Rapid Prototyping of 3D Biochips for Cell Motility Studies Using Two-Photon Polymerization

The study of cellular migration dynamics and strategies plays a relevant role in the understanding of both physiological and pathological processes. An important example could be the link between cancer cell motility and tumor evolution into metastatic stage. These strategies can be strongly influenced by the extracellular environment and the consequent mechanical constrains. In this framework, the possibility to study the behavior of single cells when subject to specific topological constraints could be an important tool in the hands of biologists. Two-photon polymerization is a sub-micrometric additive manufacturing technique that allows the fabrication of 3D structures in biocompatible resins, enabling the realization of ad hoc biochips for cell motility analyses, providing different types of mechanical stimuli. In our work, we present a new strategy for the realization of multilayer microfluidic lab-on-a-chip constructs for the study of cell motility which guarantees complete optical accessibility and the possibility to freely shape the migration area, to tailor it to the requirements of the specific cell type or experiment. The device includes a series of micro-constrictions that induce different types of mechanical stress on the cells during their migration. We show the realization of different possible geometries, in order to prove the versatility of the technique. As a proof of concept, we present the use of one of these devices for the study of the motility of murine neuronal cancer cells under high physical confinement, highlighting their peculiar migration mechanisms

Intermediate filaments ensure resiliency of single carcinoma cells, while active contractility of the actin cortex determines their invasive potential

During the epithelial-to-mesenchymal transition, the intracellular cytoskeleton undergoes severe reorganization which allows epithelial cells to transition into a motile mesenchymal phenotype. Among the different cytoskeletal elements, the intermediate filaments keratin (in epithelial cells) and vimentin (in mesenchymal cells) have been demonstrated to be useful and reliable histological markers. In this study, we assess the potential invasiveness of six human breast carcinoma cell lines and two mouse fibroblasts cells lines through single cell migration assays in confinement. We find that the keratin and vimentin networks behave mechanically the same when cells crawl through narrow channels and that vimentin protein expression does not strongly correlate to single cells invasiveness. Instead, we find that what determines successful migration through confining spaces is the ability of cells to mechanically switch from a substrate-dependent stress fibers based contractility to a substrate-independent cortical contractility, which is not linked to their tumor phenotype

Search for eccentric black hole coalescences during the third observing run of LIGO and Virgo

Despite the growing number of confident binary black hole coalescences observed through gravitational waves so far, the astrophysical origin of these binaries remains uncertain. Orbital eccentricity is one of the clearest tracers of binary formation channels. Identifying binary eccentricity, however, remains challenging due to the limited availability of gravitational waveforms that include effects of eccentricity. Here, we present observational results for a waveform-independent search sensitive to eccentric black hole coalescences, covering the third observing run (O3) of the LIGO and Virgo detectors. We identified no new high-significance candidates beyond those that were already identified with searches focusing on quasi-circular binaries. We determine the sensitivity of our search to high-mass (total mass M>70 M⊙) binaries covering eccentricities up to 0.3 at 15 Hz orbital frequency, and use this to compare model predictions to search results. Assuming all detections are indeed quasi-circular, for our fiducial population model, we place an upper limit for the merger rate density of high-mass binaries with eccentricities 0<e≤0.3 at 0.33 Gpc−3 yr−1 at 90\% confidence level

Ultralight vector dark matter search using data from the KAGRA O3GK run

Among the various candidates for dark matter (DM), ultralight vector DM can be probed by laser interferometric gravitational wave detectors through the measurement of oscillating length changes in the arm cavities. In this context, KAGRA has a unique feature due to differing compositions of its mirrors, enhancing the signal of vector DM in the length change in the auxiliary channels. Here we present the result of a search for U(1)B−L gauge boson DM using the KAGRA data from auxiliary length channels during the first joint observation run together with GEO600. By applying our search pipeline, which takes into account the stochastic nature of ultralight DM, upper bounds on the coupling strength between the U(1)B−L gauge boson and ordinary matter are obtained for a range of DM masses. While our constraints are less stringent than those derived from previous experiments, this study demonstrates the applicability of our method to the lower-mass vector DM search, which is made difficult in this measurement by the short observation time compared to the auto-correlation time scale of DM

Search for gravitational-lensing signatures in the full third observing run of the LIGO-Virgo network

Gravitational lensing by massive objects along the line of sight to the source causes distortions of gravitational wave-signals; such distortions may reveal information about fundamental physics, cosmology and astrophysics. In this work, we have extended the search for lensing signatures to all binary black hole events from the third observing run of the LIGO--Virgo network. We search for repeated signals from strong lensing by 1) performing targeted searches for subthreshold signals, 2) calculating the degree of overlap amongst the intrinsic parameters and sky location of pairs of signals, 3) comparing the similarities of the spectrograms amongst pairs of signals, and 4) performing dual-signal Bayesian analysis that takes into account selection effects and astrophysical knowledge. We also search for distortions to the gravitational waveform caused by 1) frequency-independent phase shifts in strongly lensed images, and 2) frequency-dependent modulation of the amplitude and phase due to point masses. None of these searches yields significant evidence for lensing. Finally, we use the non-detection of gravitational-wave lensing to constrain the lensing rate based on the latest merger-rate estimates and the fraction of dark matter composed of compact objects

Rapid Prototyping of 3D Biochips for Cell Motility Studies Using Two-Photon Polymerization

The study of cellular migration dynamics and strategies plays a relevant role in the understanding of both physiological and pathological processes. An important example could be the link between cancer cell motility and tumor evolution into metastatic stage. These strategies can be strongly influenced by the extracellular environment and the consequent mechanical constrains. In this framework, the possibility to study the behavior of single cells when subject to specific topological constraints could be an important tool in the hands of biologists. Two-photon polymerization is a sub-micrometric additive manufacturing technique that allows the fabrication of 3D structures in biocompatible resins, enabling the realization of ad hoc biochips for cell motility analyses, providing different types of mechanical stimuli. In our work, we present a new strategy for the realization of multilayer microfluidic lab-on-a-chip constructs for the study of cell motility which guarantees complete optical accessibility and the possibility to freely shape the migration area, to tailor it to the requirements of the specific cell type or experiment. The device includes a series of micro-constrictions that induce different types of mechanical stress on the cells during their migration. We show the realization of different possible geometries, in order to prove the versatility of the technique. As a proof of concept, we present the use of one of these devices for the study of the motility of murine neuronal cancer cells under high physical confinement, highlighting their peculiar migration mechanisms

Intermediate filaments ensure resiliency of single carcinoma cells, while active contractility of the actin cortex determines their invasive potential

During the epithelial-to-mesenchymal transition, the intracellular cytoskeleton undergoes severe reorganization which allows epithelial cells to transition into a motile mesenchymal phenotype. Among the different cytoskeletal elements, the intermediate filaments keratin (in epithelial cells) and vimentin (in mesenchymal cells) have been demonstrated to be useful and reliable histological markers. In this study, we assess the potential invasiveness of six human breast carcinoma cell lines and two mouse fibroblasts cells lines through single cell migration assays in confinement. We find that the keratin and vimentin networks behave mechanically the same when cells crawl through narrow channels and that vimentin protein expression does not strongly correlate to single cells invasiveness. Instead, we find that what determines successful migration through confining spaces is the ability of cells to mechanically switch from a substrate-dependent stress fibers based contractility to a substrate-independent cortical contractility, which is not linked to their tumor phenotype

Intermediate filaments ensure resiliency of single carcinoma cells, while active contractility of the actin cortex determines their invasive potential

During the epithelial-to-mesenchymal transition, the intracellular cytoskeleton undergoes severe reorganization which allows epithelial cells to transition into a motile mesenchymal phenotype. Among the different cytoskeletal elements, the intermediate filaments keratin (in epithelial cells) and vimentin (in mesenchymal cells) have been demonstrated to be useful and reliable histological markers. In this study, we assess the potential invasiveness of six human breast carcinoma cell lines and two mouse fibroblasts cells lines through single cell migration assays in confinement. We find that the keratin and vimentin networks behave mechanically the same when cells crawl through narrow channels and that vimentin protein expression does not strongly correlate to single cells invasiveness. Instead, we find that what determines successful migration through confining spaces is the ability of cells to mechanically switch from a substrate-dependent stress fibers based contractility to a substrate-independent cortical contractility, which is not linked to their tumor phenotype