scholarly journals A DNA-based optical force sensor for live-cell applications

2021 ◽  
Author(s):  
Christina Jayachandran ◽  
Arindam Ghosh ◽  
Meenakshi Prabhune ◽  
Jonathan Bath ◽  
Andrew J. Turberfield ◽  
...  
Keyword(s):  
Force Sensor ◽  
Live Cell ◽  
Optical Force ◽  
Fluorescence Lifetime Imaging ◽  
Mechanical Forces ◽  
Actin Networks ◽  
Non Invasive ◽  
Novel Approach ◽  
Cell Force

Mechanical forces are relevant for many biological processes, from wound healing or tumour formation to cell migration and differentiation. Cytoskeletal actin is largely responsible for responding to forces and transmitting them in cells, while also maintaining cell shape and integrity. Here, we describe a novel approach to employ a FRET-based DNA force sensor in vitro and in cellulo for non-invasive optical monitoring of intracellular mechanical forces. We use fluorescence lifetime imaging to determine the FRET efficiency of the sensor, which makes the measurement robust against intensity variations. We demonstrate the applicability of the sensor by monitoring cross-linking activity in in vitro actin networks by bulk rheology and confocal microscopy. We further demonstrate that the sensor readily attaches to stress fibers in living cells which opens up the possibility of live-cell force measurements.

Quantifying effects of cyclic stretch on cell–collagen substrate adhesiveness of vascular endothelial cells

2018 ◽  
Vol 232 (5) ◽  
pp. 531-541 ◽  
Author(s):  
Ramin Omidvar ◽  
Mohammad Tafazzoli-Shadpour ◽  
Farbod Mahmoodi-Nobar ◽  
Shohreh Azadi ◽  
Mohammad-Mehdi Khani
Keyword(s):  
Endothelial Cells ◽  
Vascular Endothelial Cells ◽  
Cyclic Stretch ◽  
Cell Stiffness ◽  
Atomic Force ◽  
Novel Approach ◽  
Cell Tissue ◽  
Tensile Forces ◽  
Cell Force

Vascular endothelium is continuously subjected to mechanical stimulation in the form of shear forces due to blood flow as well as tensile forces as a consequence of blood pressure. Such stimuli influence endothelial behavior and regulate cell–tissue interaction for an optimized functionality. This study aimed to quantify influence of cyclic stretch on the adhesive property and stiffness of endothelial cells. The 10% cyclic stretch with frequency of 1 Hz was applied to a layer of endothelial cells cultured on a polydimethylsiloxane substrate. Cell–substrate adhesion of endothelial cells was examined by the novel approach of atomic force microscope–based single-cell force spectroscopy and cell stiffness was measured by atomic force microscopy. Furthermore, the adhesive molecular bonds were evaluated using modified Hertz contact theory. Our results show that overall adhesion of endothelial cells with substrate decreased after cyclic stretch while they became stiffer. Based on the experimental results and theoretical modeling, the decrease in the number of molecular bonds after cyclic stretch was quantified. In conclusion, in vitro cyclic stretch caused alterations in both adhesive capacity and elastic modulus of endothelial cells through mechanotransductive pathways as two major determinants of the function of these cells within the cardiovascular system.

scholarly journals Raman Imaging and Fluorescence Lifetime Imaging Microscopy for Diagnosis of Cancer State and Metabolic Monitoring

Cancers ◽  
2021 ◽  
Vol 13 (22) ◽  
pp. 5682
Author(s):  
Lucas Becker ◽  
Nicole Janssen ◽  
Shannon L. Layland ◽  
Thomas E. Mürdter ◽  
Anne T. Nies ◽  
...  
Keyword(s):  
Raman Spectroscopy ◽  
Fluorescence Lifetime ◽  
Drug Efficacy ◽  
Raman Microspectroscopy ◽  
Fluorescence Lifetime Imaging Microscopy ◽  
Fluorescence Lifetime Imaging ◽  
Lifetime Imaging ◽  
Non Invasive

Hurdles for effective tumor therapy are delayed detection and limited effectiveness of systemic drug therapies by patient-specific multidrug resistance. Non-invasive bioimaging tools such as fluorescence lifetime imaging microscopy (FLIM) and Raman-microspectroscopy have evolved over the last decade, providing the potential to be translated into clinics for early-stage disease detection, in vitro drug screening, and drug efficacy studies in personalized medicine. Accessing tissue- and cell-specific spectral signatures, Raman microspectroscopy has emerged as a diagnostic tool to identify precancerous lesions, cancer stages, or cell malignancy. In vivo Raman measurements have been enabled by recent technological advances in Raman endoscopy and signal-enhancing setups such as coherent anti-stokes Raman spectroscopy or surface-enhanced Raman spectroscopy. FLIM enables in situ investigations of metabolic processes such as glycolysis, oxidative stress, or mitochondrial activity by using the autofluorescence of co-enzymes NADH and FAD, which are associated with intrinsic proteins as a direct measure of tumor metabolism, cell death stages and drug efficacy. The combination of non-invasive and molecular-sensitive in situ techniques and advanced 3D tumor models such as patient-derived organoids or microtumors allows the recapitulation of tumor physiology and metabolism in vitro and facilitates the screening for patient-individualized drug treatment options.

scholarly journals Near Infrared Spectrum for Non-Invasive Glucose Measurement

2019 ◽  
Vol 11 (1) ◽  
Author(s):  
Ghozzi Dorsaf
Keyword(s):  
Glucose Concentration ◽  
Near Infrared ◽  
Glucose Monitoring ◽  
Nir Spectroscopy ◽  
Well Being ◽  
Glucose Measurement ◽  
Non Invasive ◽  
Novel Approach ◽  
Clarke Error Grid

This paper deals with novel approach for non-invasive glucose monitoring based on NIR spectroscopy. The technique was demonstrated on 300 human serums of different concentrations range of 08-297 mg/dl. In order to compare the proposed approach to a standard one, a regression analysis was performed and used to predict glucose concentration overall range of values. In vitro experiments showed a strong correlation between noninvasively device result and real glucose concentration. The correlation was 0.97 and Clarke error grid analysis showed that 97.33% of the measured fall within the clinically acceptable regions. Results showed that the created model can open a new path to a real-time, painless and a portable device that would well-being the lives of millions of diabetics in the world.

scholarly journals Label-free imaging of macrophage phenotypes and phagocytic activity in the human dermis in vivo using two-photon excited FLIM

2021 ◽  
Author(s):  
Marius Kröger ◽  
Jörg Scheffel ◽  
Evgeny A. Shirshin ◽  
Johannes Schleusener ◽  
Martina C Meinke ◽  
...  
Keyword(s):  
Fluorescence Lifetime Imaging ◽  
Label Free ◽  
Immune Effector ◽  
Effector Cells ◽  
Two Photon ◽  
Non Invasive ◽  
Human Dermis ◽  
Main Components

Macrophages (MΦs) are important immune effector cells that promote (M1 MΦs) or inhibit (M2 MΦs) inflammation and are involved in numerous physiological and pathogenic immune responses. Their precise role and relevance, however, is not fully understood because of the lack of non-invasive quantification methods. Here, we show that two-photon excited fluorescence lifetime imaging (TPE-FLIM), a label-free non-invasive method, can visualize MΦs in human dermis in vivo. We demonstrate in vitro that human dermal MΦs exhibit specific TPE-FLIM properties that distinguish them from the main components of the extracellular matrix and other dermal cells. We visualized MΦs, their phenotypes and phagocytosis in the skin of healthy individuals in vivo using TPE-FLIM. Additionally, machine learning identified M1 and M2 MΦs with a sensitivity of 0.88±0.04 and 0.82±0.03 and a specificity of 0.89±0.03 and 0.90±0.03, respectively. In clinical research, TPE-FLIM can advance the understanding of the role of MΦs in health and disease.

scholarly journals Validation of nasospheroids to assay CFTR functionality and modulator responses in cystic fibrosis

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Maite Calucho ◽  
Silvia Gartner ◽  
Paula Barranco ◽  
Paula Fernández-Álvarez ◽  
Raquel García Pérez ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
3D Culture ◽  
Live Cell ◽  
Functional Study ◽  
Wild Type ◽  
Non Invasive ◽  
Cftr Protein ◽  
Cystic Fibrosis Transmembrane ◽  
Fluid Reservoir

AbstractThe availability of a simple, robust and non-invasive in vitro airway model would be useful to study the functionality of the cystic fibrosis transmembrane regulator (CFTR) protein and to personalize modulator therapy for cystic fibrosis (CF) patients. Our aim was to validate a CFTR functional study using nasospheroids, a patient-derived nasal cell 3D-culture. We performed live-cell experiments in nasospheroids obtained from wild-type individuals and CF patients with different genotypes and phenotypes. We extended the existing method and expanded the analysis to upgrade measurements of CFTR activity using forskolin-induced shrinking. We also tested modulator drugs in CF samples. Immobilizing suspended-nasospheroids provided a high number of samples for live-cell imaging. The diversity observed in basal sizes of nasospheroids did not affect the functional analysis of CFTR. Statistical analysis with our method was simple, making this protocol easy to reproduce. Moreover, we implemented the measurement of inner fluid reservoir areas to further differentiate CFTR functionality. In summary, this rapid methodology is helpful to analyse response to modulators in CF samples to allow individualized treatment for CF patients.

Follicular fluid and assisted reproductive technology programs outcomes (literature review)

GYNECOLOGY ◽  
2020 ◽  
Vol 21 (6) ◽  
pp. 36-40
Author(s):  
Anna G. Burduli ◽  
Natalia A. Kitsilovskaya ◽  
Yuliya V. Sukhova ◽  
Irina A. Vedikhina ◽  
Tatiana Y. Ivanets ◽  
...  
Keyword(s):  
Clinical Practice ◽  
Assisted Reproductive Technology ◽  
Follicular Fluid ◽  
Economic Cost ◽  
Reproductive Technology ◽  
Art Programs ◽  
Vitro Fertilization ◽  
Non Invasive ◽  
Established Fact

The review presents data on metabolites in the follicular fluid (FF) from the perspective of reproductive medicine and their use in order to predict outcomes of assisted reproductive technology (ART) programs. It considers various components of this biological medium (hormones, lipids, melatonin, etc.) with an assessment of their predictive value in prognosis of the effectiveness of in vitro fertilization (IVF) programs. The data on experimental directions in this field and the prospects for their use in clinical practice are presented. The article emphasizes that the growing clinical need and the unsolved problem of increasing the effectiveness of ART programs determine the need for further studies of the FF composition. Materials and methods. The review includes data related to this topic from foreign and Russian articles found in PubMed which were published in recent years. Results. Given the established fact of a direct effect of FF composition on growth and maturation of oocytes, and further, on the fertilization process, various FF metabolites are actively investigated as non-invasive markers of quality of oocytes/embryos. The article provides data on the experimental directions in this field and the prospects for their use in clinical practice. However, clinical studies of a relation between various FF metabolites levels and outcomes of IVF programs are contradictory. Conclusion. Owing large economic cost for treatment of infertility with IVF, there is need for expansion and intensification of studies to identify and use reliable predictors in prognosis of ART programs outcomes.

Picosecond acoustics in vegetal cells: Non-invasive in vitro measurements at a sub-cell scale

Ultrasonics ◽  
2010 ◽  
Vol 50 (2) ◽  
pp. 202-207 ◽  
Author(s):  
B. Audoin ◽  
C. Rossignol ◽  
N. Chigarev ◽  
M. Ducousso ◽  
G. Forget ◽  
...  
Keyword(s):  
Non Invasive ◽  
In Vitro Measurements

scholarly journals Non-invasive skin sampling of tryptophan/kynurenine ratio in vitro towards a skin cancer biomarker

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Skaidre Jankovskaja ◽  
Johan Engblom ◽  
Melinda Rezeli ◽  
György Marko-Varga ◽  
Tautgirdas Ruzgas ◽  
...  
Keyword(s):  
Skin Cancer ◽  
Relative Increase ◽  
Cancer Biomarker ◽  
Cancer Diagnostics ◽  
Sampling Time ◽  
Skin Permeability ◽  
Experimental Conditions ◽  
Non Invasive

AbstractThe tryptophan to kynurenine ratio (Trp/Kyn) has been proposed as a cancer biomarker. Non-invasive topical sampling of Trp/Kyn can therefore serve as a promising concept for skin cancer diagnostics. By performing in vitro pig skin permeability studies, we conclude that non-invasive topical sampling of Trp and Kyn is feasible. We explore the influence of different experimental conditions, which are relevant for the clinical in vivo setting, such as pH variations, sampling time, and microbial degradation of Trp and Kyn. The permeabilities of Trp and Kyn are overall similar. However, the permeated Trp/Kyn ratio is generally higher than unity due to endogenous Trp, which should be taken into account to obtain a non-biased Trp/Kyn ratio accurately reflecting systemic concentrations. Additionally, prolonged sampling time is associated with bacterial Trp and Kyn degradation and should be considered in a clinical setting. Finally, the experimental results are supported by the four permeation pathways model, predicting that the hydrophilic Trp and Kyn molecules mainly permeate through lipid defects (i.e., the porous pathway). However, the hydrophobic indole ring of Trp is suggested to result in a small but noticeable relative increase of Trp diffusion via pathways across the SC lipid lamellae, while the shunt pathway is proposed to slightly favor permeation of Kyn relative to Trp.

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