Coherent light scattering from cellular dynamics in living tissues.

This review examines the biological physics of intracellular transport probed by the coherent optics of dynamic light scattering from optically thick living tissues. Cells and their constituents are in constant motion, composed of a broad range of speeds spanning many orders of magnitude that reflect the wide array of functions and mechanisms that maintain cellular health. From the organelle scale of tens of nanometers and upward in size, the motion inside living tissue is actively driven rather than thermal, propelled by the hydrolysis of bioenergetic molecules and the forces of molecular motors. Active transport can mimic the random walks of thermal Brownian motion, but mean-squared displacements are far from thermal equilibrium and can display anomalous diffusion through Lévy or fractional Brownian walks. Despite the average isotropic three-dimensional environment of cells and tissues, active cellular or intracellular transport of single light-scattering objects is often pseudo-one-dimensional, for instance as organelle displacement persists along cytoskeletal tracks or as membranes displace along the normal to cell surfaces, albeit isotropically oriented in three dimensions. Coherent light scattering is a natural tool to characterize such tissue dynamics because persistent directed transport induces Doppler shifts in the scattered light. The many frequency-shifted partial waves from the complex and dynamic media interfere to produce dynamic speckle that reveals tissue-scale processes through speckle contrast imaging and fluctuation spectroscopy. Low-coherence interferometry, dynamic optical coherence tomography, diffusing-wave spectroscopy, diffuse-correlation spectroscopy, differential dynamic microscopy and digital holography offer coherent detection methods that shed light on intracellular processes. In health-care applications, altered states of cellular health and disease display altered cellular motions that imprint on the statistical fluctuations of the scattered light. For instance, the efficacy of medical therapeutics can be monitored by measuring the changes they induce in the Doppler spectra of livingex vivocancer biopsies.

Immune checkpoint inhibitor-related colitis in patients on immunotherapy for cancer.

OBJECTIVES: Immune checkpoint inhibitors, a revolutionary class of cancer immunotherapy drugs, have transformed cancer treatment by bolstering antitumor immunity for various advanced-stage solid cancers. The US Food and Drug Administration has approved 7 immune checkpoint inhibitors that target 3 major immune checkpoint proteins: cytotoxic T-lymphocyte-associated protein 4, programmed cell death 1 protein, and programmed cell death 1 ligand 1. In addition to their remarkable efficacy, however, these inhibitors have been observed causing immune-related adverse events, particularly immune checkpoint inhibitor-related colitis, which often results in severe or life-threatening clinical issues. METHODS: The diagnosis of immune checkpoint inhibitor-related colitis relies on incorporation of clinical evaluation as well as endoscopic and histopathologic examination, with exclusion of other potential etiologies. RESULTS: The common histopathologic manifestations of immune checkpoint inhibitor-related colitis are acute active colitis, chronic active colitis, microscopic colitis (collagenous or lymphocytic), and ischemic colitis, with patterns overlapping. Notably, enterocyte apoptosis is a unique feature of immune checkpoint inhibitor toxicity. The proposed mechanisms for the pathogenesis of immune checkpoint inhibitor-related colitis are primarily associated with autoimmune-type dysregulation and gut microbiome alteration. This review summarizes the clinical and pathologic characteristics of immune checkpoint inhibitor-related colitis and elucidates its underlying pathogenic mechanisms. CONCLUSIONS: Future successful management of this form of colitis relies on our comprehension of the intricate interplay between tumoral and systemic immune responses to immune checkpoint inhibitors and innovative approaches to modify these responses, along with specific immune cell populations, to preclude immune-related adverse events while achieving antitumor therapeutic outcomes.

Comparative oncology chemosensitivity assay for personalized medicine using low-coherence digital holography of dynamic light scattering from cancer biopsies.

Nearly half of cancer patients who receive standard-of-care treatments fail to respond to their first-line chemotherapy, demonstrating the pressing need for improved methods to select personalized cancer therapies. Low-coherence digital holography has the potential to fill this need by performing dynamic contrast OCT on living cancer biopsies treated ex vivo with anti-cancer therapeutics. Fluctuation spectroscopy of dynamic light scattering under conditions of holographic phase stability captures ultra-low Doppler frequency shifts down to 10 mHz caused by light scattering from intracellular motions. In the comparative preclinical/clinical trials presented here, a two-species (human and canine) and two-cancer (esophageal carcinoma and B-cell lymphoma) analysis of spectral phenotypes identifies a set of drug response characteristics that span species and cancer type. Spatial heterogeneity across a centimeter-scale patient biopsy sample is assessed by measuring multiple millimeter-scale sub-samples. Improved predictive performance is achieved for chemoresistance profiling by identifying red-shifted sub-samples that may indicate impaired metabolism and removing them from the prediction analysis. These results show potential for using biodynamic imaging for personalized selection of cancer therapy.

Biodynamic prediction of neoadjuvant chemotherapy response: Results from a prospective multicenter study of predictive accuracy among muscle-invasive bladder cancer patients.

BACKGROUND: Biodynamic signatures (temporal patterns of microscopic motion within a 3-dimensional tumor explant) offer phenomic biomarkers that are highly predictive for therapeutic response. OBJECTIVE: By utilizing motility contrast tomography, which provides a simple, fast assessment of motion patterns in living tissue, we evaluated the predictive accuracy of a biodynamic drug response classifier in muscle-invasive bladder cancer (MIBC) patients undergoing neoadjuvant chemotherapy (NAC). DESIGN, SETTING, AND PARTICIPANTS: One hundred five consecutive bladder cancer patients suspected of having MIBC were screened in a multi-institutional prospective observational study (NCT03739177) from July 2018 to June 2020, of whom, 30 completed NAC and radical cystectomy. INTERVENTION(S): Biodynamic signatures from treatment-naïve fresh bladder tumor specimens obtained after transurethral resection were measured in living tumor fragments challenged by standard-of-care cytotoxins. Patients received gemcitabine and cisplatin or dose-dense methotrexate, vinblastine, doxorubicin, and cisplatin per institutional guidelines and were followed through radical cystectomy. OUTCOMES MEASUREMENTS AND STATISTICAL ANALYSIS: A 4-level classifier was developed to predict pathologic complete response (pCR) vs. incomplete response utilizing a one-left-out cross-validation protocol to minimize over-fitting. Area under the curve evaluated predictive utility. RESULTS: Thirty percent (9 of 30) achieved pCR. Utilizing the 4-level classifier, biodynamically "favored" (scoring ≥ 3) and "strongly favored" (scoring 4) regimens accurately predicted pCR at rates of 66.7% (4 of 6 patients) and 100% (4 of 4 patients), respectively. Biodynamically "favored" scores predicted pCR with 88% sensitivity and 95% negative predictive value, P < 0.0001. Only 5.0% (1 of 20 patients) achieved pCR from regimens scoring 1 or 2, indicating poor to no response from NAC. Area under the receiver operating curve was 96% (95% Confidence Interval: 79%-99%, P < 0.0001). Future direction involves validating this model prospectively. PRINCIPAL CONCLUSIONS: Biodynamic scoring accurately predicts response in MIBC patients receiving NAC and holds promise to substantially improve the scope of appropriate management intervention.

Inverse problems in blood flow modeling: A review.

Mathematical and computational modeling of the cardiovascular system is increasingly providing non-invasive alternatives to traditional invasive clinical procedures. Moreover, it has the potential for generating additional diagnostic markers. In blood flow computations, the personalization of spatially distributed (i.e., 3D) models is a key step which relies on the formulation and numerical solution of inverse problems using clinical data, typically medical images for measuring both anatomy and function of the vasculature. In the last years, the development and application of inverse methods has rapidly expanded most likely due to the increased availability of data in clinical centers and the growing interest of modelers and clinicians in collaborating. Therefore, this work aims to provide a wide and comparative overview of literature within the last decade. We review the current state of the art of inverse problems in blood flows, focusing on studies considering fully dimensional fluid and fluid-solid models. The relevant physical models and hemodynamic measurement techniques are introduced, followed by a survey of mathematical data assimilation approaches used to solve different kinds of inverse problems, namely state and parameter estimation. An exhaustive discussion of the literature of the last decade is presented, structured by types of problems, models and available data.

Validation of 4D Flow based relative pressure maps in aortic flows.

While the clinical gold standard for pressure difference measurements is invasive catheterization, 4D Flow MRI is a promising tool for enabling a non-invasive quantification, by linking highly spatially resolved velocity measurements with pressure differences via the incompressible Navier-Stokes equations. In this work we provide a validation and comparison with phantom and clinical patient data of pressure difference maps estimators. We compare the classical Pressure Poisson Estimator (PPE) and the new Stokes Estimator (STE) against catheter pressure measurements under a variety of stenosis severities and flow intensities. Specifically, we use several 4D Flow data sets of realistic aortic phantoms with different anatomic and hemodynamic severities and two patients with aortic coarctation. The phantom data sets are enriched by subsampling to lower resolutions, modification of the segmentation and addition of synthetic noise, in order to study the sensitivity of the pressure difference estimators to these factors. Overall, the STE method yields more accurate results than the PPE method compared to catheterization data. The superiority of the STE becomes more evident at increasing Reynolds numbers with a better capacity of capturing pressure gradients in strongly convective flow regimes. The results indicate an improved robustness of the STE method with respect to variation in lumen segmentation. However, with heuristic removal of the wall-voxels, the PPE can reach a comparable accuracy for lower Reynolds' numbers.

Common-path interferometer for digital holographic Doppler spectroscopy of living biological tissues.

SIGNIFICANCE: Common-path interferometers have the advantage of producing ultrastable interferometric fringes compared with conventional interferometers, such as Michelson or Mach-Zehnder that are sensitive to environmental instabilities. Isolating interferometric measurements from mechanical disturbances is important in biodynamic imaging because Doppler spectroscopy of intracellular dynamics requires extreme stability for phase-sensitive interferometric detection to capture fluctuation frequencies down to 10 mHz. AIM: The aim of this study was to demonstrate that Doppler spectra produced from a common-path interferometer using a grating and a spatial filter (SF) are comparable to, and more stable than, spectra from conventional biodynamic imaging. APPROACH: A common-path interferometer using a holographic diffraction grating and an SF was employed with a low-coherence source. Simulations evaluated the spatial resolution. DLD-1 (human colon adenocarcinoma) spheroids were used as living target tissue samples. Power spectra under external vibrations and drug-response spectrograms were compared between common-path and Fourier-domain holographic systems. RESULTS: The common-path holography configuration shows enhanced interferometric stability against mechanical vibrations through common-mode rejection while maintaining sensitivity to Doppler frequency fluctuations caused by intracellular motions. CONCLUSIONS: A common-path interferometer using a grating and an SF can provide enhanced interferometric stability in tissue-dynamics spectroscopy for drug screening assays.

Biodynamic digital holographic speckle microscopy for oocyte and embryo metabolic evaluation.

Assisted reproductive technologies seek to improve the success rate of pregnancies. Morphology scoring is a common approach to evaluate oocyte and embryo viability prior to embryo transfer in utero, but the efficacy of the method is low. We apply biodynamic imaging, based on dynamic light scattering and low-coherence digital holography, to assess the metabolic activity of oocytes and embryos. A biodynamic microscope, developed to image small and translucent biological specimens, is inserted into the bay of a commercial inverted microscope that can switch between conventional microscopy channels and biodynamic microscopy. We find intracellular Doppler spectral features that act as noninvasive proxies for embryo metabolic activity that may relate to embryo viability.

Doppler imaging detects bacterial infection of living tissue.

Living 3D in vitro tissue cultures, grown from immortalized cell lines, act as living sentinels as pathogenic bacteria invade the tissue. The infection is reported through changes in the intracellular dynamics of the sentinel cells caused by the disruption of normal cellular function by the infecting bacteria. Here, the Doppler imaging of infected sentinels shows the dynamic characteristics of infections. Invasive Salmonella enterica serovar Enteritidis and Listeria monocytogenes penetrate through multicellular tumor spheroids, while non-invasive strains of Escherichia coli and Listeria innocua remain isolated outside the cells, generating different Doppler signatures. Phase distributions caused by intracellular transport display Lévy statistics, introducing a Lévy-alpha spectroscopy of bacterial invasion. Antibiotic treatment of infected spheroids, monitored through time-dependent Doppler shifts, can distinguish drug-resistant relative to non-resistant strains. This use of intracellular Doppler spectroscopy of living tissue sentinels opens a new class of microbial assay with potential importance for studying the emergence of antibiotic resistance.

Cancer Holography for Personalized Medicine.

Digital holography can measure the 3D physiology and motion of cancer cells, allowing identification of effective chemotherapies for patients.

Biodynamic signatures from ex vivo bone marrow aspirates are associated with chemotherapy-induced neutropenia in cancer-bearing dogs.

BACKGROUND: Neutropenia is the most common dose-limiting side effect of cytotoxic chemotherapy in cancer-bearing dogs. Biodynamic imaging (BDI) is a functional imaging technology that measures dynamic light scattering from living, three-dimensional tissues to characterize intracellular motion within those tissues. Previous studies have associated BDI biomarkers with tumour sensitivity to chemotherapy agents in dogs with naturally occurring cancer. We hypothesized that BDI, performed ex vivo on bone marrow aspirate samples, would identify dynamic biomarkers associated with the occurrence of specific degrees of neutropenia in tumour-bearing dogs receiving doxorubicin chemotherapy. MATERIALS AND METHODS: Bone marrow aspirates were collected from 10 dogs with naturally occurring cancers prior to initiation of doxorubicin treatment. BDI was performed on bone marrow samples treated ex vivo with doxorubicin at 0.1, 1, 10 and 100 µM along with 0.1% DMSO as a control. Dogs then were treated with doxorubicin (30 mg/m2 , intravenously). Peripheral blood neutrophil counts were obtained on the day of treatment and again 7 days later. Receiver operating characteristic curves identified provisional breakpoints for BDI biomarkers that correlated with specific changes in neutrophil counts between the two time points. RESULTS: Provisional breakpoints for several BDI biomarkers were identified, specifying dogs with the largest proportionate change in neutrophils and with neutropenia that was grade 2 or higher following doxorubicin treatment. CONCLUSIONS: Biodynamic imaging of bone marrow aspirates may identify those dogs at greater risk for neutropenia following doxorubicin chemotherapy. This approach may be useful for pre-emptively modifying chemotherapy dosing in dogs to avoid unacceptable side effects.

Intracellular optical doppler phenotypes of chemosensitivity in human epithelial ovarian cancer.

Development of an assay to predict response to chemotherapy has remained an elusive goal in cancer research. We report a phenotypic chemosensitivity assay for epithelial ovarian cancer based on Doppler spectroscopy of infrared light scattered from intracellular motions in living three-dimensional tumor biopsy tissue measured in vitro. The study analyzed biospecimens from 20 human patients with epithelial ovarian cancer. Matched primary and metastatic tumor tissues were collected for 3 patients, and an additional 3 patients provided only metastatic tissues. Doppler fluctuation spectra were obtained using full-field optical coherence tomography through off-axis digital holography. Frequencies in the range from 10 mHz to 10 Hz are sensitive to changes in intracellular dynamics caused by platinum-based chemotherapy. Metastatic tumor tissues were found to display a biodynamic phenotype that was similar to primary tissue from patients who had poor clinical outcomes. The biodynamic phenotypic profile correctly classified 90% [88-91% c.i.] of the patients when the metastatic samples were characterized as having a chemoresistant phenotype. This work suggests that Doppler profiling of tissue response to chemotherapy has the potential to predict patient clinical outcomes based on primary, but not metastatic, tumor tissue.

Probing complex geophysical geometries with chattering dust.

The modern energy economy and environmental infrastructure rely on the flow of fluids through fractures in rock. Yet this flow cannot be imaged directly because rocks are opaque to most probes. Here we apply chattering dust, or chemically reactive grains of sucrose containing pockets of pressurized carbon dioxide, to study rock fractures. As a dust grain dissolves, the pockets burst and emit acoustic signals that are detected by distributed sets of external ultrasonic sensors that track the dust movement through fracture systems. The dust particles travel through locally varying fracture apertures with varying speeds and provide information about internal fracture geometry, flow paths and bottlenecks. Chattering dust particles have an advantage over chemical sensors because they do not need to be collected, and over passive tracers because the chattering dust delineates the transport path. The current laboratory work has potential to scale up to near-borehole applications in the field.

Tissue dynamics spectroscopic imaging: functional imaging of heterogeneous cancer tissue.

SIGNIFICANCE: Tumor heterogeneity poses a challenge for the chemotherapeutic treatment of cancer. Tissue dynamics spectroscopy captures dynamic contrast and can capture the response of living tissue to applied therapeutics, but the current analysis averages over the complicated spatial response of living biopsy samples. AIM: To develop tissue dynamics spectroscopic imaging (TDSI) to map the heterogeneous spatial response of tumor tissue to anticancer drugs. APPROACH: TDSI is applied to tumor spheroids grown from cell lines and to ex vivo living esophageal biopsy samples. Doppler fluctuation spectroscopy is performed on a voxel basis to extract spatial maps of biodynamic biomarkers. Functional images and bivariate spatial maps are produced using a bivariate color merge to represent the spatial distribution of pairs of signed drug-response biodynamic biomarkers. RESULTS: We have mapped the spatial variability of drug responses within biopsies and have tracked sample-to-sample variability. Sample heterogeneity observed in the biodynamic maps is associated with histological heterogeneity observed using inverted selective-plane illumination microscopy. CONCLUSION: We have demonstrated the utility of TDSI as a functional imaging method to measure tumor heterogeneity and its potential for use in drug-response profiling.

Hypopharyngeal Dedifferentiated Liposarcoma in the MDM2 Era: A Case Report and Short Review.

Pharyngeal liposarcomas are very rare; still more rare are dedifferentiated liposarcomas in the pharynx. An 82-year-old man presented with dysphagia, voice changes, weight loss, nasal regurgitation of liquids, and coughing spells. A 3.5 cm mass was identified in the hypopharynx. The mass was biopsied and diagnosed as a benign fibroepithelial polyp. Continued symptoms and airway obstruction prompted a pharyngectomy, and the mass was then diagnosed as dedifferentiated liposarcoma. Due to infrequency and subtle histological findings, liposarcomas of the head and neck can be misdiagnosed and recur.

Simulation of hypoxia PET-tracer uptake in tumours: Dependence of clinical uptake-values on transport parameters and arterial input function.

Poor radiotherapy outcome is in many cases related to hypoxia, due to the increased radioresistance of hypoxic tumour cells. Positron emission tomography may be used to non-invasively assess the oxygenation status of the tumour using hypoxia-specific radiotracers. Quantification and interpretation of these images remains challenging, since radiotracer binding and oxygen tension are not uniquely related. Computer simulation is a useful tool to improve the understanding of tracer dynamics and its relation to clinical uptake parameters currently used to quantify hypoxia. In this study, a model for simulating oxygen and radiotracer distribution in tumours was implemented to analyse the impact of physiological transport parameters and of the arterial input function (AIF) on: oxygenation histograms, time-activity curves, tracer binding and clinical uptake-values (tissue-to-blood ratio, TBR, and a composed hypoxia-perfusion metric, FHP). Results were obtained for parallel and orthogonal vessel architectures and for vascular fractions (VFs) of 1% and 3%. The most sensitive parameters were the AIF and the maximum binding rate (Kmax). TBR allowed discriminating VF for different AIF, and FHP for different Kmax, but neither TBR nor FHP were unbiased in all cases. Biases may especially occur in the comparison of TBR- or FHP-values between different tumours, where the relation between measured and actual AIF may vary. Thus, these parameters represent only surrogates rather than absolute measurements of hypoxia in tumours.

Biodynamic optical assay for embryo viability.

Early stage porcine parthenogenetic embryos were evaluated for metabolic activity using a biodynamic microscope (BDM) that images dynamic light scattering using low-coherence digital holography. The microscope has a 45-deg illumination configuration that reduces specular background for the imaging of small translucent samples. The off-axis illumination is compatible with coherence-gated imaging because of volumetric light scattering in which the coherence plane is tilted at half the illumination angle in a three-dimensional tissue target. The BDM was used to profile the viability of porcine parthenotes with normal and with inhibited mitochondrial adenosine triphosphate (ATP) production using Doppler fluctuation spectroscopy. The ATP concentrations in the parthenotes, which are indicative of developmental potential, were validated by a conventional bioluminescence assay. Biodynamic classifications achieved ∼80 % accuracy correlating sample ATP treatment, providing a quick, label-free surrogate measurement to replace invasive metabolic assays as a candidate for evaluating quality of early embryos in the assisted reproductive technology setting.

Doppler fluctuation spectroscopy of intracellular dynamics in living tissue.

Intracellular dynamics in living tissue are dominated by active transport driven by bioenergetic processes far from thermal equilibrium. Intracellular constituents typically execute persistent walks. In the limit of long mean free paths, the persistent walks are ballistic, exhibiting a "Doppler edge" in light scattering fluctuation spectra. At shorter transport lengths, the fluctuations are described by lifetime-broadened Doppler spectra. Dynamic light scattering from transport in the ballistic, diffusive, or the crossover regimes is derived analytically, including the derivation of autocorrelation functions through a driven damped harmonic oscillator analog for light scattering from persistent walks. The theory is validated through Monte Carlo simulations. Experimental evidence for the Doppler edge in three-dimensional (3D) living tissue is obtained using biodynamic imaging based on low-coherence interferometry and digital holography.

Intracellular Doppler Spectroscopy detects altered drug response in SKOV3 tumor spheroids with silenced or inhibited P-glycoprotein.

Intracellular Doppler spectroscopy is a form of low-coherence digital holography based upon Doppler detection of scattered light that measures drug response/resistance in tumor spheroids, xenografts, and clinical biopsies. Multidrug resistance (MDR) is one of the main causes of ineffective cancer treatment. One MDR mechanism is mediated by the MDR1 gene that encodes the drug efflux pump P-glycoprotein (Pgp). Overexpression of Pgp in some cancers is associated with poor chemotherapeutic response. This paper uses intracellular Doppler spectroscopy to detect Pgp-mediated changes to drug response in 3D tissues grown from an ovarian cancer cell line (SKOV3). The SKOV3 cell line was incrementally exposed to cisplatin to create a cell line with increased Pgp expression (SKOV3cis). Subsequently, MDR1 in a subset of these cells was silenced in SKOV3cis using shRNA to create a doxycycline inducible, Pgp-silenced cell line (SKOV3cis-sh). A specific Pgp inhibitor, zosuquidar, was used to study the effects of Pgp inhibition on the Doppler spectra. Increased drug sensitivity was observed with Pgp silencing or inhibition as determined by drug IC50s of paclitaxel-response of silenced Pgp and doxorubicin-response of inhibited Pgp, respectively. These results indicate that intracellular Doppler spectroscopy can detect changes in drug response due to silencing or inhibition of a single protein associated with drug resistance with important consequences for personalized medicine.

Reducing the impact of geometric errors in flow computations using velocity measurements.

Numerical blood flow simulations are typically set up from anatomical medical images and calibrated using velocity measurements. However, the accuracy of the computational geometry itself is limited by the resolution of the anatomical image. We first show that applying standard no-slip boundary conditions on inaccurately extracted boundaries can cause large errors in the results, in particular the pressure gradient. In this work, we therefore propose to augment the flow model calibration by slip/transpiration boundary conditions, whose parameters are then estimated using velocity measurements. Numerical experiments show that this methodology can considerably improve the accuracy of the estimated pressure gradients and 3D velocity fields when the vessel geometry is uncertain.