Phase Contrast Research Articles

Quantitative differential phase contrast imaging system using FPGA for cellular imaging

Abstract Quantitative differential phase contrast (DPC) microscopy, a label-free method, allows cell analysis without needing labels. We introduce a compact, FPGA-based qDPC microscope module that enhances imaging efficiency by controlling essential functions such as digital illumination, rapid image capture, and phase reconstruction. This module integrates seamlessly with existing commercial microscopes, eliminating the need for additional optical components. A deep learning model is implemented to improve functionality, converting intensity images to phase data and requiring only a single image pair for isotropic phase reconstruction. Our approach holds promise for enhancing cell imaging and virology methodologies for epidemiological investigations. Our model demonstrates high accuracy in predicting quantitative phase maps, with a Structural Similarity Index (SSIM) of ~0.9 and a
Peak Signal-to-Noise Ratio (PSNR) of ~30. The FPGA-driven system achieves a tenfold increase in imaging speed, is advantageous for real time cell imaging, and offers convenience-through its unified programming language and hardware user interface. Additionally, our system shows significant potential for practical and scalable applications in basic research and clinical settings.

Differential Phase Contrast Imaging to Predict MSC Immune Function.

Mesenchymal stromal cells (MSCs) exhibit significant immunomodulatory potential, making them promising candidates for cell-based therapies in autoimmune and inflammatory diseases. However, the heterogeneity of MSC cultures and a lack of robust, predictive potency assays have hindered their clinical translation. In this study, the potential of single-cell morphological imaging during MSC expansion is explored as a method to estimate indoleamine-2,3-dioxygenase (IDO) protein and enzyme activity, a common immunosuppressive capacity measure. Fluorescence and label-free quantitative differential phase contrast (qDPC) imaging is employed to non-invasively extract morphological features from live MSCs during biomanufacturing with machine learning (ML) regression models to predict single cell IDO activity. qDPC imaging characterization is extended to estimate a previously established consensus model of MSC potency based on their IDO activity and immune suppression on T cells. These findings establish a foundation for scalable, non-destructive monitoring of MSC immunomodulatory capacity, facilitating the future development of quality control strategies for MSC manufacturing and clinical applications.

Mesenchymal Stem Cell-conditioned Medium Attenuated CoCl2-induced Injury of Renal Tubular Epithelial Cells by Inhibiting NCOA1, HIF-1α, and Sox9.

Renal interstitial fibrosis (RIF) constitutes the ultimate pathological alteration in nearly all chronic kidney diseases (CKD). Mesenchymal stem cell conditioned medium (MSC-CM) exhibits an alleviating impact on renal fibrosis; however, the underlying mechanism remains unclear. The objective of this study was to explore whether MSC-CM regulates the expression of α-smooth muscle actin (α-SMA), Transforming growth factor-β1 (TGF-β1), Hypoxia-inducible factor-1α (HIF-1α), Nuclear receptor coactivators (NCOA1), and SRY-related high mobility (Sox9). Rat renal tubular epithelial cells (RTECs), NRK-52E, were treated with diverse concentrations of Cobalt chloride (CoCl2) for 24 hours. The survival rate and protein expression of NRK-52E cells exposed to different concentrations of CoCl2 were determined to identify the final concentration. Three groups of NRK-52E cells were employed in the experiment: the normal control group, the 400 μM CoCl2 group, and the MSC-CM + 400 μM CoCl2 group. The cell morphology was observed by an inverted phase contrast microscope and scanning electron microscope, and the protein expressions of α-SMA, TGF-β1, HIF-1α, NCOA1, and Sox9 were detected. The microscopic findings demonstrated that MSC-CM was able to decrease the degree of cytochemical hypoxia damage in NRK-52E cells induced by CoCl2. Immunofluorescence and Western blot analyses also affirmed a similar tendency. The upregulation of α-SMA, TGF-β1, HIF-1α, NCOA1, and Sox9 triggered by CoCl2 could be inhibited following MSC-CM intervention. Our findings indicate that MSC-CM exerts a protective effect on RTECs by down-regulating α-SMA, TGF-β1, HIF-1α, NCOA1, and Sox9.

Origanum syriacum Induces Apoptosis in Lung Cancer Cells by Altering the Ratio of Bax/Bcl2.

The lung cancer is the leading cause of death worldwide. Although methods such as surgery, chemotherapy, radiotherapy, and immunotherapy are used for treatment, these treatments are sometimes inadequate. In addition, the number of chemotherapeutic agents used is very limited, and it is very important to use new natural agents that can increase the effect of these methods used in treatment. The present study was designed to determine the suppression of proliferation and induction of apoptosis activities and phenolic content of Origanum syriacum methanol extract (OsME) on lung cancer cells (A549). For this purpose, the cell viability of A549 cells exposed to OsME was first determined. The morphological changes of the cell were observed by an inverted phase contrast microscope. Moreover, the percentage of apoptotic and necrotic cells was determined by FACS with AnnexinV/Propodium iodide staining. Additionally, proapoptotic Bax and antiapoptotic Bcl-2 mRNA levels were determined by Real-time PCR. Phenolic compounds of OsME were detected by LC-MS-MS. It was observed that the viability and proliferation of lung cancer cells decreased after the treatment of different concentrations of OsME. At a concentration of 200 mg/ml of OsME, most of the cell membrane structures were observed to disintegrate. Meanwhile, a 25 μg/ml concentration of OsME increased the Bax expression and percentage of late apoptotic cells. Vanillic acid and luteolin were identified as the main phenolic compounds of OsME. OsME exhibited antiproliferation activity on A549 cells and induced apoptosis at low doses.

Heterologous interference between Mayaro and Chikungunya viruses in Vero cells.

Heterologous interference between Mayaro and Chikungunya viruses in Vero cells.

Green synthesis of magnesium oxide nanoparticles from Solanum lycopersicum and evaluation of their antidiabetic effect in rats

This study investigates the green synthesis of magnesium oxide nanoparticles (MgO NPs) using Solanum lycopersicum (tomato) extract and evaluates their antidiabetic effect in rats. Diabetic neuropathy (DN) being the common and debilitating complication of diabetes mellitus was therapeutically targeted. The synthesized MgO NPs were characterized using UV-Vis spectrophotometry, phase contrast microscopy, FTIR, SEM-EDS, particle size and zeta potential analyser, and TEM. The antidiabetic neuropathic effect of the Solanum lycopersicum-derived MgO NPs was assessed in streptozotocin-induced diabetic rats by measuring biochemical parameters (blood glucose, HbA1c, and insulin levels), performing histopathological analysis of various brain regions, and conducting behavioral tests to evaluate nociception and motor activity. The results indicate the successful green synthesis of MgO NPs from Solanum lycopersicum and demonstrate their potential to ameliorate diabetic neuropathy in rats, suggesting a promising antidiabetic effect.

Controllable Preparation of Millimeter-Scale In-Plane Charged Domain Wall in a Lead-Free Ferroelectric Film.

Ferroelectric domain walls are emerging as active components in nanoelectronics, offering a transformative paradigm for non-volatile memory and logic technologies. In particular, in-plane charged domain walls stand out for their potential to support unique functionalities, such as quantum confinement and tunneling effects, unlocking new possibilities for advanced device applications. Here, a facile approach is developed for fabricating large-scale continuous in-plane charged domain walls in Na0.5Bi0.5TiO3 ferroelectric films through thermally optimized growth control. Using Differential Phase Contrast Scanning Transmission Electron Microscopy, an electric field fluctuation is identified across the charged domain wall. This fluctuation is accompanied by oxygen octahedron distortions, which allow the lattice to redistribute charges, as confirmed by integrated Differential Phase Contrast analyses. Additionally, Electron Energy Loss Spectroscopy reveals variations in titanium oxidation states across the domain wall, which compensates for the charge accumulation and enhances the stability of the charged domain wall. This work presents a wafer-scale fabrication strategy for in-plane charged domain walls and provides atomic-scale insights into their stabilization mechanisms, offering a foundation for their integration into domain-wall-based nanodevices and on-chip applications.

MRI Assessment of Energy Loss Within the Thoracic Aorta and Its Impact on Cardiac Function in Fontan Patients After Aortic Reconstruction.

In Fontan patients undergoing aortic reconstruction, concerns regarding the impact of aortic function on ventricular function exist. 4D Flow MRI was used to compare energy loss (EL) within the thoracic aorta in patients with and without aortic reconstruction. Retrospective case control. Eighty-nine patients underwent 4D Flow MRI: group A (n = 36), Fontan patients without aortic reconstruction (9.9 (1.0-26.7) years since Fontan completion); group B (n = 42), Fontan patients with aortic reconstruction (11.8 (1.0-26.4) years since Fontan completion); and group C (n = 11), patients with biventricular circulation without aortic reconstruction. Balanced SSFP cine and time-resolved 3D phase contrast (4D Flow) sequences at 1.5 T or 3 T. Peak and average aortic EL in the thoracic aorta as well as peak aortic velocity and flow volume were assessed. Correlations between EL indexed to aortic forward flow volume and volumetric ventricular parameters and peak aortic velocity were assessed. Kruskal-Wallis test, chi-square test and Spearman's correlation coefficient were used. Peak and average EL were significantly larger in group B than in groups A and C (peak EL (mW); A: 1.45 (0.22-9.81), B: 3.09 (0.51-12.49), C: 2.10 (1.20-3.45); average EL (mW); A: 0.46 (0.07-2.63), B: 1.13 (0.13-4.67), C: 0.76 (0.40-1.98)). Group B had significantly larger ventricular end-diastolic volume index (EDVi, 108 mL/m2) and end-systolic volume index (ESVi, 53 mL/m2), significantly lower ejection fraction (EF, 51%) and significantly greater end-diastolic myocardial mass (MM, 50 g/m2) of the systemic ventricle than group A (EDVi: 86 mL/m2, ESVi: 34 mL/m2, EF: 58%, end-diastolic MM: 43 g/m2). In Fontan patients, indexed average aortic EL correlated positively with aortic peak velocity (R = 0.68) and with years after Fontan completion (R = 0.60). Fontan patients who underwent aortic reconstruction had increased aortic EL, even in the absence of significant residual aortic stenosis. Level 3. Stage 3.

Aortic Function in a Longitudinal 4D Flow MRI Study in Marfan Syndrome Patients Receiving Resveratrol.

New treatment strategies are required to reduce aortic events in Marfan syndrome (MFS). Resveratrol is a dietary supplement that intervenes in aortic wall cellular metabolism and may benefit MFS patients. To evaluate whether treatment with Resveratrol affects aorta hemodynamics derived from 4D flow MRI in MFS. Prospective single-arm open-label multicenter trial. 46 MFS patients (mean age 36 ± 9 years, 23 female), 20 with and 26 without a history of aortic root surgery (RR and native MFS), and 25 age- and sex-matched healthy controls. 3T, temporally resolved 3D phase contrast (4D flow MRI) and 3D mDixon sequences. MFS patients underwent 4D flow MRI before (baseline) and after 1 year of 500 mg daily Resveratrol treatment (follow-up). Velocity magnitude and wall shear stress (WSS) in six regions in the thoracic aorta were evaluated. Incidence maps showing abnormal hemodynamics compared to healthy controls were generated. Pulse wave velocity (PWV) was assessed in 45 subjects. The relationships between MRI parameters and annual aortic growth from the 3D mDixon scans and age were investigated. Student's paired and unpaired t-tests, Fisher's exact tests, McNemar's exact test, and Pearson correlation coefficients (r). Statistical significance was defined as p < 0.05. No significant changes in any of the hemodynamic parameters were observed: inner descending aorta WSS (Pa), native: from 1.08 ± 0.21 to 1.08 ± 0.17, p = 0.818, RR: from 1.02 ± 0.29 to 0.98 ± 0.21, p = 0.270; velocity (m/s), native: from 0.64 ± 0.10 to 0.65 ± 0.10, p = 0.359, RR: 0.68 ± 0.15, to 0.67 ± 0.12, p = 0.629; abnormally directed WSS incidence: native: from 17 (65%) to 14 (54%), p = 0.453, RR: 13 (65%) to 13 (65%), p = 1.000; PWV (m/s) native: from 7.7 ± 1.9 to 7.9 ± 1.8, p = 0.582, RR: 9.2 ± 1.6 to 8.2 ± 3.2, p = 0.184. PWV correlated with age in RR MFS patients (r = 0.59, n = 19). No significant changes in aortic hemodynamics derived from 4D flow MRI were observed after 1 year of Resveratrol treatment. 1. Stage 4.

Phase Imaging Methods in the Scanning Transmission Electron Microscope.

Scanning transmission electron microscopy (STEM) has become an essential tool for investigating materials, providing detailed characterization at nano and atomic scales. By combining subangstrom resolution Z-contrast imaging with analytical X-ray and electron spectroscopies, STEM allows the visualization of atomic arrangements, crystal defects, and interfaces, understanding sample properties and correlating these with the functionalities of materials and devices. Recent advancements in phase imaging techniques, including differential phase contrast (DPC) and electron ptychography, have further enhanced STEM capabilities. These methods allow direct imaging of electromagnetic fields, the study of beam-sensitive materials with high dose efficiency, or resolving the 3D structure of materials, proving invaluable for investigating intricate nanoscale phenomena. This review introduces phase imaging methods in the STEM and explores how the most recent innovations are driving progress in nanoscience, deepening material insights and shaping next-generation applications in electronics, energy storage, and catalysis.

Revealing neurovascular coupling at a high spatial and temporal resolution in the living human retina.

Neurovascular coupling (NVC) regulates local blood flow in response to neuronal activity, yet its precise characterization at the capillary level has been hindered by the lack of a noninvasive, high-resolution imaging method. Here, we introduce the adaptive optics rolling slit ophthalmoscope, a unique noninvasive, label-free, high-speed, cellular-resolution clinical imaging system for assessing retinal NVC in the living human eye. Using an off-axis phase contrast approach combined with camera-based confocal slit gating, our method provides large field-of-view imaging of arterial and venular walls, enabling the study of vascular dynamics with unprecedented spatiotemporal precision. Our findings highlight that this level of precision is essential for accurately distinguishing NVC-driven vasodilation from spontaneous fluctuations, such as vasomotion and the cardiac cycle. By bridging the gap between fundamental neurovascular research and clinical applications, this approach offers a powerful tool for neuroscience research and early disease detection and monitoring of neurodegenerative and vascular disorders.

The Effect of Rosavin, a Characteristic Compound of Rhodiola rosea, on BMP-2 Induction and Osteoblast Proliferation In Vitro

Rosavin, a glycoside isolated from Rhodiola rosea, exhibits various biological activities, including potential modulation of metabolic pathways. Despite promising findings in animal models, its effects on many human bone cells remain unexplored. This study aimed to investigate, for the first time, the in vitro effects of rosavin on human osteoblasts (HOBs), focusing on BMP-2 expression, cell morphology, and culture confluence as indicators of osteogenic activity. HOB cultures were treated with 50 µM or 100 µM rosavin for 21 days. BMP-2 expression was measured by ELISA, collagen production was assessed via Sirius Red staining, and cell morphology and confluence were evaluated using phase-contrast microscopy. A significant increase in BMP-2 expression was observed in the 100 µM rosavin group compared to the mineralization control (p &lt; 0.05), particularly on days 14 and 21. Both rosavin-treated groups exhibited higher confluence than controls, with the 50 µM group showing unexpectedly greater confluence than the 100 µM group. Rosavin at 50 µM also promoted a cuboidal morphology characteristic of active HOBs. The presence of collagen validated both the successful progression of the mineralization process and the correct implementation of the experimental protocol. Rosavin enhances BMP-2 expression and supports HOB proliferation and morphological maturation in vitro. These findings suggest its potential as a supportive agent in the prevention or treatment of metabolic bone diseases. Further research is necessary to determine its bioavailability, safety profile, and therapeutic relevance in clinical settings.

Free-breathing cardiovascular magnetic resonance flow quantification can be an alternative to standard breath-holding approach

Cardiovascular magnetic resonance (CMR) evaluation of valvular heart disease is an important diagnostic tool when echocardiography is inconclusive. Phase contrast flow quantification is usually performed during breath hold (BH), which can be challenging in patients suffering from dyspnea and heart failure. The purpose of the present study is to compare a free-breathing (FB) with the conventional BH approach for flow quantification in the aortic, pulmonary and tricuspid valves in 20 healthy subjects (HS) and 25 patients with tricuspid regurgitation (TR). Aortic (AoFF) and pulmonary forward flow volume (PuFF), and tricuspid inflow volume (TrIF) were evaluated. Mean, standard deviation (SD) and limits of agreement (LoA) were calculated. There were good agreements between phase contrast flow volumes obtained by FB and BH approach. Mean difference ± SD / LoA for AoFF during BH versus FB were 1 ± 6 / -10 to 13 ml. The corresponding for PuFF were 1 ± 6 / -11 to 13 ml, and for TrIF − 3 ± 6 / -15 to 9 ml, respectively. Thus, free-breathing CMR flow acquisition can be an important alternative in the assessment of stroke volume, valvular regurgitant volume and be useful in all patients with difficulties to hold their breath.

How X-ray dark-field imaging relates to small-angle X-ray scattering measurements

Dark-field (DF) imaging is a recent X-ray imaging modality which is promising because it gives access to information not resolved in conventional transmission X-ray imaging. The DF technique was first introduced as a loss of visibility of the grating interferometry modulations. DF signal is now measured with all the different X-ray phase contrast setups such as beam tracking or modulation-based imaging. Using a dedicated setup [Magnin et al. (2023). Opt. Lett. 48, 5839–5842], we present in the present article combined measurements of small-angle X-ray scattering and DF signal on the same material. We confirm that DF imaging is sensitive to multiple refraction from a sample, as can be found in the literature on lung imaging, but we show that the DF signal is also sensitive to scattering events. Finally, we measure a porous membrane that creates both types of signal (scattering and refraction), showing that, contrary to existing models, it is difficult to be quantitative about DF.

Comparative cardiomyocyte differentiation potential of rat adipose-derived mesenchymal stem cells from two anatomical sites: metabolomic profiling and pathway analysis

BackgroundAdipose-derived mesenchymal stem cells (AD-MSCs) have emerged as a promising source for cardiac regenerative therapy due to their multipotency and ease of isolation. However, the impact of anatomical origin on their cardiomyocyte differentiation potential remains unclear. Metabolic analysis provides valuable real-time insights into the cellular metabolic state, capturing dynamic changes in metabolite concentrations that reflect both internal cellular mechanisms and external stimuli. This approach allows us to identify specific metabolic pathways activated during cardiomyocyte differentiation, offering a deeper understanding of how the anatomical origin of stem cells influences their differentiation potential and metabolic flexibility. Such insights are critical for optimizing stem cell-based therapies for cardiac regeneration. This study aimed to compare the differentiation capacity of AD-MSCs derived from peri-ovarian and peri-renal adipose tissue, with a focus on metabolic adaptations during cardiomyocyte differentiation.MethodsAD-MSCs were isolated from peri-ovarian and peri-renal fat of Sprague-Dawley rats and characterized by morphology, immunophenotyping, and multilineage differentiation potential. Cardiomyocyte differentiation was induced using 5-azacytidine, and morphological changes were assessed via phase-contrast microscopy and immunofluorescence staining for cardiac troponin T (cTnT). Untargeted metabolomic profiling was performed using gas chromatography-mass spectrometry (GC-MS), followed by principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and pathway enrichment analysis.ResultsBoth peri-ovarian and peri-renal AD-MSCs exhibited similar fibroblast-like morphology, MSC-specific marker expression (CD44, CD90, CD29), and multilineage differentiation potential. Following cardiomyocyte induction, both groups displayed morphological changes indicative of differentiation and strong cTnT expression. Metabolomic analysis of the cardiogenic differentiation samples identified distinct metabolic adaptations between the two AD-MSC sources. Peri-ovarian AD-MSCs exhibited a broader metabolic reprogramming, with increased engagement of glycolysis, fructose metabolism, glycerolipid metabolism, and the TCA cycle, suggesting enhanced metabolic flexibility and energy efficiency. In contrast, peri-renal AD-MSCs relied more on galactose metabolism, indicating an alternative energy strategy during differentiation.ConclusionThe anatomical origin of AD-MSCs influences their metabolic landscape during cardiomyocyte differentiation. Peri-ovarian AD-MSCs demonstrated greater metabolic adaptability, potentially favoring their differentiation capacity, making them a promising candidate for cardiac regenerative applications.

Assembly of Branched Chain Amino Acids to Toxic Fibrils may be Related to Pathogenesis of Maple Syrup Urine Disease.

Inborn errors of metabolisms (IEMs) are group of diseases caused by mutations in single genes, leading to buildup of metabolites, toxic or disrupt normal cellular function. The etiological relation of metabolic disorders has been uncovered through study of metabolite amyloids. Various metabolites that accumulate in IEMs have been reported to self-assemble into organized structures. These structures exhibit similar physicochemical properties as proteinaceous amyloid fibrils. Our study illustrates the aggregation properties of branched chain amino acids (BCAAs), isoleucine, leucine, and valine that accumulate in maple syrup urine disease (MSUD) to investigate their propensities to assemble into amyloid-like fibrils. The structural morphologies of BCAAs are studied via microscopic techniques. Further, characterization techniques are employed to understand the physicochemical properties of the self-assemblies and their underlying mechanism. The amyloid-like nature of these aggregates is confirmed using thioflavin T and congo red assays. The (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay reveals BCAAs are cytotoxic and significantly decrease cell viability. This study plays a key role in understanding the physicochemical properties of MSUD in the context of amyloid diseases, possibly paving the way for the development of its therapeutic solutions in the future.

Modifications to the membrane filter method with phase-contrast microscopy (PCM) for fibre-counting to support a new occupational exposure limit value.

Modifications to the membrane filter method with phase-contrast microscopy (PCM) for fibre-counting to support a new occupational exposure limit value.

Radial-balanced phase transfer functions for accurate retrieval in quantitative differential phase contrast microscopy

Radial-balanced phase transfer functions for accurate retrieval in quantitative differential phase contrast microscopy

The cochlear apex demystified: Implications from synchrotron radiation phase-contrast imaging and microscopy for cochlear implantation.

Due to the complex organization of the human cochlear apex, further analysis of the tonotopic relationship between the organ of Corti (OC) and spiral ganglion (SG) is required in relation to cochlear implantation. In this study, the human SG nerve fiber organization and ultrastructure were assessed using semi-thin light microscopy sectioning and three-dimensional (3D) synchrotron radiation phase-contrast imaging (SR-PCI). A fresh human temporal bone underwent high-resolution SR-PCI with a dual-detector system. Orthogonal sectioning, cropping, and tissue segmentation were used to create high-resolution 3D reconstructions. Peripheral dendrites were traced from the basilar membrane to the SG, and a tonotopic map was constructed using Greenwood's function. Results were compared and validated against novel high-resolution microscopy data of a sectioned human cochlea. Only the basal and initial middle turn of the cochlea displayed a well-defined Rosenthal's canal (RC), and after 450 degrees, this converged into a central modiolar space. The OC and SG tonotopic maps remained closely aligned for angular depths up to approximately 650 degrees, after which the SG frequencies became significantly more spatially compact relative to the OC. In the central modiolus, the apical 1.37 mm of the SG contained over four octaves of tonotopic representation. In comparison, the compressed apical SG represented 9.6 mm of the OC (28% of the overall length) over the same tonotopic range. These results were validated with microscopy, which revealed that this apical SG contained around 8000 neurons and represented 960 inner hair cells along the OC. This is the first study to present the detailed cellular organization and 3D tonotopic arrangement of the human SG within the central modiolus. For low frequency stimulation, rate-based coding may be required to augment tonotopic mapping in the compressed SG regions. In addition, the OC tonotopic map has significantly less compression and could potentially be targeted directly for place-based coding.

Real-space observation of polarization induced charges at nanoscale ferroelectric interfaces.

Unique electrical properties emerging at nanoscale ferroelectric interfaces originate from the polarization induced charges. However, real-space characterization of polarization induced charges at nanoscale ferroelectric interfaces has been extremely challenging. Here, directly observing the nanoscale electric field by tilt-scan averaged differential phase contrast scanning transmission electron microscopy enables us to measure the spatially varying total charge density profiles across both head-to-head and tail-to-tail domain walls in a ferroelectric crystal. Combined with atomic column displacement measurements, the spatial distribution of polarization bound charges and screening charges across the domain walls can be disentangled. Our results reveal the true charge states of the nanoscale ferroelectric interfaces, providing an opportunity for experimentally exploring the interplay between atomic-scale local polarization structures and their charge states in ferroelectric interfaces.