Journal: Scientific Reports

Article Title: YAP promotes the early development of temporomandibular joint bony ankylosis by regulating mesenchymal stem cell function

doi: 10.1038/s41598-024-63613-8

Figure Lengend Snippet: Transwell migration assays show that the migration ability of YAP-silenced MSCs is significantly reduced. ( A ) Transwell migration assay under a microscope (scale bar = 100 μm). ( B ) The graphical data represent the number of migrated cells per field (n = 4, ** P < 0.01, *** P < 0.001). MSCs mesenchymal stem cells.

Article Snippet: A 24‐well plate with Transwell chambers (Beyotime, China) with an 8.0 μm pore size was used for this experiment.

Techniques: Migration, Transwell Migration Assay, Microscopy

Journal: Nature

Article Title: Asparagine bioavailability governs metastasis in a model of breast cancer

doi: 10.1038/nature25465

Figure Lengend Snippet: a, Representative images of the lungs of mice that were intravenously injected with Asns-silenced or -expressing 4T1-T cells as described in Fig. 2a. b, Quantification of Matrigel invasion capacity for Asns-silenced and -expressing 4T1-T cells (n = 3 replicates per cell line). c, Quantification of mCherry-positive 4T1-T cells after roughly 50% of cells were infected with mCherry-expressing constructs harbouring shRNAs targeting Renilla luciferase and Asns. Cells were grown during the 24-h period that the Matrigel invasion assay described in Fig. 2b was being performed (n = 3 replicates per cell line). d, Violet cell-labelling intensity of Asns-silenced and -expressing 4T1-T cells, relative to the initial population. Cells were grown during the 24-h period that the Matrigel invasion assay described in Fig. 2b was being performed (n = 3 replicates per cell line). e, Free amino-acid quantification by HPLC for each amino acid in Asns-expressing and -silenced cells. Shown are the log-fold changes for each amino acid (n = 3 replicates per cell line). f, Quantification of mCherry-positive 4T1-T cells after roughly 50% of cells were infected with mCherry-expressing constructs harbouring shRNAs targeting Renilla luciferase and Asns. After infection, cells were grown in medium supplemented with l-asparagine or d-asparagine and mCherry percentages were measured at 48 and 96 h (n = 3 replicates per cell line). g, Quantification of Matrigel invasion for Asns-silenced and -expressing cells when assayed in medium supplemented with and without l-asparagine (n = 3 invasion chambers per cell line).

Article Snippet: Individual in vitro invasion assay The in vitro invasive capacity of cells was measured using six-well BioCoat Matrigel invasion plates.

Techniques: Injection, Expressing, Infection, Construct, Luciferase, Invasion Assay

Journal: Nature

Article Title: Asparagine bioavailability governs metastasis in a model of breast cancer

doi: 10.1038/nature25465

Figure Lengend Snippet: a, Volume measurements of tumours resulting from orthotopic injection of Asns-silenced and -expressing parental 4T1 cells (n = 10 mice per cell line, edges of the box are the 25th and 75th percentiles and error bars extend to the values q3 + w(q3 − q1) and q1 − w(q3 − q1), in which w is 1.5 and q1 and q3 are the 25th and 75th percentiles, which is also the case for b–g). b, Quantification of lung metastases corresponding to the tumours described in a (rank-sum P < 0.002). c, Volume measurements of tumours resulting from orthotopic injection of parental 4T1 cells with basal (Empty) or enforced expression of Asns (n = 10 mice per cell line). d, Quantification of lung metastases corresponding to the tumours described in c (rank-sum P < 5.0 × 10−5). e, Average diameters of the metastases of each mouse described in d (rank-sum P < 0.001). f, Volume measurements for tumours resulting from orthotopic injection of MDA-MB-231 cells with basal (Empty) or enforced expression of ASNS (n = 10 mice per cell line). g, Quantification of lung metastases corresponding to the tumours described in f (rank-sum P < 0.005). h, Quantification of Matrigel invasion for the MDA-MB-231- derived cell lines described in f (n = 3 invasion chambers per cell line). i, Representative images of the collection wells for the invasion assays described in h. See Source Data.

Article Snippet: Individual in vitro invasion assay The in vitro invasive capacity of cells was measured using six-well BioCoat Matrigel invasion plates.

Techniques: Biomarker Discovery, Injection, Expressing, Derivative Assay

Journal: Nature

Article Title: Asparagine bioavailability governs metastasis in a model of breast cancer

doi: 10.1038/nature25465

Figure Lengend Snippet: a, Quantification of parental 4T1 cell invasion rates, as measured by the Matrigel invasion assay, in culture medium supplemented with the indicated NEAAs (n = 5 invasion chambers, rank-sum P < 0.01). b, Representative H&E-stained lung sections from animals injected with Asns-silenced or -expressing 4T1-T cells. Animals were administered l-asparaginase or PBS (n = 10 mice per condition). c, Quantification of lung metastases in animals injected with Asns-silenced or -expressing 4T1-T cells. Animals were administered a diet with either 0%, 0.6%, or 4% asparagine content for the duration of the experiment (n = 10 mice per condition, ranksum P < 0.05 for Asns-silenced versus -expressing cells across all diets, for each cell line with 0% versus 4% diets, for shRenilla and shAsns-1 infected cells with 0% versus 0.6% diet, and for unsilenced cells with 0.6% versus 4% diet). d, Mass-spectrometric quantification of the asparagine levels in the mammary gland, blood serum, and lungs of animals administered l-asparaginase or PBS (relative abundance normalized by total metabolite peak area, n > 8 tissue sections per condition, rank-sum P < 0.005 for PBS versus l-asparaginase across all tissues, rank-sum P < 0.05 for mammary gland versus lung, and rank-sum P < 0.0005 for serum versus lung and serum versus mammary gland). See Source Data.

Article Snippet: Individual in vitro invasion assay The in vitro invasive capacity of cells was measured using six-well BioCoat Matrigel invasion plates.

Techniques: Invasion Assay, Staining, Injection, Expressing, Infection

Journal: Nature

Article Title: Asparagine bioavailability governs metastasis in a model of breast cancer

doi: 10.1038/nature25465

Figure Lengend Snippet: a, HPLC quantification of cellular free amino-acid percentages for parental 4T1 cells when the medium is supplemented with each of the NEAAs lacking in the DMEM culture medium (n = 3 replicates per cell line). b, Quantification of MDA-MB-231 Matrigel invasion rates under the same conditions as described in Fig. 3a (n = 5 invasion chambers per condition, rank-sum P < 0.001). c, HPLC quantification of cellular free aminoacid percentages for MDA-MB-231 cells when cultured in the medium conditions described in a (n = 3 replicates per cell line). d, Violet celllabelling intensity of parental 4T1 cells when grown in asparagine-lacking or -supplemented medium for the same period that the Matrigel invasion assay described in Fig. 3a was being performed (n = 3 replicates per cell line). e, Violet cell-labelling intensity of MDA-MB-231 cells when grown in asparagine-lacking or -supplemented medium for the same period that the Matrigel invasion assay described in b was being performed (n = 3 replicates per cell line).

Article Snippet: Individual in vitro invasion assay The in vitro invasive capacity of cells was measured using six-well BioCoat Matrigel invasion plates.

Techniques: Cell Culture, Invasion Assay