c57bl 6 bl 6 mice  (Janvier Labs)

Journal: Nature Communications

Article Title: Differential ER stress responses during lactation and postpartum outcomes in mice depending on their mitochondrial genotype

doi: 10.1038/s41467-025-61666-5

Figure Lengend Snippet: A – H RNA seq analysis was performed on 3-4 months old, estrus cycle matched non-lactating (NL) virgin female mice and 3-4 months old lactating (L) BL/6 C57 or BL/6 NZB female mice on day 8, 14 and 21 of lactation. Venn diagram indicating the number of genes significantly (adjusted P < 0.05) upregulated or downregulated either in BL/6 C57 or BL/6 NZB female mice at day 8, 14, 21 of lactation compared to their non lactating (NL) controls ( n = 3 mice/group) ( A , C , E , G ). Graphs showing the trend of genes exclusively upregulated or downregulated at day 8, 14 and 21 in BL/6 C57 or BL/6 NZB mice ( B , D , F , H ). I Venn diagram indicating the number of genes significantly upregulated ( p < 0.05) in BL/6 C57 mice or BL/6 NZB mice at day 21 of lactation compared to their non lactating (NL) control ( n = 3 mice/group) (middle). The genes exclusively upregulated in BL/6 C57 or BL/6 NZB mice were subjected to pathway analysis using EnrichR and are represented in the graphs according to their combined score. All pathways shown are statistically significant. J Electron Microscopy was performed on the mammary glands of 3–4 months old BL/6 C57 or BL/6 NZB mice at day 10 and day 21 of lactation ( n = 5 mice/group). Pictures were taken from 8 to 10 different sections per mice. The scale bar represents 500 nm or 200 nm. K Graph of milk volumes per gland collected from either 3–4 months old BL/6 C57 or BL/6 NZB mice on day 8, 14 and 21 of lactation. Student’s unpaired t test, two-tailed, *** p < 0.001 at Day 8; *** p < 0.0008 at Day 14 and *** p < 0.0004 at Day 21; mean ± SD; n = 6 mice/group. L Graph showing average weight of mice per litter at day 8, 14, 21 of lactation. Student’s unpaired t test, two-tailed; mean ± SD; n = 6 litters/group. Source data are provided as a Source Data file. Mice used in all experiments were 2–3 months old at the beginning of the experiments.

Article Snippet: The BL/6 C57 (C57BL/6 J) and NZB NZB (NZB/OlaHsd) strains were purchased from ENVIGO, Netherlands .

Techniques: RNA Sequencing, Control, Electron Microscopy, Two Tailed Test

Journal: Nature Communications

Article Title: Differential ER stress responses during lactation and postpartum outcomes in mice depending on their mitochondrial genotype

doi: 10.1038/s41467-025-61666-5

Figure Lengend Snippet: A , B Western blot analysis of indicated cell cycle markers and p53 were performed in mammary glands from BL/6 C57 or BL/6 NZB 3-4 months old, estrus cycle matched non lactating (NL) virgin mice and 3–4 months old lactating (Day 21) female mice ( A ). Graphs of quantification of western blot analysis ( B ). Student’s unpaired t test, two-tailed, *** p < 0.0006, ** p < 0.0029 for cyclin E; **** p < 0.0001, *** p < 0.0002 for cyclin D1; *** p < 0.006, ** p < 0.0017 for cdk2; **** p < 0.001, ** p < 0.0010, ** p < 0.0045 for cdk4; *** p < 0.006, ** p < 0.0017 for cdk7; * p < 0.0111, ** p < 0.0018 for p53; mean ± SD; n = 9 mice/group. The representative blot shows 3 individual mice per group. In panels a and b, data are representative of two independent experiments. C Western Blot analysis of cell cycle markers in mammary organoids derived from 3-4 months old, estrus cycle matched BL/6 C57 or BL/6 NZB virgin female mice before (BI) or after induction (AI) with 1 µg/ml prolactin, 1 µg/ml hydrocortisone and 40 µg/ml oxytocin on the 6th day of organoid culture. D Graphs showing the quantification of western blot analysis in C . Student’s unpaired t test, two-tailed, **** p < 0.0001 for cyclin E; *** p < 0.007 for cyclin D1; *** p < 0.0007 for cdk2; ** p < 0.0014 for cdk4; ** p < 0.0015 for cdk7; ** p < 0.0018 for p53; mean ± SD; n = 9 mice/group. Number of experiments=3. E–J Mammary organoids from 3 to 4 months old, estrus cycle matched BL/6 C57 or BL/6 NZB female virgin mice induced with 1 µg/ml prolactin, 1 µg/ml hydrocortisone and 40 µg/ml oxytocin were fixed and stained for pRB, K-14, Actin and Dapi. ( E, G, I ) Representative image of pRB, K-14, Actin, Dapi and overlay. The scale bar represents 50 µm. n = 9 mice/group. Number of experiments=3. F , H , J Graph showing pRB stained region, K-14 surface area compared to total surface area, Actin-stained area which were analyzed using image J software. Student’s unpaired t test, one-tailed, * p < 0.0358 (F); Student’s unpaired t test, two-tailed, * p < 0.0245 (H); Student’s unpaired t test, one-tailed, * p < 0.0276 (J); mean ± SD; n = 9 mice/group. Number of experiments=3. K Primary mammary organoids isolated from 3 to 4 months old, estrus cycle matched, non-lactating BL/6 C57 or BL/6 NZB virgin female mice were grown for 6 days before induction with 1 µg/ml prolactin and 40 µg/ml oxytocin. Images show representative organoids at the indicated time. The scale bar represents 100 µm. n = 9 mice/group. Number of experiments=3. Source data are provided as a Source Data file. Mice used in all experiments were 2–3 months old at the beginning of the experiments.

Article Snippet: The BL/6 C57 (C57BL/6 J) and NZB NZB (NZB/OlaHsd) strains were purchased from ENVIGO, Netherlands .

Techniques: Western Blot, Two Tailed Test, Derivative Assay, Staining, Software, One-tailed Test, Isolation

Journal: Nature Communications

Article Title: Differential ER stress responses during lactation and postpartum outcomes in mice depending on their mitochondrial genotype

doi: 10.1038/s41467-025-61666-5

Figure Lengend Snippet: A Diagram of the experimental design in indicated female mice ( n = 10 mice/group). SMART images of mice were obtained from Servier. B , C 1×10 5 EO771 cells were injected in inguinal mammary glands of 3-4 months old BL/6 C57 or BL/6 NZB mice on day 17 of lactation. Graph depicts the mean tumor volume over time ( B ). Quantification of tumor volumes at day 32 ( C ). Quantification at each time point represents the average of 10 BL/6 C57 female mice relative to the average value of 10 BL/6 NZB female mice. Student’s unpaired t test, two-tailed, **** p < 0.0001; mean ± SD; n = 10 mice/group. D Diagram of the experimental design in indicated genotypes ( n = 10 mice/group). SMART images of mice were obtained from Servier . E , F H&E section of mammary glands from 5-6 months old, estrus cycle matched BL/6 C57 or BL/6 NZB female mice harvested after 45 days post completion of 21 days of lactation. Scale bar represents 200 µm. Graphs showing number of ducts or size of ducts analyzed using image J software. Student’s unpaired t test, two-tailed, * p < 0.05, **** p < 0.0001; mean ± SD; n = 10 mice/group. G Representative whole mount of mammary gland from 5-6 months old, estrus cycle matched BL/6 C57 or BL/6 NZB female mice harvested after 45 days post-partum after completion of 21 days of lactation. The scale bar represents 0.5 cm. Graph showing branching of ducts were analyzed using image J software. Student’s unpaired t test, one-tailed, **** p < 0.0001; mean ± SD; n = 10 mice/group. H Representative image of IHC of Ki67 in mammary glands derived from 5-6 months old, estrus cycle matched BL/6 C57 or BL/6 NZB mice harvested after 45 days post-partum after completion of 21 days of lactation. The scale bar represents 200 µm. Graph showing number of Ki67 stained cells were analyzed using image J software. Student’s unpaired t test, two-tailed, **** p < 0.0001; mean ± SD; n = 10 mice/group. I Schematic of patient’s selection and pathways identified in the human study comparing post-partum breast cancer (PPBC) to never pregnant breast cancer (NPBC). SMART images of women were obtained from Servier J Heatmap of the 14 transcription factors of the post-partum breast cancer (PPBC) regulon at day 21 of lactation in 3–4 months old BL/6 C57 and BL/6 NZB female mice. Source data are provided as a Source Data file. Mice used in all experiments were 2-3 months old at the beginning of the experiments.

Article Snippet: The BL/6 C57 (C57BL/6 J) and NZB NZB (NZB/OlaHsd) strains were purchased from ENVIGO, Netherlands .

Techniques: Injection, Two Tailed Test, Software, One-tailed Test, Derivative Assay, Staining, Selection

Journal: Nature Communications

Article Title: Differential ER stress responses during lactation and postpartum outcomes in mice depending on their mitochondrial genotype

doi: 10.1038/s41467-025-61666-5

Figure Lengend Snippet: Mammary glands from 3 to 4 months old, estrus cycle matched non-lactating (NL) female virgin mice and 3–4 months old day 21 lactating (D21) female mice were subjected to Single cell RNASeq ( n = 3 mice/group). A UMAP plot of Luminal Alveolar (Luminal AV), Hormone sensing (HS) and myoepithelial cell clusters identified by expression of characteristic markers. B Relative proportion of epithelial cell types per genotype in non-lactating (NL) and day 21 lactating (D21) female mice. C Hierarchical clustering heatmaps of differential gene expression of luminal HS cells in non-lactating (NL) BL/6 C57 and BL/6 NZB . D Hierarchical clustering heatmaps of differential gene expression of luminal HS cells in lactating BL/6 C57 and BL/6 NZB females. E Number of PPBC-like cells (defined by high cell cycle score and high regulon positivity, see text) in non-lactating (NL) and lactating day 21 (D21) BL/6 C57 and BL/6 NZB females. Student’s unpaired t test, one-tailed, * p < 0.0253 NL vs D21 in BL/6 NZB females; Student’s unpaired t test, two-tailed, * p < 0.0458 D21 of BL/6 C57 vs D21 of BL/6 NZB females; mean ± SD; n = 3 mice/group. F Representative image of PPBC-like positive cell cluster. G Number of luminal HS cells expressing p53 in non-lactating (NL) and lactating day 21 (D21) BL/6 C57 and BL/6 NZB females. Ordinary one-way ANOVA, ** p < 0.0014 NL vs D21 in BL/6 NZB females; * p < 0.0111 D21 of BL/6 C57 vs D21 of BL/6 NZB females; mean ± SD; n = 3 mice/group H List of the top 40 differentially up-regulated genes in luminal HS cells in BL/6 C57 relative to BL/6 NZB female mice at day 21 of lactation. Bold indicated p53 transcriptional targets. I Pathways related to the expression of the 40 genes shown in H. Databases where these pathways were identified are color coded. Source data are provided as a Source Data file. Mice used in all experiments were 2-3 months old at the beginning of the experiments.

Article Snippet: The BL/6 C57 (C57BL/6 J) and NZB NZB (NZB/OlaHsd) strains were purchased from ENVIGO, Netherlands .

Techniques: Expressing, Gene Expression, One-tailed Test, Two Tailed Test

Journal: Nature Communications

Article Title: Differential ER stress responses during lactation and postpartum outcomes in mice depending on their mitochondrial genotype

doi: 10.1038/s41467-025-61666-5

Figure Lengend Snippet: Western blot analysis of mammary glands from 3-4 months old, estrus cycle matched virgin non-lactating (NL) and 3-4 months old lactating (L) at day 8, 14 and 21 in BL/6 C57 (Left) or BL/6 NZB (right) female mice ( n = 9 mice/group) for the indicated markers of ER-stress ( A ),mitochondrial-stress ( C ) and apoptosis ( E ). Representative westerns show the result of 3 (out of 9) individual BL/6 C57 or BL/6 NZB mice at each time point compared to their non lactating counterpart. Quantification of ER-stress markers ( B ), mitochondrial-stress markers ( D ) and apoptosis markers ( F ). Student’s unpaired t test, two-tailed, * p < 0.05, ** p < 0.01, *** p < 0.001; mean ± SD. A detailed p value has been provided in the Source data as a Source Data file. Mice used in all experiments were 2-3 months old at the beginning of the experiments. The representative blot shows 3 individual mice per group. In panels a to f, data are representative of two independent experiments.

Article Snippet: The BL/6 C57 (C57BL/6 J) and NZB NZB (NZB/OlaHsd) strains were purchased from ENVIGO, Netherlands .

Techniques: Western Blot, Two Tailed Test

Journal: Nature Communications

Article Title: Differential ER stress responses during lactation and postpartum outcomes in mice depending on their mitochondrial genotype

doi: 10.1038/s41467-025-61666-5

Figure Lengend Snippet: A Organoids of mammary glands from 3-4 months old, estrus cycle matched, non-lactating BL/6 NZB virgin female mice were treated with 20 µg/ml δ-tocotrienol starting from the 4 th day followed by induction with 1 µg/ml prolactin, 1 µg/ml hydrocortisone and 40 µg/ml oxytocin on the 6 th day. 22 hr post induction, the organoids were subjected to western blot analysis for the indicated markers. Graphs show the quantification of western blot analysis. Student’s unpaired t test, two-tailed, ** p < 0.01; ** p < 0.01, *** p < 0.001, **** p < 0.0001; mean ± SD; n = 9 mice/group. Number of experiments=3. The detailed p values are provided in the source data file. B Organoids of mammary glands from 3-4 months old, estrus cycle matched, non-lactating BL/6 C57 virgin female mice were treated with 30 µM Salubrinal from the 4 th day, then induced with 1 µg/ml prolactin, 1 µg/ml hydrocortisone and 40 µg/ml oxytocin on the 6 th day. 22 hr post induction, the organoids were subjected to western blot analysis for the indicated markers. Graphs show the quantification of western blot analysis. Student’s unpaired t test, two-tailed,* ** p < 0.001, **** p < 0.0001; mean ± SD; n = 9 mice/group. Number of experiments=3. A detailed p value is provided in the source data file ( C – H ), Mammary glands from 5-6 months old, estrus cycle matched BL/6 NZB mice collected 45 days post completion of 21 days of lactation treated with either vehicle (olive oil) or δ-tocotrienol (200 mg/kg body weight in olive oil, twice daily) by oral gavage starting from 10 th day of lactation for 4 weeks ( n = 10 mice/Group). C , Schematic representation of experimental design. SMART images of mice were obtained from Servier . D , E H&E section of mammary glands from BL/6 NZB mice without (Vehicle) or with δ-tocotrienol. Scale bar represents 200 µm. Images were taken with a 20X ( D ) and 40X ( E ) objectives respectively. Graphs showing number of ducts or size of ducts analyzed using image J software. Student’s t test, two-tailed, **** p < 0.0001; mean ± SD; n = 10 mice/group. F Representative whole mount of mammary gland from above mentioned mice. The scale bar represents 0.5 cm. Graph showing thickness of ducts were analyzed using image J software. Student’s t test, two-tailed, **** p < 0.0001; mean ± SD; n = 10 mice/group. G Representative image of IHC of Ki67 in mammary glands derived from above mentioned mice. The scale bar represents 200 µm. Graph showing number of Ki67 stained cells were analyzed using image J software. Student’s t test, two-tailed, **** p < 0.0001; mean ± SD; n = 10 mice/group. H Western blot analysis of ER Stress markers and cell cycle markers were performed in mammary glands of above-mentioned BL/6 NZB mice treated without (Vehicle) or with δ-tocotrienol. Graphs of quantification of western blot analysis. Student’s t test, two-tailed, *** p < 0.0006, **** p < 0.0001; mean ± SD; n = 10 mice/group. The representative blot shows 5 individual mice per group. In panel h, data are representative of two independent experiments. I Diagram of the experimental design in each group, n = 10 mice/group. SMART images of mice were obtained from Servier. J , K BL/6 NZB mice were treated with vehicle (olive oil) or δ- tocotrienol (200 mg/kg body weight in olive oil, twice daily) by oral gavage starting from 10 th day of lactation and 1×10 5 EO771 cells were injected in inguinal mammary glands of mice on day 17 of lactation. The mice were harvested 32 days post-partum after completion of 21 days of lactation. J Graph of mean tumor volumes per mice at the given time points. Quantification at each time point represents the average of 10 BL/6 NZB mice treated with vehicle relative to the average value of 10 BL/6 NZB mice treated with δ-tocotrienol. K Graph of tumor volumes per gland on the day of sacrifice (day 32 post implantation). Student’s t test, two-tailed, **** p < 0.0001; mean ± SD; n = 10 mice/group. Source data are provided as a Source Data file. Mice used in all experiments were 2-3 months old at the beginning of the experiments.

Article Snippet: The BL/6 C57 (C57BL/6 J) and NZB NZB (NZB/OlaHsd) strains were purchased from ENVIGO, Netherlands .

Techniques: Western Blot, Two Tailed Test, Software, Derivative Assay, Staining, Injection