#METABOLOMICS WORKBENCH sbeh91Jema_20250327_162802 DATATRACK_ID:5800 STUDY_ID:ST003855 ANALYSIS_ID:AN006333 PROJECT_ID:PR002412 VERSION 1 CREATED_ON April 9, 2025, 9:04 pm #PROJECT PR:PROJECT_TITLE Mammary gland metabolism and its relevance to the fetoplacental expression of PR:PROJECT_TITLE cytokine signaling in Caveolin-1 null mice PR:PROJECT_SUMMARY Mice lacking Caveolin-1 (Cav1), a major protein of the lipid raft of plasma PR:PROJECT_SUMMARY membrane, show dysregulated cellular proliferation of mammary gland and an PR:PROJECT_SUMMARY abnormal fetoplacental communication during pregnancy. The aim of this study is PR:PROJECT_SUMMARY to better understand the functional links of mammary gland metabolism with gene PR:PROJECT_SUMMARY expression of the placenta and fetus. Untargeted metabolomics analysis was PR:PROJECT_SUMMARY performed to examine changes in mammary gland metabolism due to the absence of PR:PROJECT_SUMMARY Cav1. The metabolomics analysis detected a total of 141 metabolites. We PR:PROJECT_SUMMARY identified 81 metabolites that showed differential rank order in the level of PR:PROJECT_SUMMARY expression in the mammary glands of Cav1-null compared to control mice. PR:PROJECT_SUMMARY Differential metabolomics analysis identified 11 metabolites that were PR:PROJECT_SUMMARY significantly changed in the mammary gland due to the absence of Cav1. PR:PROJECT_SUMMARY Integrative metabolomics and transcriptomics analyses were applied to untangle PR:PROJECT_SUMMARY functional links of metabolic pathways of the mammary gland with the gene PR:PROJECT_SUMMARY expression changes of the placenta and fetus. The findings of this study show PR:PROJECT_SUMMARY that metabolism and gene expression of the mammary gland are significantly PR:PROJECT_SUMMARY impacted due to the loss of Cav1. Genes associated with specific metabolic and PR:PROJECT_SUMMARY signaling pathways show coordinated expression changed in the placenta, mammary PR:PROJECT_SUMMARY gland and fetal brain in Cav1-null mice. The cytokine signaling pathway emerges PR:PROJECT_SUMMARY as a key player of the molecular crosstalk among the mammary gland, placenta and PR:PROJECT_SUMMARY fetal brain. By interrogating the single-nuclei gene expression data of placenta PR:PROJECT_SUMMARY and fetal brain previously generated from Cav1-null mice, the study further PR:PROJECT_SUMMARY reveals that these metabolic and signaling genes are differentially regulated in PR:PROJECT_SUMMARY specific cell types of the placenta and fetal brain. The findings of this study PR:PROJECT_SUMMARY expand our understanding about the role of mammary gland metabolism in the PR:PROJECT_SUMMARY regulation of fetoplacental communication in mammalian pregnancy. PR:INSTITUTE Universiy of Missouri, Columbia PR:LAST_NAME Behura PR:FIRST_NAME Susanta PR:ADDRESS 920 East Campus, Columbia, Missouri, 65211, USA PR:EMAIL behuras@missouri.edu PR:PHONE 573-882-1722 #STUDY ST:STUDY_TITLE Mammary gland metabolism and its relevance to the fetoplacental expression of ST:STUDY_TITLE cytokine signaling in Caveolin-1 null mice ST:STUDY_SUMMARY Mice lacking Caveolin-1 (Cav1), a major protein of the lipid raft of plasma ST:STUDY_SUMMARY membrane, show dysregulated cellular proliferation of mammary gland and an ST:STUDY_SUMMARY abnormal fetoplacental communication during pregnancy. The aim of this study is ST:STUDY_SUMMARY to better understand the functional links of mammary gland metabolism with gene ST:STUDY_SUMMARY expression of the placenta and fetus. Untargeted metabolomics analysis was ST:STUDY_SUMMARY performed to examine changes in mammary gland metabolism due to the absence of ST:STUDY_SUMMARY Cav1. The metabolomics analysis detected a total of 141 metabolites. We ST:STUDY_SUMMARY identified 81 metabolites that showed differential rank order in the level of ST:STUDY_SUMMARY expression in the mammary glands of Cav1-null compared to control mice. ST:STUDY_SUMMARY Differential metabolomics analysis identified 11 metabolites that were ST:STUDY_SUMMARY significantly changed in the mammary gland due to the absence of Cav1. ST:STUDY_SUMMARY Integrative metabolomics and transcriptomics analyses were applied to untangle ST:STUDY_SUMMARY functional links of metabolic pathways of the mammary gland with the gene ST:STUDY_SUMMARY expression changes of the placenta and fetus. The findings of this study show ST:STUDY_SUMMARY that metabolism and gene expression of the mammary gland are significantly ST:STUDY_SUMMARY impacted due to the loss of Cav1. Genes associated with specific metabolic and ST:STUDY_SUMMARY signaling pathways show coordinated expression changed in the placenta, mammary ST:STUDY_SUMMARY gland and fetal brain in Cav1-null mice. The cytokine signaling pathway emerges ST:STUDY_SUMMARY as a key player of the molecular crosstalk among the mammary gland, placenta and ST:STUDY_SUMMARY fetal brain. By interrogating the single-nuclei gene expression data of placenta ST:STUDY_SUMMARY and fetal brain previously generated from Cav1-null mice, the study further ST:STUDY_SUMMARY reveals that these metabolic and signaling genes are differentially regulated in ST:STUDY_SUMMARY specific cell types of the placenta and fetal brain. The findings of this study ST:STUDY_SUMMARY expand our understanding about the role of mammary gland metabolism in the ST:STUDY_SUMMARY regulation of fetoplacental communication in mammalian pregnancy. ST:INSTITUTE Universiy of Missouri, Columbia ST:LAST_NAME Behura ST:FIRST_NAME Susanta ST:ADDRESS 920 East Campus, Columbia, Missouri, 65211, USA ST:EMAIL behuras@missouri.edu ST:PHONE 573-882-1722 ST:NUM_GROUPS 2 ST:TOTAL_SUBJECTS 6 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN B6.Cg-Cav1tm1Mls/J (Cav1-null) mice; C57BL/6J (Control) mice #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data SUBJECT_SAMPLE_FACTORS - BLMAM-1 Sample source:Mammary gland | Genotype:C57BL6/J | Sample type:Control RAW_FILE_NAME(Raw file name)=BLMAM-1.CDF SUBJECT_SAMPLE_FACTORS - BLMAM-2 Sample source:Mammary gland | Genotype:C57BL6/J | Sample type:Control RAW_FILE_NAME(Raw file name)=BLMAM-2.CDF SUBJECT_SAMPLE_FACTORS - BLMAM-3 Sample source:Mammary gland | Genotype:C57BL6/J | Sample type:Control RAW_FILE_NAME(Raw file name)=BLMAM-3.CDF SUBJECT_SAMPLE_FACTORS - CVMAM-1 Sample source:Mammary gland | Genotype:B6.Cg-Cav1tm1Mls/J | Sample type:Cav1-null RAW_FILE_NAME(Raw file name)=CVMAMA-1.CDF SUBJECT_SAMPLE_FACTORS - CVMAM-2 Sample source:Mammary gland | Genotype:B6.Cg-Cav1tm1Mls/J | Sample type:Cav1-null RAW_FILE_NAME(Raw file name)=CVMAMA-2.CDF SUBJECT_SAMPLE_FACTORS - CVMAM-3 Sample source:Mammary gland | Genotype:B6.Cg-Cav1tm1Mls/J | Sample type:Cav1-null RAW_FILE_NAME(Raw file name)=CVMAMA-3.CDF #COLLECTION CO:COLLECTION_SUMMARY The B6.Cg-Cav1tm1Mls/J (Cav1-null) and control (C57BL/6J) mice were purchased CO:COLLECTION_SUMMARY from the Jackson Laboratory (stock numbers: 000664 and 007083 respectively). CO:COLLECTION_SUMMARY Adult male and female mice (7 weeks old) were paired in cages to induce CO:COLLECTION_SUMMARY pregnancy. The females were checked for vaginal plug. The start of pregnancy CO:COLLECTION_SUMMARY (day 1) was considered when a vaginal plug was observed. The control and CO:COLLECTION_SUMMARY Cav1-null pregnant mice were euthanized in triplicates on gestation day 15, and CO:COLLECTION_SUMMARY the mammary gland samples were dissected from mice of both the groups (n=3 CO:COLLECTION_SUMMARY each). CO:SAMPLE_TYPE mammary gland #TREATMENT TR:TREATMENT_SUMMARY No treatment. Genotypes B6.Cg-Cav1tm1Mls/J (Cav1-null mice) C57BL/6J (Control TR:TREATMENT_SUMMARY mice) were compared. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY The samples were processed as per the attached protocol of GC-MS used by the SP:SAMPLEPREP_SUMMARY University of Missouri Metabolomics Center that provided the metabolomics SP:SAMPLEPREP_SUMMARY service. SP:SAMPLEPREP_PROTOCOL_FILENAME Protocol_CoreLab.pdf #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Agilent 6890 CH:COLUMN_NAME J&W Scientific DB-5MS (60m x 0.25mm, 0.25mm) CH:SOLVENT_A - CH:SOLVENT_B - CH:FLOW_GRADIENT - CH:FLOW_RATE 1.0 mL/min CH:COLUMN_TEMPERATURE Temperature program: 80C for 2 min, then ramped at 5C min-1 to 315C and held for CH:COLUMN_TEMPERATURE 12 min CH:METHODS_FILENAME Protocol_CoreLab.pdf #ANALYSIS AN:ANALYSIS_TYPE MS AN:ANALYSIS_PROTOCOL_FILE Protocol_CoreLab.pdf #MS MS:INSTRUMENT_NAME Agilent 5973 MS:INSTRUMENT_TYPE Single quadrupole MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS The spectral analysis was performed by the AMDIS (Automate Mass-spectral MS:MS_COMMENTS Deconvolution and Identification System), and metabolites were identified using MS:MS_COMMENTS a commercial NIST17 mass spectral library. The abundance of the metabolites was MS:MS_COMMENTS determined by the Metabolomics Ion-Based Data Extraction Algorithm (MET-IDEA). MS:MS_RESULTS_FILE ST003855_AN006333_Results.txt UNITS:uM Has m/z:Yes Has RT:Yes RT units:Minutes #END