#METABOLOMICS WORKBENCH hormel101_20170623_115935 DATATRACK_ID:1086 STUDY_ID:ST000637 ANALYSIS_ID:AN000969 PROJECT_ID:PR000457 VERSION 1 CREATED_ON June 25, 2017, 9:30 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 and ST:STUDY_TITLE Kidney Tissue (part X) 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 - ms6103-1 sample source:YCR1 urine | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-2 sample source:YCR2 urine | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-3 sample source:YCR3 urine | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-4 sample source:YCR4 urine | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-5 sample source:YCR5 urine | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-6 sample source:YCR6 urine | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-7 sample source:YCR7 urine | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-8 sample source:YCR8 urine | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-9 sample source:YCR1 outer medulla kidney | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-10 sample source:YCR1 outer medulla kidney | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-11 sample source:YCR1 outer medulla kidney | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-12 sample source:YCR1 outer medulla kidney | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-13 sample source:YCR1 outer medulla kidney | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-14 sample source:YCR1 outer medulla kidney | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-15 sample source:YCR1 outer medulla kidney | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-16 sample source:YCR1 outer medulla kidney | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-17 sample source:YCR1 kidney cortex | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-18 sample source:YCR1 kidney cortex | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-19 sample source:YCR1 kidney cortex | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-20 sample source:YCR1 kidney cortex | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-21 sample source:YCR1 kidney cortex | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-22 sample source:YCR1 kidney cortex | group:Transgenic Negative SUBJECT_SAMPLE_FACTORS - ms6103-23 sample source:YCR1 kidney cortex | group:Transgenic Het SUBJECT_SAMPLE_FACTORS - ms6103-24 sample source:YCR1 kidney cortex | group:Transgenic Het #COLLECTION CO:COLLECTION_SUMMARY The objective of the study is to identify Fumarase in hypertension. Samples are CO:COLLECTION_SUMMARY from Transgenic fumerase rats and WT littermates, 13 weeks of age #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 and kidney tissue extract (outer medulla and cortex) TCA SP:SAMPLEPREP_SUMMARY concentrations. Urine TCA concentrations in uM and tissue in nmol/vial #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE GC CH:INSTRUMENT_NAME Agilent 7890B 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 urine uM and Tissue nnol/vial MS_METABOLITE_DATA_START Samples ms6103-1 ms6103-2 ms6103-3 ms6103-4 ms6103-5 ms6103-6 ms6103-7 ms6103-8 ms6103-9 ms6103-10 ms6103-11 ms6103-12 ms6103-13 ms6103-14 ms6103-15 ms6103-16 ms6103-17 ms6103-18 ms6103-19 ms6103-20 ms6103-21 ms6103-22 ms6103-23 ms6103-24 Factors sample source:YCR1 urine | group:Transgenic Het sample source:YCR2 urine | group:Transgenic Negative sample source:YCR3 urine | group:Transgenic Negative sample source:YCR4 urine | group:Transgenic Negative sample source:YCR5 urine | group:Transgenic Het sample source:YCR6 urine | group:Transgenic Negative sample source:YCR7 urine | group:Transgenic Het sample source:YCR8 urine | group:Transgenic Het sample source:YCR1 outer medulla kidney | group:Transgenic Het sample source:YCR1 outer medulla kidney | group:Transgenic Negative sample source:YCR1 outer medulla kidney | group:Transgenic Negative sample source:YCR1 outer medulla kidney | group:Transgenic Negative sample source:YCR1 outer medulla kidney | group:Transgenic Het sample source:YCR1 outer medulla kidney | group:Transgenic Negative sample source:YCR1 outer medulla kidney | group:Transgenic Het sample source:YCR1 outer medulla kidney | group:Transgenic Het sample source:YCR1 kidney cortex | group:Transgenic Het sample source:YCR1 kidney cortex | group:Transgenic Negative sample source:YCR1 kidney cortex | group:Transgenic Negative sample source:YCR1 kidney cortex | group:Transgenic Negative sample source:YCR1 kidney cortex | group:Transgenic Het sample source:YCR1 kidney cortex | group:Transgenic Negative sample source:YCR1 kidney cortex | group:Transgenic Het sample source:YCR1 kidney cortex | group:Transgenic Het Lactate 144.5879715 163.3869868 147.9087896 161.3399022 225.9648866 234.1238469 201.1648425 158.0621429 70.41931523 109.9783564 51.69977488 49.2548717 132.07328 67.58463763 76.44302082 60.81785818 57.9169519 43.99978211 69.69404647 43.1071769 36.15357664 51.07227153 44.42440613 32.75971285 Succinate 376.6042221 236.278415 442.3836203 391.4180368 459.0337678 555.9326651 528.6472154 345.2674004 0.275828341 0.280531932 0.470663447 0.391736926 0.448602988 0.291707716 0.344452676 0.477036225 3.478982347 2.446334652 1.841759703 2.20931755 1.046144233 2.663534366 2.659472969 1.018132573 Fumarate 27.35765233 23.41227583 45.26624011 34.88986622 50.99997482 54.20344364 52.23906919 28.52839327 3.229161358 4.746492314 3.918437574 3.405514286 5.398123373 4.276174357 3.921426476 4.201088983 11.95826626 8.69627553 7.81958554 9.944829418 7.62847972 11.90207924 10.54356199 6.327963862 Oxaloacetate 0.137318163 0.277782156 0.100946071 0.458759962 0.14319496 0.151160611 0.244415636 0.192553687 0.001346323 0.001286517 0.001119809 0.001173311 0.00128582 0.001501324 0.001303872 0.001164633 0.001662698 0.001423493 0.001998015 0.001495605 0.001570409 0.001572448 0.00145025 0.0015042 Ketoglutarate 2137.728155 1948.203773 2712.604563 2613.7795 2985.030176 3109.296125 2863.740135 2132.944119 0.092530566 0.082551749 0.045158337 0.061745697 0.055641492 0.097948921 0.072821988 0.029139362 0.017588974 0.014358712 0.019168135 0.014881659 0.023538338 0.018509554 0.016137338 0.020839126 Malate 117.7696734 112.2284881 216.5292556 160.5669756 257.1223031 258.1982355 264.2839727 133.0076033 3.301894268 5.032577282 4.228710046 3.608728546 5.718956003 4.539420231 4.065815229 4.366539214 11.78633713 8.781565804 7.890108787 10.20222461 8.334685266 11.55012814 10.30338728 7.349169301 2-Hydroxyglutarate 64.37537087 49.93756336 117.4230715 96.77897908 127.9207811 134.5506116 112.8715137 58.81611837 0.037512507 0.031416767 0.066061054 0.047425589 0.078831204 0.040231245 0.045536259 0.058515866 0.163960849 0.176403442 0.194837993 0.185118253 0.153125506 0.133480684 0.149333468 0.148977156 cis-Aconitic Acid 1029.787606 1171.352411 940.4219631 1205.956607 1163.463561 1433.549009 1028.676995 1132.792685 0.134249099 0.141787272 0.163819413 0.148888996 0.220166284 0.08599724 0.139262588 0.12902585 0.732725365 0.417948698 0.220638662 0.38036185 0.335830497 0.592785388 0.427892671 0.259478777 Citrate 4970.491439 3820.926274 6610.428291 5228.086388 6974.900632 6448.182492 6424.683459 4457.366735 0.047801992 0.04670287 0.056938148 0.139964781 0.241844534 0.03493677 0.08745062 0.13123964 0.160803428 0.104763122 0.208830928 0.14963621 0.059911797 0.147088637 0.143284135 0.074969122 Isocitrate 1207.358516 1311.788051 923.5192539 1229.197558 1166.991168 1251.995026 1036.827422 1224.953264 0.009918952 0.014974751 0.014979402 0.017248293 0.048584952 0.008791285 0.018255185 0.021455575 0.031567107 0.027051222 0.032938341 0.033232757 0.017347037 0.02899638 0.037480099 0.023077482 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name Lactate Succinate Fumarate Oxaloacetate Ketoglutarate Malate 2-Hydroxyglutarate cis-Aconitic Acid Citrate Isocitrate METABOLITES_END #END