Nobuhiro Kato

Selection of high-potential embryos by culture in poly(dimethylsiloxane) microwells and time-lapse imaging

OBJECTIVE To assess the developmental kinetics of human embryos and their ability to develop to morphologically normal blastocysts. DESIGN Experimental study on human embryos donated for research using a time-lapse imaging system based on individual embryo culture in poly(dimethylsiloxane) microwells and monitored using a microscope inside the incubator. SETTING Private fertility clinic. PATIENT(S) Surplus embryos donated by couples after undergoing fertility treatment. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Blastocyst score and times required from beginning to completion of the second and third mitotic divisions. RESULT(S) The time required for completion of the second division (the three- to four-cell stage) was shorter in embryos that developed to high-scoring blastocysts (0.7 hours, n = 17) than in those forming low-scoring blastocysts (3.7 hours, n = 24). Similarly, the mean time required to completion of the third division (five- to eight-cell stage) was also significantly shorter in embryos forming high-scoring blastocysts (5.7 hours) than among those forming low-scoring blastocysts (16.9 hours). CONCLUSION(S) Individual embryos with the potential to develop to high-scoring blastocysts could be selected at 2-3 days of culture using this system by examining the times required to complete the second and third mitotic divisions.

Individually programmable cell stretching microwell arrays actuated by a Braille display

Cell culture systems are often static and are therefore nonphysiological. In vivo, many cells are exposed to dynamic surroundings that stimulate cellular responses in a process known as mechanotransduction. To recreate this environment, stretchable cell culture substrate systems have been developed, however, these systems are limited by being macroscopic and low throughput. We have developed a device consisting of 24 miniature cell stretching chambers with flexible bottom membranes that are deformed using the computer-controlled, piezoelectrically actuated pins of a Braille display. We have also developed efficient image capture and analysis protocols to quantify morphological responses of the cells to applied strain. Human dermal microvascular endothelial cells (HDMECs) were found to show increasing degrees of alignment and elongation perpendicular to the radial strain in response to cyclic stretch at increasing frequencies of 0.2, 1, and 5 Hz, after 2, 4, and 12h. Mouse myogenic C2C12 cells were also found to align in response to the stretch, while A549 human lung adenocarcinoma epithelial cells did not respond to stretch.

Resonance Characteristics Of Micro Cantilever In Liquid

The resonance characteristics of a micro cantilever are experimentally and theoretically studied. The micro cantilever is vibrated by thermal excitation using laser exposure and the frequency responses of the micro cantilever in air, acetone and water are experimentally evaluated based on an optical measurement technique. It is demonstrated that the resonance frequencies in water decrease to about one-tenth of the values in air. Also, the frequency responses are calculated using the string of beads model taking into consideration the drag force by viscous fluids. The calculation results show good agreement with the experimental results.

Force-balancing microforce sensor with an optical-fiber interferometer

A microforce sensor with a force feedback method for scanning force microscopy is presented. The force sensor is constructed by using an optical fiber and a microcantilever. The facet of the optical fiber is coated with a gold thin film 15 A thick. This gold film acts not only as a partially reflected mirror but also as an electrode of the electrostatic actuator. The interaction force between a probe tip and a sample is balanced by the electrostatic force. The deflection of the cantilever is measured by an interferometer consisting of the facet of the optical fiber and the cantilever surface. We have made the force sensor and measured some force curves of a mica surface with a SiN probe tip. The force curves obtained by this sensor are quite different from the curves by a conventional measurement without a feedback system. The force resolution of the system is 10−10 N with a bandwidth of dc∼1 kHz.

Force‐balancing force sensor with an optical lever

Scanning force microscopes (SFMs) are sometimes used to obtain a force curve, which shows the force variation as a function of tip–sample distance. In the force curve measurement, if the spring constant of the force detecting lever is small, the measured force curve has discontinuity and is different from the true force curve. In this paper, we present a new type of force balancing force sensor built in SFM. This force sensor employs an optical lever for detecting the rotation of the lever and two electrostatic force actuators with transparent electrodes. This sensor has two operating modes; with and without feedback. In the feedback mode, the force detecting lever is balanced with the electrostatic force. This system has the effect of enlarging the effective spring constant of the whole sensor. In the nonfeedback mode, this sensor acts as an ordinary force sensor. By using this sensor in both modes we will show the effectiveness of the force feedback in force curve measurements.

Formation of Hydroxyapatite Thin Films on Surface-Modified Polytetrafluoroethylene Substrates

An ArF excimer laser deposition technique is used to fabricate hydroxyapatite [Ca10(PO4)6(OH)2]-(HAp) thin films on surface-modified polytetrafluoroethylene (PTFE) substrates. The surface of PTFE is modified by a sodium-naphthalene complex in a glycol ether solvent. X-ray diffraction (XRD) studies indicate that as-deposited films prepared at substrate temperatures below the glass transition temperature of PTFE (327°C) are in an amorphous state. The crystallization of as-deposited films needed annealing for 10 h at 310°C. The structure, surface morphology and bond strength of HAp films were evaluated using XRD, the atomic force microscopy (AFM) and the Instron testing machine. The tensile bond strength of HAp films on surface-modified PTFE was 6.0 MPa, which is one order of magnitude larger than that of films on non-surface-modified PTFE, demonstrating its potential for practical applications.

System analysis of the force-feedback method for force curve measurements

The force-feedback method is a promising technique to measure accurate force curves in atomic force spectroscopy. In this article, we describe criteria of the stability for the force-feedback system containing a proportional amplifier with first-order lag. We derive the criteria from an equation of the cantilever movement with higher-order vibration modes under force interaction. The criteria predict that a proportional amplifier with a certain gain and a “very high” cutoff frequency is required to stabilize the feedback system when the critical force gradient exceeds the cantilever stiffness. To measure a force curve, including the steep part (−17 N/m) with a soft (0.2 N/m) and poor-damping (2×10−7 kg/s) cantilever, the amplifier for the force-feedback controller must have a frequency response wider than 16 MHz and gain larger than 16.8. Moreover, the feedback controller with only an integrator, which is popular in scanning tunneling microscopy and atomic force microscopy, does not contribute to reducing...

Regeneration of Tooth Enamel by Flexible Hydroxyapatite Sheet

Before a tooth erupts, the ameloblasts are lost, which means that the tooth enamel does not regenerate itself after tooth eruption. In the present study, we attempt to regenerate the tooth enamel artificially using a flexible hydroxyapatite (HAp) sheet. First, a HAp film was deposited on a soluble substrate by pulsed laser deposition (PLD) using an ArF excimer laser. Next, the HAp film was collected as a freestanding sheet by dissolving the substrate using a solvent. The HAp sheet was adhered to the extracted human teeth using a calcium phosphate solution. The variation of the crystal structure of the HAp sheet with time was investigated by X-ray diffraction analysis. Furthermore, the variation in the mechanical characteristics with time between the HAp sheet and dental enamel were evaluated using tensile and scratch tests. The results suggest that the HAp sheet became fused to the tooth enamel within approximately one week.

Reduction in Feedback Bandwidth of the Force-Controlled Atomic Force Microscope Using a Polyimide Cantilever

In this paper we present a method of reducing the bandwidth required in the force-controlled atomic force microscope (AFM) for measuring the force curves. System stability strongly depends on the quality factor of vibration (Q) of the cantilever. As polyimide has a low Q, a cantilever fabricated from it reduces the bandwidth required in the feedback system. A polyimide cantilever with dimensions of 540×155×3.4 µm3 is fabricated to evaluate its Q in vacuum, density and Youngs modulus. The measured Q is 50, which is much smaller than those of conventional cantilevers used for AFM. Using the measured mechanical properties of the polyimide, the required frequency response of the feedback system is estimated. Typical force curve measurements are confirmed by some numerical simulations.

Adhesive Strength between Flexible Hydroxyapatite Sheet and Tooth Enamel

Tooth enamel cannot be reconstructed once it is destroyed immoderately. Hydroxyapatite (HAp) thin sheet can potentially be used for a novel dental biomaterial to repair the enamel. Using a pulsed laser deposition (PLD) method, we have successfully created a flexible HAp sheet of less than a few micrometers in thickness. Due to its flexibility, the HAp sheet is tightly adhered on curved surfaces at the target site. In the present study, we newly developed double-layered sheets composed of HAp film coated with tricalcium phosphate (TCP) thin layer. The HAp/TCP sheet was adhered to the extracted human teeth using a calcium phosphate solution for 3 days. The adhesive strength between the HAp/TCP sheet and tooth enamel was evaluated by quasi-static tensile tests. Moreover, the interface structure between them was observed by a scanning electron microscopy. As a result of the mechanical evaluation, the adhesive strength was greater than approximately 2.5 MPa. The electron microscopic observation revealed that the sheet was partially fused with the enamel. These findings suggest the possibility that enamel defects are repaired using the HAp/TCP sheet for a short duration.