Glycosaminoglycans modulate long-range mechanical communication between cells in collagen networks.

Cells can sense and respond to mechanical forces in fibrous extracellular matrices (ECMs) over distances much greater than their size. This phenomenon, termed long-range force transmission, is enabled by the realignment (buckling) of collagen fibers along directions where the forces are tensile (compressive). However, whether other key structural components of the ECM, in particular glycosaminoglycans (GAGs), can affect the efficiency of cellular force transmission remains unclear. Here we developed a theoretical model of force transmission in collagen networks with interpenetrating GAGs, capturing the competition between tension-driven collagen fiber alignment and the swelling pressure induced by GAGs. Using this model, we show that the swelling pressure provided by GAGs increases the stiffness of the collagen network by stretching the fibers in an isotropic manner. We found that the GAG-induced swelling pressure can help collagen fibers resist buckling as the cells exert contractile forces. This mechanism impedes the alignment of collagen fibers and decreases long-range cellular mechanical communication. We experimentally validated the theoretical predictions by comparing the intensity of collagen fiber alignment between cellular spheroids cultured on collagen gels versus collagen­GAG cogels. We found significantly lower intensities of aligned collagen in collagen­GAG cogels, consistent with the prediction that GAGs can prevent collagen fiber alignment. The role of GAGs in modulating force transmission uncovered in this work can be extended to understand pathological processes such as the formation of fibrotic scars and cancer metastasis, where cells communicate in the presence of abnormally high concentrations of GAGs.

Introduction to the special feature: amplify Black voices in optics and photonics.

This multi-journal special issue highlights the work of Black scientists and engineers in optics and photonics to accomplish the goal of engaging the entire optics and photonics community and bring awareness to the quality of their research and contributions to the field.

Automated brightfield layerwise evaluation in three-dimensional micropatterning via two-photon polymerization.

Two-photon polymerization (TPP) is an advanced 3D fabrication technique capable of creating features with submicron precision. A primary challenge in TPP lies in the facile and accurate characterization of fabrication quality, particularly for structures possessing complex internal features. In this study, we introduce an automated brightfield layerwise evaluation technique that enables a simple-to-implement approach for in situ monitoring and quality assessment of TPP-fabricated structures. Our approach relies on sequentially acquired brightfield images during the TPP writing process and using background subtraction and image processing to extract layered spatial features. We experimentally validate our method by printing a fibrous tissue scaffold and successfully achieve an overall system-adjusted fidelity of 87.5% in situ. Our method is readily adaptable in most TPP systems and can potentially facilitate high-quality TPP manufacturing of sophisticated microstructures.

Reversible inter-degree-of-freedom optical-coherence conversion via entropy swapping.

The entropy associated with an optical field quantifies the field fluctuations and thus its coherence. Any binary optical degree-of-freedom (DoF) - such as polarization or the field at a pair of points in space - can each carry up to one bit of entropy. We demonstrate here that entropy can be reversibly swapped between different DoFs, such that coherence is converted back and forth between them without loss of energy. Specifically, starting with a spatially coherent but unpolarized field carrying one bit of entropy, we unitarily convert the coherence from the spatial DoF to polarization to produce a spatially incoherent but polarized field by swapping the entropy between the two DoFs. Next, we implement the inverse unitary operator, thus converting the coherence back to yield once again a spatially coherent yet unpolarized field. We exploit the intermediate stage between the two coherence conversions - where the spatial coherence has been converted to the polarization DoF - to verify that the field has become immune to the deleterious impact of spatial phase scrambling. Maximizing the spatial entropy protects the spatial DoF by preventing it from taking on any additional fluctuations. After the second coherence conversion, spatial coherence is readily retrieved, and the effect of spatial phase scrambling circumvented.

Single-wavelength, single-shot pulse oximetry using an LED-generated vector beam.

Photoplethysmography (PPG) is an optical technique that monitors blood oxygen saturation levels, typically with the use of pulse oximeters. Conventional pulse oximetry estimates the ratio of light absorbed at two wavelengths. Attempts have been made to improve the precision of these measurements by using polarized light, with the tradeoff of requiring multiple sequential measurements. We demonstrate a novel PPG technique that uses radially polarized light generated by a light-emitting diode (LED) to obtain single-shot, blood oxygen-saturation measurements using a single wavelength at a rate of 50 fps. Our work, to the best of our knowledge, presents both a novel use of a vector beam and a first demonstration of vector-beam generation using LEDs.

Excitation of surface plasmon polaritons by diffraction-free and vector beams.

Surface plasmon polaritons (SPPs) are traditionally excited by plane waves within the Rayleigh range of a focused transverse-magnetic (TM) Gaussian beam. Here we investigate and confirm the coupling between SPPs and two-dimensional Gaussian and Bessel-Gauss wave packets, as well as one-dimensional light sheets and space-time wave packets. We encode the incoming wavefronts with spatially varying states of polarization; then we couple the respective TM components of radial and azimuthal vector beam profiles to confirm polarization-correlation and spatial-mode selectivity. Our results do not require material optimization or multi-dimensional confinement via periodically corrugated metal surfaces to achieve coupling at a greater extent, hereby outlining a pivotal, yet commonly overlooked, path towards the development of long-range biosensors and all-optical integrated plasmonic circuits.

Space-time vector light sheets.

We introduce the space-time (ST) vector light sheet. This unique one-dimensional ST wave packet is characterized by classical entanglement (CE), a correlation between at least two non-separable intrinsic degrees-of-freedom (DoFs), which in this case are the spatiotemporal DoFs in parallel with the spatial-polarization DoFs. We experimentally confirm that the ST vector light sheet maintains the intrinsic features of the uniformly polarized ST light sheet, such as near-diffraction-free propagation and self-healing, while also maintaining the intrinsic polarization structure of common vector beams, such as those that are radially polarized and azimuthally polarized. We also show that the vector beam structure of the ST vector light sheet is maintained in the subluminal and superluminal regimes.

Relaying of the local enhanced electric-field using stacked gold bowtie nanoantennas.

We demonstrate the possibility of designing plasmonic structures that extend, and even further concentrate the field enhancement in the feed-gap region of single-layer Au bowtie nanoantennas (BNAs) hundreds of nanometers away from the initial metal-dielectric interface. The design is based on a stack of Au BNAs with progressively reduced gap distances sandwiched by thin dielectric layers. We find that this stacked BNA geometry also behaves as a near-field focusing lens of nanometer focal length. By merely controlling the number of BNAs in a stack and thickness of the accompanying dielectric layers, we show that the usual fast-damping field enhancement right above a 50 nm thick base BNA can be relayed and maintained at >104 over a distance of at least up to 200 nm above the base BNA surface. Our novel plasmonic structures offer an approach to design plasmonic nanostructures for applications in metamaterial engineering, SERS, and nonlinear optics.

Nonlinear Focal Modulation Microscopy.

We demonstrate nonlinear focal modulation microscopy (NFOMM) to achieve superresolution imaging. Traditional approaches to superresolution that utilize point scanning often rely on spatially reducing the size of the emission pattern by directly narrowing (e.g., through minimizing the detection pinhole in Airyscan, Zeiss) or indirectly peeling its outer profiles [e.g., through depleting the outer emission region in stimulated emission depletion (STED) microscopy]. We show that an alternative conceptualization that focuses on maximizing the optical system's frequency shifting ability offers advantages in further improving resolution while reducing system complexity. In NFOMM, a spatial light modulator and a suitably intense laser illumination are used to implement nonlinear focal-field modulation to achieve a transverse spatial resolution of ∼60 nm (∼λ/10). We show that NFOMM is comparable with STED microscopy and suitable for fundamental biology studies, as evidenced in imaging nuclear pore complexes, tubulin and vimentin in Vero cells. Since NFOMM is readily implemented as an add-on module to a laser-scanning microscope, we anticipate wide utility of this new imaging technique.

Application of a reflective microscope objective for multiphoton microscopy.

Reflective objectives (ROs) mitigate chromatic aberration across a broad wavelength range. Yet, a systematic performance characterisation of ROs has not been done. In this paper, we compare the performance of a 0.5 numerical-aperture (NA) reflective objective (RO) with a 0.55 NA standard glass objective (SO), using two-photon fluorescence (TPF) and second-harmonic generation (SHG). For experiments spanning ∼1 octave in the visible and NIR wavelengths, the SO leads to defocusing errors of 25-40% for TPF images of subdiffraction fluorescent beads and 10-12% for SHG images of collagen fibres. The corresponding error for the RO is ∼4% for both imaging modalities. This work emphasises the potential utility of ROs for multimodal multiphoton microscopy applications.

Application of an advanced maximum likelihood estimation restoration method for enhanced-resolution and contrast in second-harmonic generation microscopy.

Second-harmonic generation (SHG) microscopy has gained popularity because of its ability to perform submicron, label-free imaging of noncentrosymmetric biological structures, such as fibrillar collagen in the extracellular matrix environment of various organs with high contrast and specificity. Because SHG is a two-photon coherent scattering process, it is difficult to define a point spread function (PSF) for this modality. Hence, compared to incoherent two-photon processes like two-photon fluorescence, it is challenging to apply the various PSF-engineering methods to improve the spatial resolution to be close to the diffraction limit. Using a synthetic PSF and application of an advanced maximum likelihood estimation (AdvMLE) deconvolution algorithm, we demonstrate restoration of the spatial resolution in SHG images to that closer to the theoretical diffraction limit. The AdvMLE algorithm adaptively and iteratively develops a PSF for the supplied image and succeeds in improving the signal to noise ratio (SNR) for images where the SHG signals are derived from various sources such as collagen in tendon and myosin in heart sarcomere. Approximately 3.5 times improvement in SNR is observed for tissue images at depths of up to ∼480 nm, which helps in revealing the underlying helical structures in collagen fibres with an ∼26% improvement in the amplitude contrast in a fibre pitch. Our approach could be adapted to noisy and low resolution modalities such as micro-nano CT and MRI, impacting precision of diagnosis and treatment of human diseases.

Third-harmonic generation imaging of breast tissue biopsies.

We demonstrate for the first time the imaging of unstained breast tissue biopsies using third-harmonic generation (THG) microscopy. As a label-free imaging technique, THG microscopy is compared to phase contrast and polarized light microscopy which are standard imaging methods for breast tissues. A simple feature detection algorithm is applied to detect tumour-associated lymphocyte rich regions in unstained breast biopsy tissue and compared with corresponding regions identified by a pathologist from bright-field images of hematoxylin and eosin stained breast tissue. Our results suggest that THG imaging holds potential as a complementary technique for analysing breast tissue biopsies.

Resolution-enhanced surface plasmon-coupled emission microscopy.

A novel fluorescence emission difference technique is proposed for further enhancements of the lateral resolution in surface plasmon-coupled emission microscopy (SPCEM). In the proposed method, the difference between the image with phase modulation by using a 0-2π vortex phase plate (VPP) along with a diaphragm and the original image obtained from SPCEM is used to estimate the spatial distribution of the analyzed sample. By optimizing the size of the diaphragm and the subtractive factor, the lateral resolution can be enhanced by about 20% and 33%, compared with that in SPCEM with a single 0-2π VPP and conventional wide-field fluorescence microscopy, respectively. Related simulation results are presented to verify the capability of the proposed method for improving lateral resolution and reducing imaging distortion. It is believed that the proposed method has potentials to improve the performance of SPCEM, thus facilitating biological observation and research.

Iterative phase-retrieval method for generating stereo array of polarization-controlled focal spots.

This Letter introduces an iterative phase-retrieval method based on the Gerchberg-Saxton (G-S) algorithm for generating any arbitrary 3D pattern in image space, while simultaneously controlling the polarization orientation at each pixel. For proof-of-principle, we generate a stereo focal spot array with distinct polarization orientation for each spot. This method is universal for controlling the output polarization; the only requirement is that the input polarization should be spatially inhomogeneous. This work has the potential to impact coherent imaging techniques and spectroscopy.

Multifunctional plasmonic film for recording near-field optical intensity.

We demonstrate the plasmonic equivalent of photographic film for recording optical intensity in the near field. The plasmonic structure is based on gold bowtie nanoantenna arrays fabricated on SiO2 pillars. We show that it can be employed for direct laser writing of image data or recording the polarization structure of optical vector beams. Scanning electron micrographs reveal a careful sculpting of the radius of curvature and height of the triangles composing the illuminated nanoantennas, as a result of plasmonic heating, that permits spatial tunability of the resonance response of the nanoantennas without sacrificing their geometric integrity. In contrast to other memory-dedicated approaches using Au nanorods embedded in a matrix medium, plasmonic film can be used in multiple application domains. To demonstrate this functionality, we utilize the structures as plasmonic optical tweezers and show sequestering of SiO2 microparticles into optically written channels formed between exposed sections of the film. The plasmonic film offers interesting possibilities for photonic applications including optofluidic channels "without walls," in situ tailorable biochemical sensing assays, and near-field particle manipulation and sorting.

Harnessing randomness to control the polarization of light transmitted through highly scattering media.

We show that the multiple scattering events taking place inside a highly scattering medium, in conjunction with wavefront shaping, can be used to control the state of polarization of the light transmitted through a highly scattering medium. This control is achieved by using the intensity, phase, and polarization changing behavior of a scattering medium captured by a vector transmission matrix (VTM). We use a single beam incident upon a scattering medium to measure the absolute value of the VTM elements, in contrast to the multiple beams required in our previously reported approach. Further, the phase-only spatial light modulator based on a low-cost (< US$600) deformable micro-mirror array used in our work will make similar experiments accessible to other researchers.

Demonstration of flat-top focusing under radial polarization illumination.

We experimentally demonstrate the generation of a flat-top intensity distribution using a radially polarized vector beam. Our approach uses higher numerical aperture focusing than what has been previously reported for a single, fixed, vector beam. In addition, the flat-top focus generated in our scheme exhibits a polarization gradient along the radial coordinate in the focal volume, with an on-axis longitudinal field component that persists over 2λ, which is a stark difference from conventional flat-top fields, which exhibit intensity profiles that are uniformly polarized. Our experimental results are found to be in good agreement with the theoretical prediction.

Mueller matrix analysis of a biologically sourced engineered tissue construct as polarimetric phantom.

Significance: The polarimetric properties of biological tissues are often difficult to ascertain independent of their complex structural and organizational features. Conventional polarimetric tissue phantoms have well-characterized optical properties but are overly simplified. We demonstrate that an innovative, biologically sourced, engineered tissue construct better recapitulates the desired structural and polarimetric properties of native collagenous tissues, with the added benefit of potential tunability of the polarimetric response. We bridge the gap between non-biological polarimetric phantoms and native tissues. Aim: We aim to evaluate a synthesized tissue construct for its effectiveness as a phantom that mimics the polarimetric properties in typical collagenous tissues. Approach: We use a fibroblast-derived, ring-shaped engineered tissue construct as an innovative tissue phantom for polarimetric imaging. We perform polarimetry measurements and subsequent analysis using the Mueller matrix decomposition and Mueller matrix transformation methods. Scalar polarimetric parameters of the engineered tissue are analyzed at different time points for both a control group and for those treated with the transforming growth factor ( TGF ) - ß 1 . Second-harmonic generation (SHG) imaging and three-dimensional collagen fiber organization analysis are also applied. Results: We identify linear retardance and circular depolarization as the parameters that are most sensitive to the tissue culture time and the addition of TGF - ß 1 . Aside from a statistically significant increase over time, the behavior of linear retardance and circular depolarization indicates that the addition of TGF - ß 1 accelerates the growth of the engineered tissue, which is consistent with expectations. We also find through SHG images that collagen fiber organization becomes more aligned over time but is not susceptible to the addition of TGF - ß 1 . Conclusions: The engineered tissue construct exhibits changes in polarimetric properties, especially linear retardance and circular depolarization, over culture time and under TGF - ß 1 treatments. This tissue construct has the potential to act as a controlled modular optical phantom for polarimetric-based methods.

Fibroblast senescence-associated extracellular matrix promotes heterogeneous lung niche.

Senescent cell accumulation in the pulmonary niche is associated with heightened susceptibility to age-related disease, tissue alterations, and ultimately a decline in lung function. Our current knowledge of senescent cell-extracellular matrix (ECM) dynamics is limited, and our understanding of how senescent cells influence spatial ECM architecture changes over time is incomplete. Herein is the design of an in vitro model of senescence-associated extracellular matrix (SA-ECM) remodeling using a senescent lung fibroblast-derived matrix that captures the spatiotemporal dynamics of an evolving senescent ECM architecture. Multiphoton second-harmonic generation microscopy was utilized to examine the spatial and temporal dynamics of fibroblast SA-ECM remodeling, which revealed a biphasic process that established a disordered and heterogeneous architecture. Additionally, we observed that inhibition of transforming growth factor-ß signaling during SA-ECM remodeling led to improved local collagen fiber organization. Finally, we examined patient samples diagnosed with pulmonary fibrosis to further tie our results of the in vitro model to clinical outcomes. Moreover, we observed that the senescence marker p16 is correlated with local collagen fiber disorder. By elucidating the temporal dynamics of SA-ECM remodeling, we provide further insight on the role of senescent cells and their contributions to pathological ECM remodeling.

Quantitative control over the intensity and phase of light transmitted through highly scattering media.

We experimentally demonstrate the use of the transmission matrix (TM) to quantitatively control the amplitude and phase of the light transmitted through highly scattering media. This is achieved by measuring the absolute value of the TM elements. We also use the fact that the cross-correlations between the contributions of different input channels at the observation plane is important in describing the transmitted optical field. In addition, we demonstrate both quantitative control of the intensity at multiple output spatial modes, each with a different intensity, as well as a "dark" area of low intensity. Our experiments are carried out using a low cost (less than US$600) spatial binary amplitude modulator that we modify for phase-only operation, as well as a novel optical setup that enables independent control of a reference and control signal while maintaining interferometric stability. The optical implementation used in this paper will make such experiments widely accessible to many researchers. Furthermore, the results presented could serve as the foundation for many useful potential applications ranging from the biomedical sciences to optical communications.