#METABOLOMICS WORKBENCH sumnerlw_20200127_121106 DATATRACK_ID:1910 STUDY_ID:ST001310 ANALYSIS_ID:AN002181 PROJECT_ID:PR000891 VERSION 1 CREATED_ON January 30, 2020, 4:02 pm #PROJECT PR:PROJECT_TITLE Bisphenol A and bisphenol S disruptions of the mouse placenta and potential PR:PROJECT_TITLE effects on the placenta–brain axis PR:PROJECT_SUMMARY Placental trophoblast cells are potentially at risk from circulating PR:PROJECT_SUMMARY endocrine-disrupting chemicals, such as bisphenol A (BPA). To understand how BPA PR:PROJECT_SUMMARY and the reputedly more inert bisphenol S (BPS) affect the placenta, C57BL6J PR:PROJECT_SUMMARY mouse dams were fed 200 μg/kg body weight BPA or BPS daily for 2 wk and then PR:PROJECT_SUMMARY bred. They continued to receive these chemicals until embryonic day 12.5, PR:PROJECT_SUMMARY whereupon placental samples were collected and compared with unexposed controls. PR:PROJECT_SUMMARY BPA and BPS altered the expression of an identical set of 13 genes. Both PR:PROJECT_SUMMARY exposures led to a decrease in the area occupied by spongiotrophoblast relative PR:PROJECT_SUMMARY to multinucleated giant cells (GCs) within the junctional zone, markedly reduced PR:PROJECT_SUMMARY placental serotonin (5-HT) concentrations, and lowered 5-HT GC immunoreactivity. PR:PROJECT_SUMMARY Concentrations of dopamine and 5-hydroxyindoleacetic acid, the main metabolite PR:PROJECT_SUMMARY of serotonin, were increased. GC dopamine immunoreactivity was increased in BPA- PR:PROJECT_SUMMARY and BPS-exposed placentas. A strong positive correlation between 5-HT+ GCs and PR:PROJECT_SUMMARY reductions in spongiotrophoblast to GC area suggests that this neurotransmitter PR:PROJECT_SUMMARY is essential for maintaining cells within the junctional zone. In contrast, an PR:PROJECT_SUMMARY inverse correlation existed between dopamine+ GCs and reductions PR:PROJECT_SUMMARY spongiotrophoblast to GC area. These outcomes lead to the following conclusions. PR:PROJECT_SUMMARY First, BPS exposure causes almost identical placental effects as BPA. Second, a PR:PROJECT_SUMMARY major target of BPA/BPS is either spongiotrophoblast or GC within the junctional PR:PROJECT_SUMMARY zone. Third, imbalances in neurotransmitter-positive GC and an observed decrease PR:PROJECT_SUMMARY in docosahexaenoic acid and estradiol, also occurring in response to BPA/BPS PR:PROJECT_SUMMARY exposure, likely affect the placental–brain axis of the developing mouse PR:PROJECT_SUMMARY fetus. PR:INSTITUTE University of Missouri PR:DEPARTMENT Life Sciences Center PR:LABORATORY Rosenfeld Lab PR:LAST_NAME Rosenfeld PR:FIRST_NAME Cheryl PR:ADDRESS Bond Life Sciences Center, 1201 Rollins St., Columbia, MO PR:EMAIL RosenfeldC@missouri.edu PR:PHONE 573-882-5132 PR:FUNDING_SOURCE NIEHS PR:PUBLICATIONS Mao et al, Proceedings National Academy of Science, USA, PR:CONTRIBUTORS Jiude Mao, Ashish Jain, Nancy D. Denslow, Mohammad-Zaman Nouri, Sixue Chen, PR:CONTRIBUTORS Tingting Wang, Ning Zhu, Jin Koh, Saurav J. Sarma, Barbara W. Sumner, Zhentian PR:CONTRIBUTORS Lei, Lloyd W. Sumner, Nathan J. Bivens, R. Michael Roberts, Geetu Tuteja, and PR:CONTRIBUTORS Cheryl S. Rosenfeld #STUDY ST:STUDY_TITLE C57 midgestation placental metabolomics analysis ST:STUDY_TYPE BPA, BPS exposure on placenta metabolite profile ST:STUDY_SUMMARY Placental trophoblast cells are potentially at risk from circulating ST:STUDY_SUMMARY endocrine-disrupting chemicals, such as bisphenol A (BPA). To understand how BPA ST:STUDY_SUMMARY and the reputedly more inert bisphenol S (BPS) affect the placenta, C57BL6J ST:STUDY_SUMMARY mouse dams were fed 200 μg/kg body weight BPA or BPS daily for 2 wk and then ST:STUDY_SUMMARY bred. They continued to receive these chemicals until embryonic day 12.5, ST:STUDY_SUMMARY whereupon placental samples were collected and compared with unexposed controls. ST:STUDY_SUMMARY BPA and BPS altered the expression of an identical set of 13 genes. Both ST:STUDY_SUMMARY exposures led to a decrease in the area occupied by spongiotrophoblast relative ST:STUDY_SUMMARY to multinucleated giant cells (GCs) within the junctional zone, markedly reduced ST:STUDY_SUMMARY placental serotonin (5-HT) concentrations, and lowered 5-HT GC immunoreactivity. ST:STUDY_SUMMARY Concentrations of dopamine and 5-hydroxyindoleacetic acid, the main metabolite ST:STUDY_SUMMARY of serotonin, were increased. GC dopamine immunoreactivity was increased in BPA- ST:STUDY_SUMMARY and BPS-exposed placentas. A strong positive correlation between 5-HT+ GCs and ST:STUDY_SUMMARY reductions in spongiotrophoblast to GC area suggests that this neurotransmitter ST:STUDY_SUMMARY is essential for maintaining cells within the junctional zone. In contrast, an ST:STUDY_SUMMARY inverse correlation existed between dopamine+ GCs and reductions ST:STUDY_SUMMARY spongiotrophoblast to GC area. These outcomes lead to the following conclusions. ST:STUDY_SUMMARY First, BPS exposure causes almost identical placental effects as BPA. Second, a ST:STUDY_SUMMARY major target of BPA/BPS is either spongiotrophoblast or GC within the junctional ST:STUDY_SUMMARY zone. Third, imbalances in neurotransmitter-positive GC and an observed decrease ST:STUDY_SUMMARY in docosahexaenoic acid and estradiol, also occurring in response to BPA/BPS ST:STUDY_SUMMARY exposure, likely affect the placental–brain axis of the developing mouse ST:STUDY_SUMMARY fetus. ST:INSTITUTE University of Missouri ST:DEPARTMENT Life Sciences Center ST:LABORATORY Univ. of Missouri Metabolomics Center ST:LAST_NAME Sumner ST:FIRST_NAME Lloyd ST:ADDRESS 1201 Rollins Street Columbia, Missouri 65211-7310 ST:EMAIL sumnerlw@missouri.edu ST:PHONE 573-882-5486 ST:NUM_GROUPS 3 treatment X 2 sex = 6 ST:TOTAL_SUBJECTS 40 ST:PUBLICATIONS Mao et al, Proceedings National Academy of Science, USA, 2020 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:GENOTYPE_STRAIN C57BL6J SU:AGE_OR_AGE_RANGE Embryonic day 12.5 SU:GENDER Male and female SU:ANIMAL_ANIMAL_SUPPLIER Jackson Labs SU:ANIMAL_HOUSING polypropylene cages #FACTORS #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 - 3017-BPA-F Exposure:BPA | Gender:female RAW_FILE_NAME=3017-BPA-F.D SUBJECT_SAMPLE_FACTORS - 3015-BPA-F Exposure:BPA | Gender:female RAW_FILE_NAME=3015-BPA-F.D SUBJECT_SAMPLE_FACTORS - 2027-BPA-F Exposure:BPA | Gender:female RAW_FILE_NAME=2027-BPA-F.D SUBJECT_SAMPLE_FACTORS - 2015-BPA-F Exposure:BPA | Gender:female RAW_FILE_NAME=2015-BPA-F.D SUBJECT_SAMPLE_FACTORS - 2021-BPA-F Exposure:BPA | Gender:female RAW_FILE_NAME=2021-BPA-F.D SUBJECT_SAMPLE_FACTORS - 3018-BPA-F Exposure:BPA | Gender:female RAW_FILE_NAME=3018-BPA-F.D SUBJECT_SAMPLE_FACTORS - 3017-BPA-M Exposure:BPA | Gender:male RAW_FILE_NAME=3017-BPA-M.D SUBJECT_SAMPLE_FACTORS - 3015-BPA-M Exposure:BPA | Gender:male RAW_FILE_NAME=3015-BPA-M.D SUBJECT_SAMPLE_FACTORS - 2027-BPA-M Exposure:BPA | Gender:male RAW_FILE_NAME=2027-BPA-M.D SUBJECT_SAMPLE_FACTORS - 2009-BPA-M Exposure:BPA | Gender:male RAW_FILE_NAME=2009-BPA-M.D SUBJECT_SAMPLE_FACTORS - 3018-BPA-M Exposure:BPA | Gender:male RAW_FILE_NAME=3018-BPA-M.D SUBJECT_SAMPLE_FACTORS - 2015-BPA-M Exposure:BPA | Gender:male RAW_FILE_NAME=2015-BPA-M.D SUBJECT_SAMPLE_FACTORS - 2021-BPA-M Exposure:BPA | Gender:male RAW_FILE_NAME=2021-BPA-M.D SUBJECT_SAMPLE_FACTORS - 2013-BPS-F Exposure:BPS | Gender:female RAW_FILE_NAME=2013-BPS-F.D SUBJECT_SAMPLE_FACTORS - 2025-BPS-F Exposure:BPS | Gender:female RAW_FILE_NAME=2025-BPS-F.D SUBJECT_SAMPLE_FACTORS - 2001-BPS-F Exposure:BPS | Gender:female RAW_FILE_NAME=2001-BPS-F.D SUBJECT_SAMPLE_FACTORS - 3004-BPS-F Exposure:BPS | Gender:female RAW_FILE_NAME=3004-BPS-F.D SUBJECT_SAMPLE_FACTORS - 2019-BPS-F Exposure:BPS | Gender:female RAW_FILE_NAME=2019-BPS-F.D SUBJECT_SAMPLE_FACTORS - 2007-BPS-F Exposure:BPS | Gender:female RAW_FILE_NAME=2007-BPS-F.D SUBJECT_SAMPLE_FACTORS - 2009-BPS-F Exposure:BPS | Gender:female RAW_FILE_NAME=2009-BPS-F.D SUBJECT_SAMPLE_FACTORS - 3002-BPS-F Exposure:BPS | Gender:female RAW_FILE_NAME=3002-BPS-F.D SUBJECT_SAMPLE_FACTORS - 2031-BPS-F Exposure:BPS | Gender:female RAW_FILE_NAME=2031-BPS-F.D SUBJECT_SAMPLE_FACTORS - 2025-BPS-M Exposure:BPS | Gender:male RAW_FILE_NAME=2025-BPS-M.D SUBJECT_SAMPLE_FACTORS - 2001-BPS-M Exposure:BPS | Gender:male RAW_FILE_NAME=2001-BPS-M.D SUBJECT_SAMPLE_FACTORS - 3004-BPS-M Exposure:BPS | Gender:male RAW_FILE_NAME=3004-BPS-M.D SUBJECT_SAMPLE_FACTORS - 2019-BPS-M Exposure:BPS | Gender:male RAW_FILE_NAME=2019-BPS-M.D SUBJECT_SAMPLE_FACTORS - 2013-BPS-M Exposure:BPS | Gender:male RAW_FILE_NAME=2013-BPS-M.D SUBJECT_SAMPLE_FACTORS - 3002-BPS-M Exposure:BPS | Gender:male RAW_FILE_NAME=3002-BPS-M.D SUBJECT_SAMPLE_FACTORS - 2007-BPS-M Exposure:BPS | Gender:male RAW_FILE_NAME=2007-BPS-M.D SUBJECT_SAMPLE_FACTORS - 2031-BPS-M Exposure:BPS | Gender:male RAW_FILE_NAME=2031-BPS-M.D SUBJECT_SAMPLE_FACTORS - 2029-Control-F Exposure:CTRL | Gender:female RAW_FILE_NAME=2029-Control-F.D SUBJECT_SAMPLE_FACTORS - 2005-Control-F Exposure:CTRL | Gender:female RAW_FILE_NAME=2005-Control-F.D SUBJECT_SAMPLE_FACTORS - 3009-Control-F Exposure:CTRL | Gender:female RAW_FILE_NAME=3009-Control-F.D SUBJECT_SAMPLE_FACTORS - 3008-Control-F Exposure:CTRL | Gender:female RAW_FILE_NAME=3008-Control-F.D SUBJECT_SAMPLE_FACTORS - 2017-Control-F Exposure:CTRL | Gender:female RAW_FILE_NAME=2017-Control-F.D SUBJECT_SAMPLE_FACTORS - 3009-Control-M Exposure:CTRL | Gender:male RAW_FILE_NAME=3009-Control-M.D SUBJECT_SAMPLE_FACTORS - 2005-Control-M Exposure:CTRL | Gender:male RAW_FILE_NAME=2005-Control-M.D SUBJECT_SAMPLE_FACTORS - 2029-Control-M Exposure:CTRL | Gender:male RAW_FILE_NAME=2029-Control-M.D SUBJECT_SAMPLE_FACTORS - 3008-Control-M Exposure:CTRL | Gender:male RAW_FILE_NAME=3008-Control-M.D SUBJECT_SAMPLE_FACTORS - 2017-Control-M Exposure:CTRL | Gender:male RAW_FILE_NAME=2017-Control-M.D #COLLECTION CO:COLLECTION_SUMMARY e12.5 placenta samples were collected immediately after dams were euthanized CO:SAMPLE_TYPE Placenta CO:COLLECTION_METHOD snap freezen in LN2 right after placenta was isolated CO:COLLECTION_LOCATION Univ. of Missouri Animal Science Research Center CO:COLLECTION_FREQUENCY 1 time from each animal CO:VOLUMEORAMOUNT_COLLECTED 50 mg CO:STORAGE_VIALS 1.5 ml vials #TREATMENT TR:TREATMENT_SUMMARY 2 weeks before breeding and during gestation, dams were fed BPA, or BPS, or TR:TREATMENT_SUMMARY vehicle till tissue collection at 12.5 DPC TR:TREATMENT_ROUTE oral TR:TREATMENT_DOSE 200 microg/kg body weight TR:TREATMENT_VEHICLE 70% ethanol dry out onto Nabisco Nilla wafer TR:ANIMAL_ENDP_EUTHANASIA yes TR:TREATMENT_COMPOUND Bisphenol A, bisphenol S #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Freshly isolated mouse placneta tissue was snap frozen in liqud nitrogen and no SP:SAMPLEPREP_SUMMARY further preparation. Samples were storred at -52C in lyophilyzer and -20C after SP:SAMPLEPREP_SUMMARY weighing. Extracts were dried under nitrogen flow and stored at -20C SP:PROCESSING_METHOD Frozen tissues were lyophilized for 24 hrs before weighing. SP:PROCESSING_STORAGE_CONDITIONS Described in summary SP:EXTRACTION_METHOD samples were extracted water, containing 25 μg/ml ribitol, vortex for 20 SP:EXTRACTION_METHOD seconds, sonicated for 15 minutes, incubated at 50C for 1h SP:EXTRACT_ENRICHMENT Sample tubes were sonicated for 15 minutes at 13000g then collected the SP:EXTRACT_ENRICHMENT supernatantant SP:EXTRACT_STORAGE -20℃ SP:SAMPLE_RESUSPENSION Sample was resuspended in 25 μl of pyridine containing 15 mg/ml methoxyamine SP:SAMPLE_RESUSPENSION hydrochloride SP:SAMPLE_DERIVATIZATION Sample was derivatized with 25 μl MSTFA SP:SAMPLE_DERIVATIZATION (N-methyl-N-(trimethyl-silyl)trifluoroacetamide) + 1% TMCS SP:SAMPLE_DERIVATIZATION (chlorotrimethylsilane) SP:SAMPLE_SPIKING Ribitol was used in the extraction solvent as an internal standard #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Agilent 6890N;Agilent 5973N CH:COLUMN_NAME DB-5MS (J&W Scientific) CH:FLOW_RATE 1mL/Min of Helium gas CH:COLUMN_TEMPERATURE 80 C for 2 min, then ramped at 5 C /min to 315 C and held at 315 C for 12 min CH:INJECTION_TEMPERATURE 280 C CH:INTERNAL_STANDARD Ribitol CH:SAMPLE_INJECTION split injection (split ration 5:1) with 1uL injection volume CH:CHROMATOGRAPHY_COMMENTS Agilent 6890N coupled to an Agilent 5973N #ANALYSIS AN:ANALYSIS_TYPE MS AN:DETECTOR_TYPE electron multiplier #MS MS:INSTRUMENT_NAME Agilent 5973 MS:INSTRUMENT_TYPE Single quadrupole MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:MS_COMMENTS 70 eV electron ionization MS:ION_SOURCE_TEMPERATURE 230 C MS:MASS_ACCURACY 0.1 m/z MS:DATAFORMAT .D MS:SCAN_RANGE_MOVERZ 50 to 650 m/z MS:MS_RESULTS_FILE ST001310_AN002181_Results.txt UNITS:relative instrument response Has m/z:Yes Has RT:Yes RT units:Minutes #END