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Author(s): V. Backman [1]; M. B. Wallace [2]; L. T. Perelman [1]; J. T. Arendt [1]; R. Gurjar [1]; M. G. Müller [1]; Q. Zhang [1]; G. Zonios [1]; E. [...]

An emerging research area in biophotonics with potentially near-term clinical applications in early stage cancer detection involves the investigation of possible correlations of the elastic light scattering properties of tissues with alterations in their cellular composition and nanostructure. Until recently, exploring these correlations has been impeded by a lack of robust and accurate mathematical models of the light scattering properties of complex structures. In this paper, we review recent progress in this area. Topics include: 1) development of accurate reduced-order expressions for the total scattering cross section spectra of a wide range of nonspherical and inhomogeneous particles; 2) rigorous finite-difference time-domain modeling results showing how the backscattering of light can be sensitive to nanometer scale features embedded within micrometer-scale particles; and 3) development of accurate reduced-order expressions for the backscattering depolarization properties of a wide range of inhomogeneous particles. These advances provide an improved science base for cellular level biophotonics, and have promise to accelerate the development of novel corresponding clinical technologies. [ABSTRACT FROM PUBLISHER]

We present a novel instrument to measure the spectral, angular, azimuthal, and polarization dependence of light backscattered by living biological tissues, thus providing the most comprehensive description of the light scattering to obtain unique quantitative information about the microarchitecture of living cells and tissues. We show the potential of this technique to characterize and diagnose early premalignant changes in the epithelia. In studies with a rodent model of colon carcinogenesis, we show that several parameters obtained using this technique, such as the number density of red blood cells in the capillary network immediately underlying the epithelium, the fractal dimension of the tissue, and the average roundness of subcellular structures, are significant for detection of precancerous changes at a very early stage of the carcinogenic process, at which no other histological or molecular markers have been identified. [ABSTRACT FROM PUBLISHER]

We present a novel instrument for imaging the angular distributions of light backscattered by biological cells and tissues. The intensities in different regions of the image are due to scatterers of different sizes. We exploit this to study scattering from particles smaller than the wavelength of light used, even when they are mixed with larger particles. We show that the scattering from subcellular structure in both normal and cancerous human cells is best fitted to inverse power-law distributions for the sizes of the scattering objects, and propose that the distribution of scattering objects may be different in normal versus cancerous cells [ABSTRACT FROM PUBLISHER]

We report an in situ method of probing the structure of living epithelial cells, based on light scattering spectroscopy with polarized light. The method makes it possible to distinguish between single backscattering from uppermost epithelial cells and multiply scattered light. The spectrum of the single backscattering component can be further analyzed to provide histological information about the epithelial cells such as the size distribution of the cell nuclei and their refractive index. These are valuable quantities' to detect and diagnose precancerous changes in human tissues [ABSTRACT FROM PUBLISHER]

This letter presents a scattered-field formulation for modeling dispersive media using the finite-difference time-domain (FDTD) method. Specifically, the auxiliary differential equation method is applied to Drude and Lorentz media for a scattered field FDTD model. The present technique can also be applied in a straightforward manner to Debye media. Excellent agreement is achieved between the FDTD-calculated and exact theoretical results for the reflection coefficient in half-space problems. [ABSTRACT FROM PUBLISHER]

The 30 articles in this special section cover a broad range among the following areas: advanced bioimaging; multifunctional endoscopy; novel approaches in biophotonic diagnostics and therapeutics; advanced biophotonic sensing techniques; laser-cell interactions; photoacoustics and terahertz biophotonics; nanobiophotonics; and novel laser, fiber-optics and electrooptics biophotonic tools, and devices. [ABSTRACT FROM PUBLISHER]

The eighteen papers in this special issue are devoted to the topic of biophotonics - new techniques that have been developed as alternatives to conventional medical methods for diagnostics, sensing, monitoring, and treatment of diseases. [ABSTRACT FROM PUBLISHER]

Most of what we know about gene transcription comes from the view of cells as molecular machines: focusing on the role of molecular modifications to the proteins carrying out transcriptional reactions at a loci-by-loci basis. This view ignores a critical reality: biological reactions do not happen in an empty space, but in a highly complex, interrelated, and dense nanoenvironment that profoundly influences chemical interactions. We explored the relationship between the physical nanoenvironment of chromatin and gene transcription in vitro. We analytically show that changes in the fractal dimension, D, of chromatin correspond to simultaneous increases in chromatin accessibility and compaction heterogeneity. Using these predictions, we demonstrate experimentally that nanoscopic changes to chromatin D within thirty minutes correlate with concomitant enhancement and suppression of transcription. Further, we show that the increased heterogeneity of physical structure of chromatin due to increase in fractal dimension correlates with increased heterogeneity of gene networks. These findings indicate that the higher order folding of chromatin topology may act as a molecular-pathway independent code regulating global patterns of gene expression. Since physical organization of chromatin is frequently altered in oncogenesis, this work provides evidence pairing molecular function to physical structure for processes frequently altered during tumorigenesis. [ABSTRACT FROM AUTHOR]

Extended abstract of a paper presented at Microscopy and Microanalysis 2012 in Phoenix, Arizona, USA, July 29 – August 2, 2012. [ABSTRACT FROM PUBLISHER]

Presents a letter to the editor regarding the frequency and distribution of the HLA class II alleles and haplotypes in the Icelandic population and Icelandic Type I diabetic patients.

Despite major importance in physics, biology, and other sciences, the optical sensing of nanoscale structures in the far zone remains an open problem due to the fundamental diffraction limit of resolution. We establish that the expected value of spectral variance (~Σ) of a far-field, diffraction-limited microscope image can quantify the refractive-index fluctuations of a label-free, weakly scattering sample at subdiffraction length scales. We report the general expression of ~Σ for an arbitrary refractive-index distribution. For an exponential refractive-index spatial correlation, we obtain a closed-form solution of ~Σ that is in excellent agreement with three-dimensional finite-difference time-domain solutions of Maxwell's equations. Sensing complex inhomogeneous media at the nanoscale can benefit fields from material science to medical diagnostics. [ABSTRACT FROM AUTHOR]

Nuclear blebs are herniations of the nucleus that occur in diseased nuclei that cause nuclear rupture leading to cellular dysfunction. Chromatin and lamins are two of the major structural components of the nucleus that maintain its shape and function, but their relative roles in nuclear blebbing remain elusive. To determine the composition of nuclear blebs, we compared the immunofluorescence intensity of DNA and lamin B in the main nucleus body to the nuclear bleb across cell types and perturbations. DNA density in the nuclear bleb was consistently decreased to about half of the nuclear body while lamin B levels in the nuclear bleb varied widely. Partial Wave Spectroscopic (PWS) microscopy recapitulated significantly decreased likelihood of high-density domains in the nuclear bleb versus body, independent of lamin B. Time lapse imaging into immunofluorescence reveals that decreased DNA density marks all nuclear blebs while decreased lamin B1 levels only occur in blebs that have recently ruptured. Thus, decreased DNA density is a better marker of a nuclear bleb than lamin B level., (© 2024. Published by The Company of Biologists Ltd.)

We propose the Self Returning Excluded Volume (SR-EV) model for the structure of chromatin based on stochastic rules and physical interactions. The SR-EV rules of return generate conformationally defined domains observed by single-cell imaging techniques. From nucleosome to chromosome scales, the model captures the overall chromatin organization as a corrugated system, with dense and dilute regions alternating in a manner that resembles the mixing of two disordered bi-continuous phases. This particular organizational topology is a consequence of the multiplicity of interactions and processes occurring in the nuclei, and mimicked by the proposed return rules. Single configuration properties and ensemble averages show a robust agreement between theoretical and experimental results including chromatin volume concentration, contact probability, packing domain identification and size characterization, and packing scaling behavior. Model and experimental results suggest that there is an inherent chromatin organization regardless of the cell character and resistant to an external forcing such as RAD21 degradation., Competing Interests: MC, MK, LA, VA, WL, EP, RN, VB, IS No competing interests declared, (© 2024, Carignano, Kroeger, Almassalha et al.)

Super-resolution microscopy has revolutionized our ability to visualize structures below the diffraction limit of conventional optical microscopy and is particularly useful for investigating complex biological targets like chromatin. Chromatin exhibits a hierarchical organization with structural compartments and domains at different length scales, from nanometers to micrometers. Single molecule localization microscopy (SMLM) methods, such as STORM, are essential for studying chromatin at the supra-nucleosome level due to their ability to target epigenetic marks that determine chromatin organization. Multi-label imaging of chromatin is necessary to unpack its structural complexity. However, these efforts are challenged by the high-density nuclear environment, which can affect antibody binding affinities, diffusivity and non-specific interactions. Optimizing buffer conditions, fluorophore stability, and antibody specificity is crucial for achieving effective antibody conjugates. Here, we demonstrate a sequential immunolabeling protocol that reliably enables three-label studies within the dense nuclear environment. This protocol couples multiplexed localization datasets with a robust analysis algorithm, which utilizes localizations from one target as seed points for distance, density and multi-label joint affinity measurements to explore complex organization of all three targets. Applying this multi-plexed algorithm to analyze distance and joint density reveals that heterochromatin and euchromatin are not-distinct territories, but that localization of transcription and euchromatin couple with the periphery of heterochromatic clusters. This work is a crucial step in molecular imaging of the dense nuclear environment as multi-label capacity enables for investigation of complex multi-component systems like chromatin with enhanced accuracy.

The emergence of aerobic respiration created unprecedented bioenergetic advantages, while imposing the need to protect critical genetic information from reactive byproducts of oxidative metabolism (i.e., reactive oxygen species, ROS). The evolution of histone proteins fulfilled the need to shield DNA from these potentially damaging toxins, while providing the means to compact and structure massive eukaryotic genomes. To date, several metabolism-linked histone post-translational modifications (PTMs) have been shown to regulate chromatin structure and gene expression. However, whether and how PTMs enacted by metabolically produced ROS regulate adaptive chromatin remodeling remain relatively unexplored. Here, we review novel mechanistic insights into the interactions of ROS with histones and their consequences for the control of gene expression regulation, cellular plasticity, and behavior., Competing Interests: Declaration of interests N.K. is a consultant for Thermo Fisher Scientific on proteomics and biological mass spectrometry applications., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Chromatin-sensitive Partial Wave Spectroscopic (csPWS) microscopy offers a non-invasive glimpse into the mass density distribution of cellular structures at the nanoscale, leveraging the spectroscopic information. Such capability allows us to analyze the chromatin structure and organization and the global transcriptional state of the cell nuclei for the study of its role in carcinogenesis. Accurate segmentation of the nuclei in csPWS microscopy images is an essential step in isolating them for further analysis. However, manual segmentation is error-prone, biased, time-consuming, and laborious, resulting in disrupted nuclear boundaries with partial or over-segmentation. Here, we present an innovative deep-learning-driven approach to automate the accurate nuclei segmentation of label-free live cell csPWS microscopy imaging data. Our approach, csPWS-seg, harnesses the Convolutional Neural Networks-based U-Net model with an attention mechanism to automate the accurate cell nuclei segmentation of csPWS microscopy images. We leveraged the structural, physical, and biological differences between the cytoplasm, nucleus, and nuclear periphery to construct three distinct csPWS feature images for nucleus segmentation. Using these images of HCT116 cells, csPWS-seg achieved superior performance with a median Intersection over Union (IoU) of 0.80 and a Dice Similarity Coefficient (DSC) score of 0.88. The csPWS-seg overcame the segmentation performance over the baseline U-Net model and another attention-based model, SE-U-Net, marking a significant improvement in segmentation accuracy. Further, we analyzed the performance of our proposed model with four loss functions: binary cross-entropy loss, focal loss, dice loss, and Jaccard loss. The csPWS-seg with focal loss provided the best results compared to other loss functions. The automatic and accurate nuclei segmentation offered by the csPWS-seg not only automates, accelerates, and streamlines csPWS data analysis but also enhances the reliability of subsequent chromatin analysis research, paving the way for more accurate diagnostics, treatment, and understanding of cellular mechanisms for carcinogenesis., Competing Interests: Disclosures The authors declare no conflicts of interest.