#METABOLOMICS WORKBENCH araskind_20150313_9045041_mwtab.txt DATATRACK_ID:222 STUDY_ID:ST000143 ANALYSIS_ID:AN000227 PROJECT_ID:PR000219 VERSION 1 CREATED_ON 01-15-2019 #PROJECT PR:PROJECT_TITLE Metabolic analysis of Parp1 ko/wt Saline & Bleo Mouse Lung Fibroblasts and Human PR:PROJECT_TITLE IPF & Normal Lung Fibroblasts PR:PROJECT_TYPE Glycolysis/TCA/Nucleotide analysis (tissue/cells) PR:PROJECT_SUMMARY Hedgehog signaling plays important roles in cell development and PR:PROJECT_SUMMARY differentiation. In this study, the ability of Sonic Hedgehog (SHH) to induce PR:PROJECT_SUMMARY myofibroblast differentiation was analyzed in isolated human lung fibroblasts, PR:PROJECT_SUMMARY and its in vivo significance was evaluated in rodent bleomycin-induced pulmonary PR:PROJECT_SUMMARY fibrosis. The results showed that SHH could induce myofibroblast differentiation PR:PROJECT_SUMMARY in human lung fibroblasts in a Smo- and Gli1-dependent manner. Gel shift PR:PROJECT_SUMMARY analysis, chromatin immunoprecipitation assay, and site-directed mutagenesis PR:PROJECT_SUMMARY revealed that a Gli1 binding consensus in the ?-SMA gene promoter was important PR:PROJECT_SUMMARY for mediating SHH-induced myofibroblast differentiation. Analysis of Hedgehog PR:PROJECT_SUMMARY reemergence in vivo revealed that of all three Hedgehog isoforms, only SHH was PR:PROJECT_SUMMARY significantly induced in bleomycin-injured lung along with Gli1. The induction PR:PROJECT_SUMMARY of SHH was only noted in epithelial cells, and its expression was undetectable PR:PROJECT_SUMMARY in lung fibroblasts or macrophages. Transforming growth factor (TGF)-? induced PR:PROJECT_SUMMARY SHH significantly in cultured alveolar epithelial cells, whereas SHH induced PR:PROJECT_SUMMARY TGF-? in lung fibroblasts. Pulmonary fibrosis and ?-smooth muscle actin (?-SMA) PR:PROJECT_SUMMARY expression were significantly reduced in mice that were Smo deficient only in PR:PROJECT_SUMMARY type I collagenexpressing cells. Thus, the reemergence of SHH in epithelial PR:PROJECT_SUMMARY cells could result in induction of myofibroblast differentiation in a PR:PROJECT_SUMMARY Smo-dependent manner and subsequent Gli1 activation of the ?-SMA promoter. PR:INSTITUTE University of Michigan PR:DEPARTMENT Deaprtment of Pathology PR:LABORATORY Sem H. Phan PR:LAST_NAME Hu PR:FIRST_NAME Biao PR:ADDRESS Ann Arbor, MI PR:EMAIL biaohu@med.umich.edu PR:PHONE 734-7635731 PR:DOI http://dx.doi.org/10.21228/M8359Z #STUDY ST:STUDY_TITLE Metabolic analysis of Normal Mouse Lung Fiboblasts with/without TGFbeta ST:STUDY_TITLE treatment (Part 1) ST:STUDY_TYPE Glycolysis/TCA/Nucleotide analysis (tissue/cells) ST:STUDY_SUMMARY This study is a part of series performed for the same researcher through ST:STUDY_SUMMARY pilot/feasibility grant program, so the publication is relevant reference for ST:STUDY_SUMMARY other studies, see the project for this study. This specific experiment is a ST:STUDY_SUMMARY small pilot study to establish method performance, it includes four biological ST:STUDY_SUMMARY replicas of identical cell cultures after the identical treatment and a single ST:STUDY_SUMMARY tissue sample. ST:INSTITUTE University of Michigan ST:DEPARTMENT Biomedical Research Core Facilities ST:LABORATORY Metabolomics core ST:LAST_NAME Kachman ST:FIRST_NAME Maureen ST:ADDRESS 6300 Brehm Tower, 1000 Wall Street, Ann Arbor, MI 48105-5714 ST:EMAIL mkachman@umich.edu ST:PHONE - #SUBJECT SU:SUBJECT_TYPE Animal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 SU:SPECIES_GROUP Mammal #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS SU0003983 S00014557 Genotype:parp1 ko | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003984 S00014558 Genotype:parp1 ko | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003985 S00014559 Genotype:parp1 ko | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003994 S00014568 Genotype:parp1 ko | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003995 S00014569 Genotype:parp1 ko | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0004004 S00014578 Genotype:parp1 ko | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0004005 S00014579 Genotype:parp1 ko | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0004006 S00014580 Genotype:parp1 ko | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003980 S00014554 Genotype:parp1 ko | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0003981 S00014555 Genotype:parp1 ko | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0003982 S00014556 Genotype:parp1 ko | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0003992 S00014566 Genotype:parp1 ko | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0003993 S00014567 Genotype:parp1 ko | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0004001 S00014575 Genotype:parp1 ko | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0004002 S00014576 Genotype:parp1 ko | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0004003 S00014577 Genotype:parp1 ko | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0003989 S00014563 Genotype:parp1 wt | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003990 S00014564 Genotype:parp1 wt | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003991 S00014565 Genotype:parp1 wt | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003998 S00014572 Genotype:parp1 wt | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003999 S00014573 Genotype:parp1 wt | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0004000 S00014574 Genotype:parp1 wt | Treatment:bleomycin SUBJECT_SAMPLE_FACTORS SU0003986 S00014560 Genotype:parp1 wt | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0003987 S00014561 Genotype:parp1 wt | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0003988 S00014562 Genotype:parp1 wt | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0003996 S00014570 Genotype:parp1 wt | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0003997 S00014571 Genotype:parp1 wt | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0004007 S00014581 Genotype:parp1 wt | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0004008 S00014582 Genotype:parp1 wt | Treatment:saline SUBJECT_SAMPLE_FACTORS SU0004009 S00014583 Genotype:parp1 wt | Treatment:saline #COLLECTION CO:SAMPLE_TYPE Tissues CO:COLLECTION_SUMMARY - #TREATMENT TR:TREATMENT_SUMMARY - #SAMPLEPREP SP:SAMPLEPREP_PROTOCOL_FILENAME Gly-TCA-nucleotides_analysis_protocol-2015-03-09.docx SP:SAMPLEPREP_SUMMARY - #CHROMATOGRAPHY CH:METHODS_ID AQM011 CH:METHODS_FILENAME ALPHA_KETO_ACIDS-FULL.M.zip CH:INSTRUMENT_NAME Agilent 7890A CH:COLUMN_NAME Agilent DB5-MS (30m × 0.25mm, 0.25um) CH:CHROMATOGRAPHY_TYPE GC #ANALYSIS AN:LABORATORY_NAME MRC2 (University of Michigan) AN:ANALYSIS_TYPE MS AN:ACQUISITION_PARAMETERS_FILE ALPHA_KETO_ACIDS-FULL.M.zip AN:PROCESSING_PARAMETERS_FILE EX00294-MassHunterQuant-GlyTCA-GCMS-DataAnalysis-Method.m.zip #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Agilent 5975C MS:INSTRUMENT_TYPE Single quadrupole MS:MS_TYPE EI MS:ION_MODE POSITIVE MS:TUBE_LENS_VOLTAGE Positive #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS uM MS_METABOLITE_DATA_START Samples S00014557 S00014558 S00014559 S00014568 S00014569 S00014578 S00014579 S00014580 S00014554 S00014555 S00014556 S00014566 S00014567 S00014575 S00014576 S00014577 S00014563 S00014564 S00014565 S00014572 S00014573 S00014574 S00014560 S00014561 S00014562 S00014570 S00014571 S00014581 S00014582 S00014583 Factors Genotype:parp1 ko | Treatment:bleomycin Genotype:parp1 ko | Treatment:bleomycin Genotype:parp1 ko | Treatment:bleomycin Genotype:parp1 ko | Treatment:bleomycin Genotype:parp1 ko | Treatment:bleomycin Genotype:parp1 ko | Treatment:bleomycin Genotype:parp1 ko | Treatment:bleomycin Genotype:parp1 ko | Treatment:bleomycin Genotype:parp1 ko | Treatment:saline Genotype:parp1 ko | Treatment:saline Genotype:parp1 ko | Treatment:saline Genotype:parp1 ko | Treatment:saline Genotype:parp1 ko | Treatment:saline Genotype:parp1 ko | Treatment:saline Genotype:parp1 ko | Treatment:saline Genotype:parp1 ko | Treatment:saline Genotype:parp1 wt | Treatment:bleomycin Genotype:parp1 wt | Treatment:bleomycin Genotype:parp1 wt | Treatment:bleomycin Genotype:parp1 wt | Treatment:bleomycin Genotype:parp1 wt | Treatment:bleomycin Genotype:parp1 wt | Treatment:bleomycin Genotype:parp1 wt | Treatment:saline Genotype:parp1 wt | Treatment:saline Genotype:parp1 wt | Treatment:saline Genotype:parp1 wt | Treatment:saline Genotype:parp1 wt | Treatment:saline Genotype:parp1 wt | Treatment:saline Genotype:parp1 wt | Treatment:saline Genotype:parp1 wt | Treatment:saline Alanine 151.6573 70.4766 93.5807 111.4252 139.9279 167.2569 205.0150 90.1871 7.7040 82.4592 30.6312 25.5918 76.4968 95.2423 155.7659 128.3495 90.5514 63.8067 72.5396 181.4594 108.1661 105.7941 55.5827 35.3740 80.7843 134.3805 113.6705 123.8192 75.1341 44.2565 alpha-Ketoglutarate 7.1708 7.0555 4.8385 8.8545 6.4383 7.5234 8.4837 6.2941 1.2637 10.8507 6.3356 7.6072 11.2267 10.7315 15.2899 15.2920 6.4578 7.2312 6.8530 7.5326 6.4014 6.5092 8.6642 7.3092 11.8568 16.6321 13.3534 12.8941 14.1187 9.2709 Aspartate 2076.4871 797.7439 1433.4756 1242.7894 1616.6492 349.7478 2231.0293 1320.6448 188.9277 1959.6168 650.3874 683.4647 1950.4351 1642.2770 3256.4948 3183.1172 1255.9257 730.8475 1014.8004 802.7163 884.0014 853.8910 1169.0253 846.5137 2134.1592 2205.6471 2677.9243 2065.5850 2104.9137 1540.6600 Fumarate 17.0088 11.6572 9.0879 12.3454 15.2743 19.2316 17.1799 9.3336 0.9722 11.5210 4.9353 4.8900 15.1165 17.4668 32.6831 23.4230 9.8914 14.7183 10.4935 25.6170 17.2919 13.5918 7.2624 5.0299 13.8872 23.7657 21.7977 15.3666 21.3472 10.0972 Glutamate 2105.7375 1099.1217 1469.9426 1842.7389 3276.6622 627.2081 2256.9272 1173.1735 160.3561 1701.1911 576.3003 1143.0470 1493.4309 1303.6415 2794.0158 2330.5132 1427.3856 1568.6731 1539.0837 1239.6388 1353.0150 1105.3898 956.7064 737.4067 1818.2787 1923.7723 2136.3574 1550.7936 1499.0787 1386.2061 Glutamine Lactate 251.1005 158.1682 130.6085 166.7536 222.1973 892.3098 261.5111 101.4923 8.1847 116.0776 48.8403 36.7356 117.2674 121.9344 219.1280 146.0706 121.9681 152.3817 134.0089 1458.5798 257.9958 222.2661 87.8774 32.1373 88.4476 161.6987 152.1650 114.2767 116.7061 36.3362 Oxaloacetate Pyruvate 0.8096 0.4030 1.0915 1.6420 2.0036 4.1500 3.7822 2.0267 0.0281 1.1693 0.1397 0.2605 1.6177 3.1275 5.1975 2.1320 0.8377 1.1696 0.8625 6.7898 2.3476 1.7819 0.5193 0.2656 4.7473 2.8914 0.9335 1.5561 1.1019 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name pubchem_id inchi_key kegg_id other_id other_id_type ri ri_type moverz_quant Alanine 5950 C00041 ALA UM_Target_ID alpha-Ketoglutarate 51 C00026 AKG UM_Target_ID Aspartate 5960 C00049 Asp UM_Target_ID Fumarate 444972 C00122 Fum UM_Target_ID Glutamate 611 Glu UM_Target_ID Glutamine 5961 C00064 Gln UM_Target_ID Lactate 107689 C00186 Lac UM_Target_ID Oxaloacetate 970 C00036 OAA UM_Target_ID Pyruvate 1060 C00022 Pyr UM_Target_ID METABOLITES_END #END