostatic agents have been widely used in surgical procedures. Above all, membranetype hemostatic agents are most frequently used in clinical practice because they are easy to handle and work by application of pressure. We developed a carboxymethyl cellulose (CMC) nonwoven sheet as a hemostatic agent by carboxymethylating a continuous filament cellulose nonwoven sheet. The CMC nonwoven sheet was able to absorb water and dissolve in it. The rates of absorption and dissolution depended on the degree of carboxymethylation. After dissolving in blood, CMC accelerated clot development (possibly owing to the incorporation of CMC into fibrin fibers) and increased the viscosity of the blood, both of which would contribute to the improved blood clotting of an injured surface. In vivo experiments using a rat tail cutting method showed that a CMC nonwoven sheet shortened the bleeding time of the tail when applied to the cut surface. These results suggest that a CMC nonwoven sheet could be used as a novel membrane-type hemostatic agent.

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Postoperative peritoneal adhesions cause pelvic pain, bowel obstruction, and infertility. Repeated hepatic resections with presence of adhesions induced by previous hepatectomies are time–consuming and hazardous. We recently developed a new hepatectomy–induced postoperative adhesion rat model to evaluate the anti-adhesion efficacy of commercially available sheet materials (Seprafilm^® and Interceed^®) and the recently reported hyaluronan–based in situ cross–linkable hydrogels. Adhesions between the liver surface and the diaphragm and adhesions around the hepatic hilum were less severe, but were not remarkably reduced by the anti-adhesion materials.

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We present a numerical method to simulate the permeation behavior of oil–in–water (O/W) emulsions with soluble surfactants through a membrane pore. Our numerical frame work is based on the phase–field model, which enables us to capture the thermodynamic and hydrodynamic effects of surfactants on oil droplet motion. We investigated the effects of surfactant concentration on the permeation behavior of oil droplets. Existence of surfactants made demulsification behavior complicated because oil droplets covered with surfactants are deformed easily by permeation flow.

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The patients in an advanced renal failure are generally treated with a dialyzer as a substitute for their kidney. The antithrombogenicity is highly required to the hollow fiber membrane inside a dialyzer. We developed “Toraylight” NV which was improved antithrombogenicity. There are three viewpoints, that is, optimizing the header shape of the dialyzer, improvement of the quality of the hollow fiber membrane form and the antithrombogenicity of hollow fiber membranes.
Applying the antithrombogenic polymer (NV polymer) to the PSf membrane surface in the nanometer order, we created an extremely antithrombogenic hollow fiber membrane for hemodialysis, which made the number of the platelet adhesion less than a hundredth compared with a conventional PSf membrane with PVP polymer.

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