InTech Acta Medica Okayama 2012 Use of Silicone Elastomer-Based Microfluidic Devices and Systems in Reproductive Technologies 243 262 EN Koji Matsuura Keiji Naruse No potential conflict of interest relevant to this article was reported.

Optical Society of America Acta Medica Okayama 2156-7085 3 2 2012 Scanning and non-scanning surface plasmon microscopy to observe cell adhesion sites 354 359 EN Koyo Watanabe Koji Matsuura Fukukazu Kawata Kotaro Nagata Jun Ning Hiroshi Kano We observe adhesion sites of a cell on a substrate with high resolution. Since this observation requires interfacial measurements between the cell and the substrate, we employ scanning localized surface plasmon microscopy. We experimentally show that focal adhesion sites of a mouse muscle cell can be observed without fluorescent labeling. We also show that a non-scanning surface plasmon microscope combined with the scanning localized surface plasmon microscope contributes to observing an entire cell adhesion site and identify regions of interest. No potential conflict of interest relevant to this article was reported.

Hindawi Publishing Corporation Acta Medica Okayama 2012 2012 Theoretical Study on the Detection of Tilted Lipid Bilayers Using Surface Plasmon Resonance Techniques EN Koyo Watanabe Koji Matsuura Effective refractive indices detected using surface plasmon resonance techniques are calculated as a function of the tilt angle of lipid bilayers in a multilayered model. The changes in the effective refractive indices are derived from the shift of an excitation angle of surface plasmon. To obtain effective refractive index plots, we determined refractive index changes in the lipid bilayers with 3 and 5 nm thicknesses as a function of tilt angle and obtained a relationship between the effective refractive index and tilt angle. We also showed that the effective refractive index depended on the lipid bilayers thickness and anisotropic permittivities, which vary with interchain distance. No potential conflict of interest relevant to this article was reported.

Society for Reproduction and Development Acta Medica Okayama 0916-8818 57 1 2010 Hydrophobic Silicone Elastomer Chamber for Recording Trajectories of Motile Porcine Sperms without Adsorption 163 167 EN Koji Matsuura Yuka Kuroda Keisuke Yamashita Hiroaki Funahashi Motile porcine sperms adhere to hydrophilic materials such as glass and plastics. The adsorption of sperms to a hydrophobic poly(dimethylsiloxane) (PDMS) membrane is less compared with that to glass. We investigated the linear velocity (LV) and amplitude of lateral head displacement (ALHD) of motile porcine sperm on glass and PDMS preparations using computer-assisted sperm analysis (CASA). Significant decreases were observed in the 15-min LV (P<0.05) and ALHD (P<0.05) in motile porcine sperm on glass preparations compared with those on PDMS preparations. These differences were due to adsorption of the head and/or neck to hydrophilic substrates. Because of the elasticity of PDMS, we propose that a PDMS membrane should be used for CASA. To investigate the dynamics of motile porcine sperms with microfluidics, we do not recommend plasma treatment to bond PDMS and glass in the microchannel preparation; instead, we suggest that a PDMS molding process without plasma treatment be used for preparation of microfluidic channels. No potential conflict of interest relevant to this article was reported.

Elsevier Acta Medica Okayama 1472-6483 24 1 2012 Screening of sperm velocity by fluid mechanical characteristics of a cyclo-olefin polymer microfluidic sperm-sorting device 109 115 EN Koji Matsuura Mami Takenami Yuka Kuroda Toru Hyakutake Shinichiro Yanase Keiji Naruse The microfluidic sperm-sorting (MFSS) device is a promising advancement for assisted reproductive technology. Previously, poly(dimethylsiloxiane) and quartz MFSS devices were developed and used for intracytoplasmic sperm injection. However, these disposable devices were not clinically suitable for assisted reproduction, so a cyclo-olefin polymer MFSS (COP-MFSS) device was developed. By micromachining, two microfluidic channels with different heights and widths (chip A: 0.3 x 0.5 mm; chip B: 0.1 x 0.6 mm) were prepared. Sorted sperm concentrations were similar in both microfluidic channels. Linear-velocity distribution using the microfluidic channel of chip B was higher than that of chip A. Using confocal fluorescence microscopy, it was found that the highest number of motile spermatozoa swam across the laminar flow at the bottom of the microfluidic channel. The time required to swim across the laminar flow was longer at the bottom and top of the microfluidic channels than in the middle because of the low fluid velocity. These results experimentally demonstrated that the width of microfluidic channels should be increased in the region of laminar flow from the semen inlet to the outlet for unsorted spermatozoa to selectively recover spermatozoa with high linear velocity. No potential conflict of interest relevant to this article was reported.

Elsevier Acta Medica Okayama 0015-0282 94 3 2010 Blastocyst quality scoring based on morphologic grading correlates with cell number 1135 1137 EN Koji Matsuura Nobuyoshi Hayashi Chisato Takiue Rei Hirata Toshihiro Habara Keiji Naruse Blastocyst quality score (BQS), first reported by Rehman et al., is a numerical blastocyst-morphology grading system based on the criteria established by Gardner and Schoolcraft. We demonstrate a positive correlation between the calculated BQS score and cell number by staining thawed human embryos and suggest that BQS can be applied to evaluate culture systems clinically. No potential conflict of interest relevant to this article was reported.

Elsevier Acta Medica Okayama 0093-691X 74 5 2010 Application of a microfluidic sperm sorter to the in-vitro fertilization of porcine oocytes reduced the incidence of polyspermic penetration 863 870 EN Hikaru Sano Koji Matsuura Keiji Naruse Hiroaki Funahashi The objective of this study was to use a microfluidic sperm sorter (MFSS), designed to isolate motile human spermatozoa with laminar flows (no centrifugation), for porcine IVF. Boar spermatozoa were diluted at 1 x 10(8) with a diluent containing 20% seminal fluid and flowed with modified TCM-199 (mM199, with 5 mM caffeine) to introduce motile sperm into the exit chamber for IVF. In Experiment 1, after flowing for 5 min, sperm concentration varied significantly among specific sites within the MFSS collecting chamber (range, 0.8 +/- 0.5 x 10(4) to 575.0 +/- 56.3 x 10(4) cells/mL; mean +/- SEM). In Experiment 2, when porcine IVM oocytes were placed at three locations in the MFSS exit chamber (where only motile spermatozoa accumulated) and subsequently cultured in caffeine-free mM199 for 8 h, sperm penetration rate was not significantly different among places (86.1 +/- 10.5 to 100%), but the monospermic penetration rate was lower (P < 0.05) in oocytes 3.5 mm from the exit position (12.5 +/- 4.8%) than those at 7.5 mm (53.1 +/- 6.0%) or further (41.9 +/- 2.8%) from the exit. In Experiment 3, the normal fertilization index (ratio of monospermic oocytes to number of oocytes examined) 8 h after insemination was higher (P < 0.05) in the MFSS-IVF system (0.375 +/- 0.040) than both standard IVF and transient IVF (0.222 +/- 0.028 and 0.189 +/- 0.027, respectively, with co-culture for 8 h and for 5 min). Developmental competence of fertilized oocytes (blastocyst formation) was higher (P < 0.05) in the MFSS-IVF system (40.9 +/- 2.3%) than in either standard or transient IVF (22.6 +/- 1.4 and 33.7 +/- 3.5%). In conclusion, brief co-culture of porcine oocytes with spermatozoa gradually accumulated in the MFSS chamber improved the efficiency of producing monospermic fertilized embryos and blastocysts. Furthermore, efficiencies were significantly affected by oocyte location within the chamber. No potential conflict of interest relevant to this article was reported.

Society for Reproduction and Development Acta Medica Okayama 0916-8818 56 5 2010 In-vitro Culture with a Tilting Device in Chemically Defined Media During Meiotic Maturation and Early Development Improves the Quality of Blastocysts Derived from In-vitro Matured and Fertilized Porcine Oocytes 552 557 EN Takayuki Koike Koji Matsuura Keiji Naruse Hiroaki Funahashi Under physiological conditions, mammalian oocytes and embryos appear to be stimulated not only chemically but also mechanically, such as by compression, shear stress and/or friction force in the follicle and female reproductive tract. The present study was undertaken to examine the effects of kinetic culture with a tilting device in chemically defined media during in vitro maturation (IVM) of porcine oocytes and in vitro culture (IVC) following in vitro fertilization (IVF) on the early developmental competence and quality of blastocysts. After culture in a chemically defined IVM medium, modified porcine oocyte medium (mPOM) containing gonadotropins and dibutyryl cAMP for 20 h, the mean diameter of the cumulus-oocyte complexes (COCs) was larger in the tilting culture than in the static controls, whereas the diameter of the oocytes did not differ. When culture of the COCs was continued additionally in a fresh medium without gonadotropins and dibutyryl cAMP for 24 h, the incidences of oocytes completing GVBD and developing to the metaphase-II stage did not differ between the tilting and static culture systems. Furthermore, the sperm penetration after IVF and developmental competence of the oocytes to the blastocyst stage were not different between the tilting and static systems during IVM and IVC. However, tilting culture during both IVM and IVC had a significant positive effect on the number of cells per blastocyst (P<0.05). These observations indicate that tilting culture during IVM and IVC in chemically defined media improves the quality of blastocyst, as determined by the number of cells per blastocyst, without any effects on penetrability and developmental competence. No potential conflict of interest relevant to this article was reported.

Springer US Acta Medica Okayama 1387-2176 11 1 2009 Application of a numerical simulation to improve the separation efficiency of a sperm sorter 25 33 EN Toru Hyakutake Yuki Hashimoto Shinichiro Yanase Koji Matsuura Keiji Naruse This paper describes a study in which numerical simulations were applied to improve the separation efficiency of a microfluidic-based sperm sorter. Initially, the motion of 31 sperm were modeled as a sinusoidal wave. The modeled sperm were expected to move while vibrating in the fluid within the microchannel. In this analysis, the number of sperm extracted at the outlet channel and the rate of movement of the highly motile sperm were obtained for a wide range of flow velocities within the microchannel. By varying the channel height, and the width and the position of the sperm-inlet channel, we confirmed that the separation efficiency was highly dependent on the fluid velocity within the channel. These results will be valuable for improving the device configuration, and might help to realize further improvements in efficiency in the future. No potential conflict of interest relevant to this article was reported.

The Royal Society of Chemistry Acta Medica Okayama 1473-0197 9 22 2009 Fabricating small-scale, curved, polymeric structures with convex and concave menisci through interfacial free energy equilibrium 3306 3309 EN Chao-Min Cheng Koji Matsuura I-Jan Wang Yuka Kuroda Philip R. LeDuc Keiji Naruse Polymeric curved structures are widely used in imaging systems including optical fibers and microfluidic channels. Here, we demonstrate that small-scale, poly(dimethylsiloxane) (PDMS)-based, curved structures can be fabricated through controlling interfacial free energy equilibrium. Resultant structures have a smooth, symmetric, curved surface, and may be convex or concave in form based on surface tension balance. Their curvatures are controlled by surface characteristics (i.e., hydrophobicity and hydrophilicity) of the molds and semi-liquid PDMS. In addition, these structures are shown to be biocompatible for cell culture. Our system provides a simple, efficient and economical method for generating integrateable optical components without costly fabrication facilities. No potential conflict of interest relevant to this article was reported.

Elsevier Acta Medica Okayama 1472-6483 20 3 2010 Improved development of mouse and human embryos using a tilting embryo culture system 358 364 EN Koji Matsuura Nobuyoshi Hayashi Yuka Kuroda Chisato Takiue Rei Hirata Mami Takenami Yoko Aoi Nanako Yoshioka Toshihiro Habara Tetsunori Mukaida Keiji Naruse Mammalian embryos experience not only hormonal but also mechanical stimuli, such as shear stress, compression and friction force in the Fallopian tube before nidation. In order to apply mechanical stimuli to embryos in a conventional IVF culture system, the tilting embryo culture system (TECS) was developed. The observed embryo images from the TECS suggest that the velocities and shear stresses of TECS embryos are similar to those experienced in the oviduct. Use of TECS enhanced the development rate to the blastocyst stage and significantly increased the cell number of mouse blastocysts (P < 0.05). Although not statistically significant, human thawed embryos showed slight improvement in development to the blastocyst stage following culture in TECS compared with static controls. Rates of blastocyst formation following culture in TECS were significantly improved in low-quality embryos and those embryos cultured under suboptimal conditions (P < 0.05). The TECS is proposed as a promising approach to improve embryo development and blastocyst formation by exposing embryos to mechanical stimuli similar to those in the Fallopian tube. No potential conflict of interest relevant to this article was reported.