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11
المؤلفون: Irina Veith (10499821), Arianna Mencattini (3088794), Valentin Picant (10499824), Marco Serra (6058550), Marine Leclerc (10499827), Maria Colomba Comes (9539597), Fathia Mami-Chouaib (10499830), Jacques Camonis (49694), Stéphanie Descroix (728307), Hamasseh Shirvani (10499833), Fatima Mechta-Grigoriou (666210), Gérard Zalcman (310344), Maria Carla Parrini (2697100), Eugenio Martinelli (350161)
مصطلحات موضوعية: Biophysics, Cell Biology, Genetics, Pharmacology, Biotechnology, Immunology, Cancer, Hematology, Biological Sciences not elsewhere classified, Chemical Sciences not elsewhere classified, Information Systems not elsewhere classified, cancer cells, 3 D tumor ecosystems, cell co-culture videos, patio t emporal, doxorubicin chemotherapy drug, CTL, apoptotic cell death, cytotoxic cancer death, breast cancer chemo-resistance, biomimetic 3 D hydrogels, novel video analysis algorithms, drug resistance mechanisms, autologous tumor-infiltrating cytot., CAF, cancer death, cell culture techniques
الوصف: (AVI)
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12
المؤلفون: Irina Veith (10499821), Arianna Mencattini (3088794), Valentin Picant (10499824), Marco Serra (6058550), Marine Leclerc (10499827), Maria Colomba Comes (9539597), Fathia Mami-Chouaib (10499830), Jacques Camonis (49694), Stéphanie Descroix (728307), Hamasseh Shirvani (10499833), Fatima Mechta-Grigoriou (666210), Gérard Zalcman (310344), Maria Carla Parrini (2697100), Eugenio Martinelli (350161)
مصطلحات موضوعية: Biophysics, Cell Biology, Genetics, Pharmacology, Biotechnology, Immunology, Cancer, Hematology, Biological Sciences not elsewhere classified, Chemical Sciences not elsewhere classified, Information Systems not elsewhere classified, cancer cells, 3 D tumor ecosystems, cell co-culture videos, patio t emporal, doxorubicin chemotherapy drug, CTL, apoptotic cell death, cytotoxic cancer death, breast cancer chemo-resistance, biomimetic 3 D hydrogels, novel video analysis algorithms, drug resistance mechanisms, autologous tumor-infiltrating cytot., CAF, cancer death, cell culture techniques
الوصف: (AVI)
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13صورة
المؤلفون: Irina Veith (10499821), Arianna Mencattini (3088794), Valentin Picant (10499824), Marco Serra (6058550), Marine Leclerc (10499827), Maria Colomba Comes (9539597), Fathia Mami-Chouaib (10499830), Jacques Camonis (49694), Stéphanie Descroix (728307), Hamasseh Shirvani (10499833), Fatima Mechta-Grigoriou (666210), Gérard Zalcman (310344), Maria Carla Parrini (2697100), Eugenio Martinelli (350161)
مصطلحات موضوعية: Biophysics, Cell Biology, Genetics, Pharmacology, Biotechnology, Immunology, Cancer, Hematology, Biological Sciences not elsewhere classified, Chemical Sciences not elsewhere classified, Information Systems not elsewhere classified, cancer cells, 3 D tumor ecosystems, cell co-culture videos, patio t emporal, doxorubicin chemotherapy drug, CTL, apoptotic cell death, cytotoxic cancer death, breast cancer chemo-resistance, biomimetic 3 D hydrogels, novel video analysis algorithms, drug resistance mechanisms, autologous tumor-infiltrating cytot., CAF, cancer death, cell culture techniques
الوصف: From multi-channel videos of ToC co-cultures, the cancer cells (pre-stained in red) are localized and tracked in the red channel. The dying cancer cells (becoming green because of the caspase reporter) are identified in the green channel, after signal normalization and thresholding. The method monitors cancer cell deaths both in time and in space. The death signal is modeled to vanish during T time frames. Then, the spatiotemporal features of all death signals are integrated in a unique map, from which a parameter, named potential of death induction ( P death ), is computed over time (see Materials and Methods for details).
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14صورة
المؤلفون: Irina Veith (10499821), Arianna Mencattini (3088794), Valentin Picant (10499824), Marco Serra (6058550), Marine Leclerc (10499827), Maria Colomba Comes (9539597), Fathia Mami-Chouaib (10499830), Jacques Camonis (49694), Stéphanie Descroix (728307), Hamasseh Shirvani (10499833), Fatima Mechta-Grigoriou (666210), Gérard Zalcman (310344), Maria Carla Parrini (2697100), Eugenio Martinelli (350161)
مصطلحات موضوعية: Biophysics, Cell Biology, Genetics, Pharmacology, Biotechnology, Immunology, Cancer, Hematology, Biological Sciences not elsewhere classified, Chemical Sciences not elsewhere classified, Information Systems not elsewhere classified, cancer cells, 3 D tumor ecosystems, cell co-culture videos, patio t emporal, doxorubicin chemotherapy drug, CTL, apoptotic cell death, cytotoxic cancer death, breast cancer chemo-resistance, biomimetic 3 D hydrogels, novel video analysis algorithms, drug resistance mechanisms, autologous tumor-infiltrating cytot., CAF, cancer death, cell culture techniques
الوصف: A. Experimental design: breast cancer MDA-MB-231 cells are embedded in a collagen matrix in the central chamber of the chip; cells are live-imaged in transmission channel and fluorescence channels (red and green) every hour for 72 h, without or with doxorubicin (1 μM). B. Representative images of MDA-MB-231 cells after 1 h, 24 h and 48 h of culture on-chip, without drug (uppers panels) or with doxorubicin (lowers panels). Red arrows indicate living cells, whereas green arrows point at apoptotic cells. Scale bar, 100 μm. C. Time-course quantifications of the apoptosis rate, calculated in 10 h-time-intervals, showing the comparison between manual counts (black rounds) and automatic counts (red squares, T LAG = 10 h), without (left) or with (right) doxorubicin. The means +/- SEM of 3 measurements on 3 view fields from the same experiment were calculated automatically with STAMP every hour and manually every 10 hours. For statistical analysis, the measurements for each of the two conditions (manual and automated) were assembled regardless of the time variable. The non-parametric Mann-Whitney test was applied, since the data did not pass the Shapiro-Wilk normality test, and the difference resulted not significant.
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15صورة
المؤلفون: Irina Veith (10499821), Arianna Mencattini (3088794), Valentin Picant (10499824), Marco Serra (6058550), Marine Leclerc (10499827), Maria Colomba Comes (9539597), Fathia Mami-Chouaib (10499830), Jacques Camonis (49694), Stéphanie Descroix (728307), Hamasseh Shirvani (10499833), Fatima Mechta-Grigoriou (666210), Gérard Zalcman (310344), Maria Carla Parrini (2697100), Eugenio Martinelli (350161)
مصطلحات موضوعية: Biophysics, Cell Biology, Genetics, Pharmacology, Biotechnology, Immunology, Cancer, Hematology, Biological Sciences not elsewhere classified, Chemical Sciences not elsewhere classified, Information Systems not elsewhere classified, cancer cells, 3 D tumor ecosystems, cell co-culture videos, patio t emporal, doxorubicin chemotherapy drug, CTL, apoptotic cell death, cytotoxic cancer death, breast cancer chemo-resistance, biomimetic 3 D hydrogels, novel video analysis algorithms, drug resistance mechanisms, autologous tumor-infiltrating cytot., CAF, cancer death, cell culture techniques
الوصف: A. Experimental design: the lung cancer IGR-Pub cells are embedded in a collagen matrix in the central chamber of the chip, alone or together with autologous CTLs (P62 clone) at 1:1 effector to target cell (E:T) ratio; cells are live-imaged in transmission channel and fluorescence channels (red and green) every hour for 48 h. B. Representative images of IGR-Pub cells after 1 h, 24 h and 48 h of culture on-chip, alone (uppers panels) or with autologous T cells (lowers panels). Red arrows indicate living cells, whereas green arrows point at apoptotic cells. Blue arrows point at CTLs. Scale bar, 100 μm. C. Time-course quantifications of the apoptosis rate, calculated in 10 h-time-intervals, showing the comparison between manual counts (black rounds) and automatic counts (red squares, T LAG = 10 h), without (left) or with (right) T cells. The means +/- SEM of 3 measurements on 3 view fields from the same experiment were calculated automatically with STAMP every hour and manually every 10 hours. For statistical analysis, the measurements for each of the two conditions (manual and automated) were assembled regardless of the time variable. The non-parametric Mann-Whitney test was applied, since the data did not pass the Shapiro-Wilk normality test, and the difference resulted not significant.
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16صورة
المؤلفون: Irina Veith (10499821), Arianna Mencattini (3088794), Valentin Picant (10499824), Marco Serra (6058550), Marine Leclerc (10499827), Maria Colomba Comes (9539597), Fathia Mami-Chouaib (10499830), Jacques Camonis (49694), Stéphanie Descroix (728307), Hamasseh Shirvani (10499833), Fatima Mechta-Grigoriou (666210), Gérard Zalcman (310344), Maria Carla Parrini (2697100), Eugenio Martinelli (350161)
مصطلحات موضوعية: Biophysics, Cell Biology, Genetics, Pharmacology, Biotechnology, Immunology, Cancer, Hematology, Biological Sciences not elsewhere classified, Chemical Sciences not elsewhere classified, Information Systems not elsewhere classified, cancer cells, 3 D tumor ecosystems, cell co-culture videos, patio t emporal, doxorubicin chemotherapy drug, CTL, apoptotic cell death, cytotoxic cancer death, breast cancer chemo-resistance, biomimetic 3 D hydrogels, novel video analysis algorithms, drug resistance mechanisms, autologous tumor-infiltrating cytot., CAF, cancer death, cell culture techniques
الوصف: IGR-Pub lung cancer cells were co-cultured on-chip with autologous T cells at indicated E:T ratios. A. Time-course quantifications of the apoptosis rate, calculated in 4 h-time-intervals ( T LAG = 4 h), for a total duration of 48 h. B. Survival curves of cancer cells within the ToC at 1-hr time resolution. For each t i the % of surviving cells, calculated with respect to the initial number of living cells, is the average of 3 hours, centered on t i , to smoothen the fluctuations. The graphs show the means of 3 measurements on 3 view fields from the same experiment (+/- SEM). For statistical analysis, the measurements for each of the 4 conditions were assembled regardless of the time variable (n = 36 per condition). The different E:T ratio conditions were compared to the control condition of cancer cells only. In panel A, the data passed the Shapiro-Wilk normality test, therefore the unpaired t test was applied. In panel B, the data did not pass the Shapiro-Wilk normality test, therefore the non-parametric Mann-Whitney test was applied. **** indicates p<0.0001, ** indicates p<0.001, * indicates p<0.05 and ns indicates not significant.
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17صورة
المؤلفون: Irina Veith (10499821), Arianna Mencattini (3088794), Valentin Picant (10499824), Marco Serra (6058550), Marine Leclerc (10499827), Maria Colomba Comes (9539597), Fathia Mami-Chouaib (10499830), Jacques Camonis (49694), Stéphanie Descroix (728307), Hamasseh Shirvani (10499833), Fatima Mechta-Grigoriou (666210), Gérard Zalcman (310344), Maria Carla Parrini (2697100), Eugenio Martinelli (350161)
مصطلحات موضوعية: Biophysics, Cell Biology, Genetics, Pharmacology, Biotechnology, Immunology, Cancer, Hematology, Biological Sciences not elsewhere classified, Chemical Sciences not elsewhere classified, Information Systems not elsewhere classified, cancer cells, 3 D tumor ecosystems, cell co-culture videos, patio t emporal, doxorubicin chemotherapy drug, CTL, apoptotic cell death, cytotoxic cancer death, breast cancer chemo-resistance, biomimetic 3 D hydrogels, novel video analysis algorithms, drug resistance mechanisms, autologous tumor-infiltrating cytot., CAF, cancer death, cell culture techniques
الوصف: MDA-MB-231 cancer cells were co-cultured on-chip +/- cancer-associated fibroblasts (CAFs) and +/- doxorubicin (1μM) treatment. Final CAF:cancer ratio was 1:6. The graph shows the time-course quantifications of the apoptosis rate, calculated in 4 h-time-intervals ( T LAG = 4 h), for a total duration of 68 h. The graphs show the means of 3 measurements on 3 view fields from the same experiment (+/- SEM). The data illustrating the MDA-MB-231 without CAF conditions come from the same videos analyzed for Fig 2 but with different time-interval resolution. For statistical analysis, the measurements for each of the 4 conditions were assembled regardless of the time variable (n = 51 per condition). The different CAF/drug conditions were compared to the MDA-MB-231 control condition. The data passed the Shapiro-Wilk normality test, therefore the unpaired t test was applied. ** indicates p<0.001, * indicates p<0.05 and ns indicates not significant.
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18صورة
المؤلفون: Irina Veith (10499821), Arianna Mencattini (3088794), Valentin Picant (10499824), Marco Serra (6058550), Marine Leclerc (10499827), Maria Colomba Comes (9539597), Fathia Mami-Chouaib (10499830), Jacques Camonis (49694), Stéphanie Descroix (728307), Hamasseh Shirvani (10499833), Fatima Mechta-Grigoriou (666210), Gérard Zalcman (310344), Maria Carla Parrini (2697100), Eugenio Martinelli (350161)
مصطلحات موضوعية: Biophysics, Cell Biology, Genetics, Pharmacology, Biotechnology, Immunology, Cancer, Hematology, Biological Sciences not elsewhere classified, Chemical Sciences not elsewhere classified, Information Systems not elsewhere classified, cancer cells, 3 D tumor ecosystems, cell co-culture videos, patio t emporal, doxorubicin chemotherapy drug, CTL, apoptotic cell death, cytotoxic cancer death, breast cancer chemo-resistance, biomimetic 3 D hydrogels, novel video analysis algorithms, drug resistance mechanisms, autologous tumor-infiltrating cytot., CAF, cancer death, cell culture techniques
الوصف: A. Representative P death analysis on one video of breast cancer MDA-MB-231 cells cultured within ToC without (left, natural death) or with 1 μM doxorubicin (right, cytotoxic death). B. Representative P death analysis on one video of NSCLC IGR-Pub cells cultured within ToC alone (left, natural death) or together with autologous CTL (P62 clone) (right, cytotoxic death). C. Representative P death analysis on one video of breast cancer MDA-MB-231 cells co-cultured with CAFs within ToC, without (left) or with 1 μM doxorubicin (right). More video analyses on additional view fields are shown in S2 Fig .
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19صورة
المؤلفون: Irina Veith, Arianna Mencattini, Valentin Picant, Marco Serra, Marine Leclerc, Maria Colomba Comes, Fathia Mami-Chouaib, Jacques Camonis, Stéphanie Descroix, Hamasseh Shirvani, Fatima Mechta-Grigoriou, Gérard Zalcman, Maria Carla Parrini, Eugenio Martinelli
مصطلحات موضوعية: Biophysics, Cell Biology, Genetics, Pharmacology, Biotechnology, Immunology, Cancer, Hematology, Biological Sciences not elsewhere classified, Chemical Sciences not elsewhere classified, Information Systems not elsewhere classified, cancer cells, 3 D tumor ecosystems, cell co-culture videos, patio t emporal, doxorubicin chemotherapy drug, CTL, apoptotic cell death, cytotoxic cancer death, breast cancer chemo-resistance, biomimetic 3 D hydrogels, novel video analysis algorithms, drug resistance mechanisms, autologous tumor-infiltrating cytot, CAF, cancer death, cell culture techniques, socio, art
الوصف: A. Global view of the Leica DMi8 used to perform the live imaging experiments. The white arrow points at the heating black chamber, in which is maintained a temperature of 37°C. The temperature is set and maintained by a temperature controller (blue arrow). The CO 2 controller (black arrow) mixes CO 2 with atmospheric air. Through a tubing system, the air with the controlled CO 2 at 5%, after passing through a bottle half filled with water for humidification (red arrow), is injected in the microscope chamber. B. View of the chip placed in the microscope chamber. The white arrow indicates the lid of the chamber in which is injected the humidified air with CO 2 at 5%. The red arrow points at the chip to be imaged. Humidified sponges contribute to humidify the chamber and to minimize micro-evaporation phenomena (black arrows). C. Picture from the side of the chip filled with medium. At each ‘anchor’ point, three magnets (red arrows) are used to lift and attach the chip to the glass slide (black arrow). D. Top view of an empty chip with magnets. Two piles composed of three magnets (red arrows) are applied in the central part of the chip and glass slide. (PNG)
