py8119 mouse breast cancer cells  (ATCC)

Journal: bioRxiv

Article Title: Single-cell profiling of synchronous multi-organ metastasis reveals a systemic CD74 + lipid-associated macrophage niche driving polymetastatic breast cancer

doi: 10.64898/2026.01.31.701004

Figure Lengend Snippet: a. qPCR analysis of Mif expression in VO-PyMT cells transduced with control or two independent Mif-targeting short hairpin RNAs (shRNA). Bar heights show means and error bars depict standard deviation of technical triplicates. Data was reproduced in 3 independent experiments. b. Immunoblot of Mif protein in shControl- or shMif-transduced VO-PyMT cells. B-Actin is shown as loading control. Data is representative of two independent experiments. c. MIF protein concentration in cell culture supernatants of shControl and shMif-transduced VO-PyMT cells as determined by enzyme linked immunosorbent assay (ELISA). Bar heights depict means and error bars show standard deviation of two technical replicates. d. Relative Mif expression levels in Py8119 cells harboring an shControl or shMif hairpin, as determined by qPCR. Error bars show standard deviations. Data is representative of 3 separate experiments. e. Western blot analysis of MIF protein in shControl or shMif Py8119 cells. B-Actin is shown as loading control. Data is representative of two independent experiments. f. Number of spheres per well quantified in in vitro spheroid cultures of VO-PyMT or Py8119 cells upon Mif knockdown. Sphere count was performed 7 days post seeding. Ns, not significant. Two-tailed Student’s t tests were used to estimate significance. g. Representative images of spheroids quantified in (f). Scale bars: 1 mm.

Article Snippet: Py8119 mouse breast cancer cells were purchased from ATCC (cat. no. CRL-3278), and cultured in F-12K medium supplemented with 2 mM L-Glutamine, 1,500 mg/L sodium bicarbonate, 10% vol/vol FBS, and P/S.

Techniques: Expressing, Transduction, Control, shRNA, Standard Deviation, Western Blot, Protein Concentration, Cell Culture, Enzyme-linked Immunosorbent Assay, In Vitro, Knockdown, Two Tailed Test

Journal: bioRxiv

Article Title: Single-cell profiling of synchronous multi-organ metastasis reveals a systemic CD74 + lipid-associated macrophage niche driving polymetastatic breast cancer

doi: 10.64898/2026.01.31.701004

Figure Lengend Snippet: a. Quantification of metastatic burden using ex vivo BLI of FVB mice injected VO-PyMT cells transduced with control vectors (shControl) and Mif-knockdown vectors (shMif 1; shMif 2). Data was acquired at day 12 post i.c. injections. shControl, n = 11; shMif (1), n = 11; shMif (2), n = 5. Data represents results from 2 independent experiments. In each experiment, raw photon flux values (p/s) from each organ were normalized to the average photon flux (p/s) values of the shControl group. Boxes depict 25th and 75th percentiles. Median is shown as horizontal lines. Whiskers depict data range. All data points are shown as dots. P values were determined using two-tailed Mann-Whitney tests by comparing shControl ( n = 11) versus the two shMif groups combined ( n = 16). Images show representative ex vivo BLI in brain, lung, liver or lower limb bones from each experimental group. b. Quantification of multi-organ metastatic burden ex vivo using BLI in albino C57/BL6 mice injected intracardially with Py8119 breast cancer cells transduced with control (shControl) and Mif-knockdown vector (shMif 2). Representative images are shown for each organ analyzed. Ex vivo metastatic burden was quantified 10 days after cancer cell injection. shControl, n = 14; shMif (2), n = 14. Data is representative of 3 independent experiments. Data in each organ was normalized to the average photon flux (p/s) of the shControl group in each experiment. Boxes boundaries define the interquartile ranges. Horizontal lines depict median values in each group. The whiskers show data range. The dots depict data points. P values were calculated by one-tailed Mann-Whitney t tests. c. Schematic showing pre-clinical model of MIF-CD74 targeting using 4-IPP, a small molecule inhibitor of MIF binding capacity. Multi-metastatic VO-PyMT cells were injected intracardially at day 0. Seeding and micrometastatic growth was allowed for 3 days, followed by i.p. administration of either a vehicle solution or 4-IPP. At day 12 post i.c. injection of VO-PyMT cells, metastatic colonization was analyzed ex vivo in brain, lung, liver and lower limb bones. d. Box plots showing quantification of metastatic burden of experiment as determined by ex vivo BLI in brain, lung, liver and bones of FVB mice injected intracardially with VO-PyMT cells and treated with the MIF inhibitor 4-IPP as described in (c). Vehicle group, n = 14; 4-IPP group, n = 13. For bone metastatic burden, right and left lower limb bones were analyzed separately. Data represents results from 2 independent experiments. In each experiment, photon flux (p/s) in each organ was normalized to the average photon flux (p/s) in the Vehicle control group. Boxes boundaries indicate 25 th and 75 th percentiles. Median is indicated by a horizontal line in boxes. Whiskers show data range. Data points are indicated as dots. P values were calculated using one-tailed Mann-Whitney t tests.

Article Snippet: Py8119 mouse breast cancer cells were purchased from ATCC (cat. no. CRL-3278), and cultured in F-12K medium supplemented with 2 mM L-Glutamine, 1,500 mg/L sodium bicarbonate, 10% vol/vol FBS, and P/S.

Techniques: Ex Vivo, Injection, Transduction, Control, Knockdown, Two Tailed Test, MANN-WHITNEY, Plasmid Preparation, One-tailed Test, Binding Assay