Confocal Raman data analysis enables identifying apoptosis of MCF-7 cells caused by anticancer drug paclitaxel

Confocal Raman microscopy is a noninvasive, label-free imaging technique used to study apoptosis of live MCF-7 cells. The images are based on Raman spectra of cells components, and their apoptosis is monitored through diffusion of cytochrome c in cytoplasm. K-mean clustering is used to identify mitochondria in cells, and correlation analysis provides the cytochrome c distribution inside the cells. Our results demonstrate that incubation of cells for 3 h with 10 mu M of paclitaxel does not induce apoptosis in MCF-7 cells. On the contrary, incubation for 30 min at a higher concentration (100 mu M) of paclitaxel induces gradual release of the cytochrome c into the cytoplasm, indicating cell apoptosis via a caspase independent pathway. (C) 2013 Society of Photo-Optical Instrumentation Engineers (SPIE) [DOI: 10.1117/1.JBO.18.5.056010

Gas permeability: a new quantitative method to assess endodontic leakage

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Elucidating the LiFePO(4) air aging mechanism to predict its electrochemical performance

International audienceAlthough LiFePO4-based chemistry has been extensively studied and developed in the last decade due to its promise for the next generation of Li-ion battery applications, the impact of ambient air exposure and the concomitant aging mechanism is still a controversial matter. In the present study, we describe quantitatively the aging process in terms of the successive phase formation, distribution and iron local environments. Formation and growth of a disordered ferric phosphate phase are directly observed for the first time from the surface of particles toward the core, preceding the crystallization of tavorite LiFePO4(OH) as observed through the combination of amorphous phase quantification by XRD, and by Mössbauer and Electron Energy Loss spectroscopies. Structural and electrochemical characterization prove that the amorphous ferric phosphate formed in the early aging stage exhibits already tavorite composition and structure at the local scale and shed light on a crucial step of the aging mechanism. Based on the correlation we establish here between the amounts of Fe(III) and hydroxyls groups present in aged samples and specific capacity of the corresponding electrodes, we show that the degradation of active material upon storage and the electrochemical performance can be predicted easily through simple TGA measurements

Towards the Standardization of Mesenchymal Stem Cell Secretome-Derived Product Manufacturing for Tissue Regeneration

Mesenchymal stem cell secretome or conditioned medium (MSC-CM) is a combination of biomolecules and growth factors in cell culture growth medium, secreted by mesenchymal stem cells (MSCs), and the starting point of several derived products. MSC-CM and its derivatives could be applied after injuries and could mediate most of the beneficial regenerative effects of MSCs without the possible side effects of using MSCs themselves. However, before the clinical application of these promising biopharmaceuticals, several issues such as manufacturing protocols and quality control must be addressed. This review aims to underline the influence of the procedure for conditioned medium production on the quality of the secretome and its derivatives and highlights the questions considering cell sources and donors, cell expansion, cell passage number and confluency, conditioning period, cell culture medium, microenvironment cues, and secretome-derived product purification. A high degree of variability in MSC secretomes is revealed based on these parameters, confirming the need to standardize and optimize protocols. Understanding how bioprocessing and manufacturing conditions interact to determine the quantity, quality, and profile of MSC-CM is essential to the development of good manufacturing practice (GMP)-compliant procedures suitable for replacing mesenchymal stem cells in regenerative medicine

Confocal Raman Microscope for the study of anti-cancer drug delivery by dental pulp stem cells

International audienceUsing stem cells as anticancer drug carriers aims for targeting tumors and reducing the side effects of chemotherapy. Coupling the multipotential capacities of stem cells with the nanomedical properties of the drugs provides an attractive platform for cancer therapy. Raman microscopy with its high spatial resolution and sensitivity is used as a unique tool to trace drugs and cellular components without labeling. Drug uptake, transfer, and release by stem cells, in addition to the effect on tumor cells were studied by Confocal Raman Microscope.Moreover, to further understand the intra-cellular effect of paclitaxel drug exposure and stem cell treatment upon the stem and cancer cells, we have traced different enzymes and biomarkers intra-cellularly. By software-aided Raman analysis, we were able to detect and measure the enzymes: cytochrome c in the mitochondria and Matrix Metalloproteinase MMP-1, as cellular markers pre and post-treatment. This was performed by calculating the Pearson correlation coefficient for the selected reference spectrum to the spectral correlation matrix. We investigated the cellular viability by means of confocal Raman microscope, also verified by apoptosis and cytotoxic assays.Our results prove the promising use of dental pulp stem cells as cellular vehicles, and significant phenotypic change in cellular marker expression after treatment with paclitaxel. Biochemical imaging of intracellular drug uptake using Confocal Raman would help to have a better understanding of drugs mechanism of action and cellular behavior, including cellular interaction mechanisms

Dental pulp stem cells used to deliver the anticancer drug paclitaxel

International audienceAbstractBackground: Understanding stem cell behavior as a delivery tool in cancer therapy is essential for evaluating their future clinical potential. Previous in-vivo studies proved the use of mesenchymal stem cells (MSCs) for local delivery of the commonest anticancer drug, paclitaxel (PTX). Dental pulp is a relatively abundant noninvasive source of MSCs. We assess dental pulp stem cells (DPSCs), for the first time, as anticancer drug carriers. Confocal Raman microscopy is a unique tool to trace drug and cell viability without labeling.Methods: Drug uptake and cell apoptosis are identified through confocal Raman microscope. We traced translocation of cytochrome c enzyme from the mitochondria, as a biomarker for apoptosis, after testing both cancer and stem cells. The viability of stem cells was checked by means of confocal Raman microscope and by cytotoxicity assays.Results: In this study, we prove that DPSCs can be loaded in vitro with the anticancerous drug without affecting their viability, which is later released in the culture medium of breast cancer cells (MCF-7 cells) in a time-dependent fashion. The induced cytotoxic damage in MCF-7 cells was observed consequently after PTX release by DPSCs. Additionally, quantitative Raman images of intracellular drug uptake in DPSCs and MCF-7 cells were obtained. Cytotoxic assays prove the DPSCs to be more resistant to PTX as compared to bone marrow-derived MSCs, provided similar conditions.Conclusions: Applications of dental stem cells for targeted treatment of cancer could be a revolution to reduce morbidity due to chemotherapy, and to increase the efficacy of systemic cancer treatment.Keywords: Raman spectroscopy, Dental pulp stem cells, Cancer, Paclitaxel, Apoptosi

Confocal Raman Microscopy to image targeted chemotherapy

International audienceThe Side effects of chemotherapy in cancer treatments are unavoidable due to the lack of anticancer drug’s specificity and it has a painful impact on patient quality of life. Over the past 30 years, increasing efforts is done to optimize chemotherapy dosing to reduce drug toxicity while increasing its efficacy. A new study proves stem cells can act as a drug reservoir and they will release anticancer drug in its original form in nearby area of cancer cells. Stem cells, due to their capacity to uptake drug, can control the drug toxicity. Dental Pulp Stem Cells DPSCs are able to uptake Paclitaxel PTX and could release it in the culture medium gradually. The conditioned culture medium (culture medium plus released PTX from DPSC) is transferred to the breast cancer cells MCF7. Visualizing the drug uptake intracellular could provide us mechanism of action of the drug. Confocal Raman microscopy as a noninvasive label free method is being used to trace drugs intracellular [1,2]. Applying Confocal Raman Microscopy, anticancer drug uptake by MCF7 is imaged. Surprisingly MCF7 - without any direct contact with PTX- showed drug uptake. It proves the stem cells carry and deliver anticancer drug without its modification. It could be a revolution in chemotherapy to avoid the side effects and increase the drug efficacy.[1] H. Salehi et al. Label-free detection of anticancer drug paclitaxel in living cells by Confocal Raman Microscopy. Applied physics letter. 2013, 102, 113701.[2] H. Salehi et al. Confocal Raman data analysis to comparison of apoptotic and non-apoptotic MCF-7 cells caused by anticancer drug paclitaxel. Journal of Biomedical Optics. 2013,18(5), 056010

Innovative device for in-vivo quantitative assessment of different wire materials using 3D models

Orthodontic archwire materials are changing rapidly in our time. Because an ideal archwire material has not yet been found, assessment of the most effectual properties of different archwires including working range and its effect on root resorption is still an unmet clinical need. Unfortunately, most clinical studies lack standardization of subjects and force. Performing a standardized in-vivo quantitative assessment for the working range of different orthodontic archwires and its effect on root resorption was not previously attempted. This is the first study to quantitatively compare and assess these properties in a standardized split mouth design. In this work, a sample of 10 Wister rats in which one of the two upper incisors was randomly selected to receive a force of 25gram-force using CuNiTi wire while the control side received the same amount of force with NiTi. CBCT was done before and after force application to compare the working range and root resorption. The results showed a significant difference in the working range between groups favoring a larger amount of displacement in the CuNiTi(p < 0.05). Consequently, this novel approach can open new avenues for experimenting with orthodontic wire materials based on standardized measurements of not only mechanical but also biological effects. By this, we can resolve many contradicting results that currently exist in the orthodontic community which are caused by lack of standardization in clinical studies. Accordingly, we were able to obtain a reliable and accurate measurement of the working range of both clinically valuable materials

Development and characterization of ultra-porous silica films made by the sol-gel method. Application to biosensing.

International audienceThe aim of this work is to demonstrate the sensing ability of reverse-symmetry waveguides to investigate adsorption of casein and build-up of poly-L-lysine mediated casein multilayers. A first part of this study is dedicated to the elaboration and characterization of ultra-porous thin films with very low refractive indices by an appropriate sol-gel method. This will form the basis of our planar optical sensors. Optical waveguide light mode spectroscopy is a real-time and sensitive method to study protein adsorption kinetics and lipid bilayers. We used it to test the obtained waveguides for in-situ monitoring of biomolecule adsorption. As a result, significant changes in the incoupling peak position were observed during the layer-by-layer adsorption. Finally, refractive index and thickness of the adsorbed layers were established

Specific intracellular signature of SARS-CoV-2 infection using confocal Raman microscopy

International audienceAbstract SARS-CoV-2 infection remains spread worldwide and requires a better understanding of virus-host interactions. Here, we analyzed biochemical modifications due to SARS-CoV-2 infection in cells by confocal Raman microscopy. Obtained results were compared with the infection with another RNA virus, the measles virus. Our results have demonstrated a virus-specific Raman molecular signature, reflecting intracellular modification during each infection. Advanced data analysis has been used to distinguish non-infected versus infected cells for two RNA viruses. Further, classification between non-infected and SARS-CoV-2 and measles virus-infected cells yielded an accuracy of 98.9 and 97.2 respectively, with a significant increase of the essential amino-acid tryptophan in SARS-CoV-2-infected cells. These results present proof of concept for the application of Raman spectroscopy to study virus-host interaction and to identify factors that contribute to the efficient SARS-CoV-2 infection and may thus provide novel insights on viral pathogenesis, targets of therapeutic intervention and development of new COVID-19 biomarkers