#METABOLOMICS WORKBENCH hormel101_20170623_093204 DATATRACK_ID:1078 STUDY_ID:ST000631 ANALYSIS_ID:AN000963 PROJECT_ID:PR000457 VERSION 1 CREATED_ON June 25, 2017, 8:54 am #PROJECT PR:PROJECT_TITLE Metabolomic Mechanisms of Dietary Salt Effects on Blood Pressure PR:PROJECT_SUMMARY "Enhanced sensitivity of blood pressure to salt intake is observed in PR:PROJECT_SUMMARY approximately 50% of hypertensive patients, reaching 75% in African American PR:PROJECT_SUMMARY hypertensive patients. We recently discovered a novel role of abnormal cellular PR:PROJECT_SUMMARY intermediary metabolism in hypertension in the Dahl salt-sensitive (SS) rat, the PR:PROJECT_SUMMARY most commonly used polygenic, hereditary model of human salt-sensitive PR:PROJECT_SUMMARY hypertension. We propose to test the hypothesis that blood pressure sensitivity PR:PROJECT_SUMMARY to dietary salt intake in human is associated with metabolite changes in the PR:PROJECT_SUMMARY urine. Leveraging the expertise and resources at the Mayo Clinic Metabolomics PR:PROJECT_SUMMARY Resources Core, we propose to perform targeted LC/MS analysis and NMR spectra PR:PROJECT_SUMMARY generation in urine samples obtained from a subset of subjects from the Dietary PR:PROJECT_SUMMARY Approaches to Stop Hypertension – Sodium (DASH2) clinical trial and kidney PR:PROJECT_SUMMARY tissue extract and urine samples from SS rats and a newly generated transgenic PR:PROJECT_SUMMARY rat that overexpresses fumarase (SS.Fh1+). The study will be the first to PR:PROJECT_SUMMARY systematically characterize urinary metabolite profiles associated with blood PR:PROJECT_SUMMARY pressure response to salt in humans. The study is anticipated to generate new PR:PROJECT_SUMMARY insight into the mechanisms (particularly renal mechanisms) underlying PR:PROJECT_SUMMARY salt-sensitive hypertension. Findings of the proposed study could lead to an PR:PROJECT_SUMMARY expanded clinical study as well as mechanistic studies in animal models." PR:INSTITUTE Mayo Clinic PR:LAST_NAME Liang PR:FIRST_NAME Mingyu PR:ADDRESS Medical College of Wisconsin 8701 Watertown Plank Road Milwaukee, WI 53226 PR:EMAIL mliang@mcw.edu PR:PHONE 414-955-8539 #STUDY ST:STUDY_TITLE TCA Cycle Metabolites of Dietary Salt Effects on Blood Pressure in Rat Urine ST:STUDY_TITLE (part IV) ST:STUDY_SUMMARY We propose to analyze kidney tissue extract and urine samples from SS and ST:STUDY_SUMMARY SS.Fh1+ transgenic rats in addition to the analysis of urine samples from the ST:STUDY_SUMMARY DASH2 trial. The analysis of the rat samples will be highly valuable for several ST:STUDY_SUMMARY reasons. First, it will to take the findings in human subjects back to animal ST:STUDY_SUMMARY models and prepare us for further mechanistic studies. We hypothesize at least ST:STUDY_SUMMARY some of the effects of dietary salt intake on metabolite profiles in human will ST:STUDY_SUMMARY be recapitulated or altered in the SS rat. If this is confirmed, we will have a ST:STUDY_SUMMARY highly informative animal model ready for mechanistic studies in which we can ST:STUDY_SUMMARY investigate the functional contribution of specific metabolites to hypertension ST:STUDY_SUMMARY and the mechanisms involved. Second, the rat study will allow us to take ST:STUDY_SUMMARY advantage of a new and unique transgenic SS.Fh1+ model that we recently ST:STUDY_SUMMARY developed that overexpresses fumarase (Fh1) on the genetic background of the SS ST:STUDY_SUMMARY rat. Fumarase is a TCA cycle enzyme previously implicated in salt-induced ST:STUDY_SUMMARY hypertension in SS rats. ST:INSTITUTE Mayo Clinic ST:LAST_NAME Liang ST:FIRST_NAME Mingyu ST:ADDRESS Medical College of Wisconsin 8701 Watertown Plank Road Milwaukee, WI 53226 ST:EMAIL mliang@mcw.edu ST:PHONE 414-955-8539 #SUBJECT SU:SUBJECT_TYPE Rat SU:SUBJECT_SPECIES Rattus norvegicus SU:TAXONOMY_ID 10116 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS - ms5952-1 Time point:Control Date of study=8/3/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-2 Time point:Control Date of study=8/3/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-3 Time point:Control Date of study=8/3/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-4 Time point:Control Date of study=8/3/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-5 Time point:Control Date of study=8/3/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-6 Time point:Control Date of study=8/3/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-7 Time point:Control Date of study=8/3/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-8 Time point:Control Date of study=8/3/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-9 Time point:7 day HS Date of study=8/10/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-10 Time point:7 day HS Date of study=8/10/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-11 Time point:7 day HS Date of study=8/10/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-12 Time point:7 day HS Date of study=8/10/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-13 Time point:7 day HS Date of study=8/10/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-14 Time point:7 day HS Date of study=8/10/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-15 Time point:7 day HS Date of study=8/10/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-16 Time point:7 day HS Date of study=8/10/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-17 Time point:Control Date of study=9/2/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-18 Time point:Control Date of study=9/2/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-19 Time point:Control Date of study=9/2/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-20 Time point:Control Date of study=9/2/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-21 Time point:Control Date of study=9/2/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-22 Time point:Control Date of study=9/2/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-23 Time point:Control Date of study=9/2/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-24 Time point:Control Date of study=9/2/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-25 Time point:Control Date of study=9/2/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-26 Time point:Control Date of study=9/2/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-27 Time point:7 day HS Date of study=9/9/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-28 Time point:7 day HS Date of study=9/9/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-29 Time point:7 day HS Date of study=9/9/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-30 Time point:7 day HS Date of study=9/9/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-31 Time point:7 day HS Date of study=9/9/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-32 Time point:7 day HS Date of study=9/9/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-33 Time point:7 day HS Date of study=9/9/2010; gender=M; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-34 Time point:7 day HS Date of study=9/9/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-35 Time point:7 day HS Date of study=9/9/2010; gender=F; sample source=rat urine SUBJECT_SAMPLE_FACTORS - ms5952-36 Time point:7 day HS Date of study=9/9/2010; gender=F; sample source=rat urine #COLLECTION CO:COLLECTION_SUMMARY Samples are from Transgenic fumerase rats and WT littermates, 13 weeks of age, CO:COLLECTION_SUMMARY placed on low salt or high salt dyets. We are interested in any differences in CO:COLLECTION_SUMMARY the TCA and AA intermediates. #TREATMENT TR:TREATMENT_SUMMARY We will analyze kidney tissue extract and urine samples from SS rats and the TR:TREATMENT_SUMMARY newly generated SS.Fh1+ transgenic rats. The SS.Fh1+ rat was generated using a TR:TREATMENT_SUMMARY Sleeping Beauty transposon-mediated transgenic technique. Overexpression of TR:TREATMENT_SUMMARY fumarase has been confirmed in SS.Fh1+ rats. Preliminary study indicated that TR:TREATMENT_SUMMARY the development of salt-induced hypertension was altered in SS.Fh1+ rats TR:TREATMENT_SUMMARY compared to wild-type littermate SS rats. We will analyze kidney tissue extract TR:TREATMENT_SUMMARY and urine samples collected from rats maintained on a 0.4% NaCl diet or switched TR:TREATMENT_SUMMARY to a 4% NaCl diet for 7 days. The dietary protocol has been used in numerous TR:TREATMENT_SUMMARY studies to examine mechanisms underlying salt sensitivity including early TR:TREATMENT_SUMMARY responses (7 days) to salt in the SS model. In total, 64 rat samples (two rat TR:TREATMENT_SUMMARY strains, two salt levels, and 8 rats per condition) will be analyzed. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Rat urine TCA concentrations #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Agilent 7890A CH:COLUMN_NAME Agilent HP5-MS (30m × 0.25mm, 0.25 um) #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Agilent 5975C MS:INSTRUMENT_TYPE Single quadrupole MS:MS_TYPE EI MS:ION_MODE POSITIVE #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS uM MS_METABOLITE_DATA_START Samples ms5952-1 ms5952-2 ms5952-3 ms5952-4 ms5952-5 ms5952-6 ms5952-7 ms5952-8 ms5952-9 ms5952-10 ms5952-11 ms5952-12 ms5952-13 ms5952-14 ms5952-15 ms5952-16 ms5952-17 ms5952-18 ms5952-19 ms5952-20 ms5952-21 ms5952-22 ms5952-23 ms5952-24 ms5952-25 ms5952-26 ms5952-27 ms5952-28 ms5952-29 ms5952-30 ms5952-31 ms5952-32 ms5952-33 ms5952-34 ms5952-35 ms5952-36 Factors Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:Control Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Time point:7 day HS Lactate 121.8370428 323.9830588 176.0009637 151.5138174 161.1559509 142.7735244 240.7109416 448.0034726 58.7345206 64.86696052 78.29252482 79.31890972 73.27702133 98.56697829 138.7821383 91.6556478 227.8093975 217.5316889 115.5196936 86.91417157 160.1013193 118.5660886 243.7256764 383.95311 174.6952399 126.4499657 75.40695843 105.8831837 115.0870091 70.53544481 71.76623988 106.208651 218.4515234 69.82112055 200.7405387 109.4228018 Succinate 99.2996057 170.0122844 126.2426559 83.06418732 142.0350439 111.964387 219.4656715 272.0321653 27.03761761 25.18949179 45.10522927 33.017913 54.99796583 83.54079335 87.14598147 40.64901018 113.4271406 69.81592255 137.0905841 97.95176824 82.58426701 88.41309726 82.7877143 123.4418308 82.08857833 47.87592963 25.20688216 41.93986406 58.25854226 60.30933219 30.81638033 37.27471168 21.83247958 108.1775993 87.02885222 144.0841026 Fumarate 24.24363463 24.72609837 19.54562058 12.91218637 29.9096659 15.87095766 30.42395502 44.29997268 7.332107871 4.820612287 8.212504645 8.204195041 20.86932529 21.08381028 29.33639825 9.949379978 21.75077352 14.63179105 43.92036612 27.46179054 20.97561915 10.71267985 14.2940784 53.13607146 63.2771294 14.86354399 9.714580403 14.4071471 31.43414662 14.87726872 8.119773687 14.23628981 8.460692503 48.18066297 51.39143038 32.85776176 Oxaloacetate 0.509410392 2.161661189 6.771573213 2.876256604 1.392368032 1.80676983 2.503243982 3.393510193 1.714802055 0.910044588 0.812163259 0.717056444 0.698382088 1.007731905 1.10146809 1.346342473 0.492574789 2.26754096 1.268028258 1.856902496 2.128398008 1.634346239 2.796929632 1.075923288 1.551617792 2.07055415 2.34565664 0.526553821 0.6300006 1.174408933 1.515630409 0.771276456 0.763298269 0.958073205 0.730029136 1.435455569 Ketoglutarate 1092.490743 1516.949378 1241.317411 783.1996532 1021.062964 506.5997198 899.2445391 1200.643254 536.3670269 366.8365912 589.6898584 576.979829 460.3486046 539.5678333 894.4679505 257.7621708 1347.806759 891.6797468 1326.770444 841.1916018 1272.542809 832.3379793 1178.153328 1757.440993 1476.960829 826.8673245 596.2631529 858.2225872 949.4360479 614.5843141 463.5759237 764.1377449 549.1289682 860.9199463 1056.756464 1179.545139 Malate 63.87881006 79.90054229 56.39464817 37.29369649 130.0189312 62.02082194 128.4469148 194.0166219 27.07188918 15.09795756 29.13442528 28.38366951 93.97391204 86.84747069 124.8918446 42.96209835 75.14731576 47.942705 195.6903144 129.4884383 70.35590308 33.02586268 50.10494887 262.5152459 272.8387953 57.03786996 38.15797319 62.63482771 141.7884585 68.16833832 30.15531287 59.18173674 31.81760034 225.5198828 227.9057526 151.8167219 cis-Aconitate 321.8058114 451.7885223 433.4575871 268.6780711 560.157153 297.2569097 535.8255304 667.3993756 105.013712 132.4733843 115.4313725 121.9559269 203.1356199 186.3002405 171.6320246 157.4321906 306.2705054 232.2790916 309.0943193 270.33129 282.0170421 232.2114349 292.7943086 420.1524209 389.4296346 274.9333854 87.63043341 92.83760192 181.742928 209.0391889 122.9851715 105.2447102 87.92753687 122.8047878 179.627693 147.5199027 Citrate 1149.156219 2324.864878 1487.278698 644.8026608 1485.13181 1065.545293 1887.529338 3590.641158 730.2351355 438.563212 866.6778595 643.6256638 969.850851 1482.677622 1484.623586 764.1787721 1674.163973 1418.760552 2952.975391 1762.140987 3757.2817 657.5915676 2822.095882 3236.214506 3705.006818 1238.362679 1147.591015 1998.523708 2099.779029 918.4933489 867.2343117 1668.007926 1017.093221 2301.833671 2824.545188 1976.593773 Isocitrate 772.9842309 1093.095218 1026.401267 631.2193035 1149.03909 587.3706989 1118.928719 1267.672806 220.3776209 297.6452208 235.4591342 265.9325698 318.3567382 325.4077373 385.2770843 291.4189292 703.1795282 545.2759128 586.6950271 520.9851573 593.2774738 683.615703 808.546851 965.461703 711.1479104 700.0581655 168.0273567 179.223219 303.9014003 436.2109158 244.3746843 197.3372102 194.9796811 192.9298877 283.9526193 300.7173856 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Lactate Succinate Fumarate Oxaloacetate Ketoglutarate Malate cis-Aconitate Citrate Isocitrate METABOLITES_END #END