Responsiveness of voltage-gated calcium channels in SH-SY5Y human neuroblastoma cells on quasi-three-dimensional micropatterns formed with poly (l-lactic acid)

Introduction: In this study, quasi-three-dimensional (3D) microwell patterns were fabricated with poly (l-lactic acid) for the development of cell-based assays, targeting voltage-gated calcium channels (VGCCs). Methods and materials: SH-SY5Y human neuroblastoma cells were interfaced with the microwell patterns and found to grow as two dimensional (2D), 3D, and near two dimensional (N2D), categorized on the basis of the cells’ location in the pattern. The capability of the microwell patterns to support 3D cell growth was evaluated in terms of the percentage of the cells in each growth category. Cell spreading was analyzed in terms of projection areas under light microscopy. SH-SY5Y cells’ VGCC responsiveness was evaluated with confocal microscopy and a calcium fluorescent indicator, Calcium GreenTM-1. The expression of L-type calcium channels was evaluated using immunofluorescence staining with DM-BODIPY. Results: It was found that cells within the microwells, either N2D or 3D, showed more rounded shapes and less projection areas than 2D cells on flat poly (l-lactic acid) substrates. Also, cells in microwells showed a significantly lower VGCC responsiveness than cells on flat substrates, in terms of both response magnitudes and percentages of responsive cells, upon depolarization with 50 mM K+. This lower VGCC responsiveness could not be explained by the difference in L-type calcium channel expression. For the two patterns addressed in this study, N2D cells consistently exhibited an intermediate value of either projection areas or VGCC responsiveness between those for 2D and 3D cells, suggesting a correlative relation between cell morphology and VGCC responsiveness. Conclusion: These results suggest that the pattern structure and therefore the cell growth characteristics were critical factors in determining cell VGCC responsiveness and thus provide an approach for engineering cell functionality in cell-based assay systems and tissue engineering scaffolds.

A model for continuous measurement of drilled shaft diameter during construction

Non-Destructive Testing (NDT) of deep foundations has become an integral part of the industry's standard manufacturing processes. It is not unusual for the evaluation of the integrity of the concrete to include the measurement of ultrasonic wave speeds. Numerous methods have been proposed that use the propagation speed of ultrasonic waves to check the integrity of concrete for drilled shaft foundations. All such methods evaluate the integrity of the concrete inside the cage and between the access tubes. The integrity of the concrete outside the cage remains to be considered to determine the location of the border between the concrete and the soil in order to obtain the diameter of the drilled shaft. It is also economic to devise a methodology to obtain the diameter of the drilled shaft using the Cross-Hole Sonic Logging system (CSL). Performing such a methodology using the CSL and following the CSL tests is performed and used to check the integrity of the inside concrete, thus allowing the determination of the drilled shaft diameter without having to set up another NDT device.^ This proposed new method is based on the installation of galvanized tubes outside the shaft across from each inside tube, and performing the CSL test between the inside and outside tubes. From the performed experimental work a model is developed to evaluate the relationship between the thickness of concrete and the ultrasonic wave properties using signal processing. The experimental results show that there is a direct correlation between concrete thicknesses outside the cage and maximum amplitude of the received signal obtained from frequency domain data. This study demonstrates how this new method to measuring the diameter of drilled shafts during construction using a NDT method overcomes the limitations of currently-used methods. ^ In the other part of study, a new method is proposed to visualize and quantify the extent and location of the defects. It is based on a color change in the frequency amplitude of the signal recorded by the receiver probe in the location of defects and it is called Frequency Tomography Analysis (FTA). Time-domain data is transferred to frequency-domain data of the signals propagated between tubes using Fast Fourier Transform (FFT). Then, distribution of the FTA will be evaluated. This method is employed after CSL has determined the high probability of an anomaly in a given area and is applied to improve location accuracy and to further characterize the feature. The technique has a very good resolution and clarifies the exact depth location of any void or defect through the length of the drilled shaft for the voids inside the cage. ^ The last part of study also evaluates the effect of voids inside and outside the reinforcement cage and corrosion in the longitudinal bars on the strength and axial load capacity of drilled shafts. The objective is to quantify the extent of loss in axial strength and stiffness of drilled shafts due to presence of different types of symmetric voids and corrosion throughout their lengths.^

A Model for Continuous Measurement of Drilled Shaft Diameter During Construction

Non-Destructive Testing (NDT) of deep foundations has become an integral part of the industry’s standard manufacturing processes. It is not unusual for the evaluation of the integrity of the concrete to include the measurement of ultrasonic wave speeds. Numerous methods have been proposed that use the propagation speed of ultrasonic waves to check the integrity of concrete for drilled shaft foundations. All such methods evaluate the integrity of the concrete inside the cage and between the access tubes. The integrity of the concrete outside the cage remains to be considered to determine the location of the border between the concrete and the soil in order to obtain the diameter of the drilled shaft. It is also economic to devise a methodology to obtain the diameter of the drilled shaft using the Cross-Hole Sonic Logging system (CSL). Performing such a methodology using the CSL and following the CSL tests is performed and used to check the integrity of the inside concrete, thus allowing the determination of the drilled shaft diameter without having to set up another NDT device. This proposed new method is based on the installation of galvanized tubes outside the shaft across from each inside tube, and performing the CSL test between the inside and outside tubes. From the performed experimental work a model is developed to evaluate the relationship between the thickness of concrete and the ultrasonic wave properties using signal processing. The experimental results show that there is a direct correlation between concrete thicknesses outside the cage and maximum amplitude of the received signal obtained from frequency domain data. This study demonstrates how this new method to measuring the diameter of drilled shafts during construction using a NDT method overcomes the limitations of currently-used methods. In the other part of study, a new method is proposed to visualize and quantify the extent and location of the defects. It is based on a color change in the frequency amplitude of the signal recorded by the receiver probe in the location of defects and it is called Frequency Tomography Analysis (FTA). Time-domain data is transferred to frequency-domain data of the signals propagated between tubes using Fast Fourier Transform (FFT). Then, distribution of the FTA will be evaluated. This method is employed after CSL has determined the high probability of an anomaly in a given area and is applied to improve location accuracy and to further characterize the feature. The technique has a very good resolution and clarifies the exact depth location of any void or defect through the length of the drilled shaft for the voids inside the cage. The last part of study also evaluates the effect of voids inside and outside the reinforcement cage and corrosion in the longitudinal bars on the strength and axial load capacity of drilled shafts. The objective is to quantify the extent of loss in axial strength and stiffness of drilled shafts due to presence of different types of symmetric voids and corrosion throughout their lengths.