#METABOLOMICS WORKBENCH MingxinZhang_20250214_004041 DATATRACK_ID:5626 STUDY_ID:ST003746 ANALYSIS_ID:AN006152 PROJECT_ID:PR002332 VERSION 1 CREATED_ON February 15, 2025, 5:04 pm #PROJECT PR:PROJECT_TITLE Serum hormone-targeted metabolome of sleep-deprived mice PR:PROJECT_SUMMARY This project investigates the changes in serum hormone levels before and after 2 PR:PROJECT_SUMMARY days of sleep deprivation in mice, exploring the alterations in serum hormone PR:PROJECT_SUMMARY release caused by sleep deprivation. Metabolomic results indicate that in mice PR:PROJECT_SUMMARY subjected to 2 days of sleep deprivation, serum hormones such as DHT, PR:PROJECT_SUMMARY Progesterone, and Corticosterone were significantly reduced PR:INSTITUTE China Agriculture University PR:LAST_NAME Zhang PR:FIRST_NAME Mingxin PR:ADDRESS 2 West Yuanmingyuan Road, Beijing, Beijing, 100193, China PR:EMAIL zmx15310686122@163.com PR:PHONE 18800152903 #STUDY ST:STUDY_TITLE Serum hormone-targeted metabolome of sleep-deprived mice ST:STUDY_SUMMARY Sleep disturbances associate with pathogenesis of numerous chronic disorders, ST:STUDY_SUMMARY including chronic gastrointestinal diseases. However, the mechanism that ST:STUDY_SUMMARY transmits sleep disturbance-induced aberrant neural signaling from the brain to ST:STUDY_SUMMARY the gut remains elusive. We show that acute sleep deprivation (SD) impairs ST:STUDY_SUMMARY intestinal stem cell function, leading to shortening of crypt-villus ST:STUDY_SUMMARY architecture and Paneth cell loss. We identified dorsal motor nucleus of vagus ST:STUDY_SUMMARY (DMV) as the SD-sensitive central nervous system center that transmits sleep ST:STUDY_SUMMARY effects to the gut. SD aberrantly activates DMV neurons, driving excessive ST:STUDY_SUMMARY acetylcholine release from the vagus nerve into the gut. Acetylcholine triggers ST:STUDY_SUMMARY 5-hydroxytryptamine (5-HT) release by enterochromaffin cells and suppresses its ST:STUDY_SUMMARY reuptake via muscarinic receptors, thereby causing a spike in 5-HT levels. ST:STUDY_SUMMARY Elevated 5-HT induces excessive oxidative stress in intestinal stem cells ST:STUDY_SUMMARY through its receptor HTR4, promoting gut pathologies. Overall, we reveal ST:STUDY_SUMMARY SD-responsive neural circuit that controls intestinal stem cells and identify ST:STUDY_SUMMARY therapeutic strategies for mitigating SD-related gut diseases. ST:INSTITUTE China Agriculture University ST:LAST_NAME Zhang ST:FIRST_NAME Mingxin ST:ADDRESS 2 West Yuanmingyuan Road, Beijing, Beijing, 100193, China ST:EMAIL zmx15310686122@163.com ST:PHONE 18800152903 #SUBJECT SU:SUBJECT_TYPE Mammal SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 #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 1 SD0d_1 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FKPO220034829-1A SUBJECT_SAMPLE_FACTORS 2 SD0d_2 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FKPO220034830-1A SUBJECT_SAMPLE_FACTORS 3 SD0d_3 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FKPO220034831-1A SUBJECT_SAMPLE_FACTORS 4 SD0d_4 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FKPO220034832-1A SUBJECT_SAMPLE_FACTORS 5 SD0d_5 Sample source:serum | Treatment:Ctrl RAW_FILE_NAME(Raw name)=FKPO220034833-1A SUBJECT_SAMPLE_FACTORS 6 SD2d_1 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FKPO220034834-1A SUBJECT_SAMPLE_FACTORS 7 SD2d_2 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FKPO220034835-1A SUBJECT_SAMPLE_FACTORS 8 SD2d_3 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FKPO220034836-1A SUBJECT_SAMPLE_FACTORS 9 SD2d_4 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FKPO220034837-1A SUBJECT_SAMPLE_FACTORS 10 SD2d_5 Sample source:serum | Treatment:SD2d RAW_FILE_NAME(Raw name)=FKPO220034838-1A #COLLECTION CO:COLLECTION_SUMMARY Fresh blood from mice was collected via cardiac puncture before and after sleep CO:COLLECTION_SUMMARY deprivation. After allowing the blood to stand for 1 hour, it was centrifuged at CO:COLLECTION_SUMMARY 3000g for 10 minutes to obtain serumA total of 100 μL of serum sample was CO:COLLECTION_SUMMARY collected and mixed by adding 100 μL of methanol, and then 100 μL of water was CO:COLLECTION_SUMMARY added and mixed at 4 ° C The samples were centrifuged at 12000 rpm for 15 min. CO:COLLECTION_SUMMARY After that, the supernatant was removed and slowly passed through the Oasis CO:COLLECTION_SUMMARY PRiME HLB column. make The columella were washed with 5% methanol. Finally, the CO:COLLECTION_SUMMARY animal hormones were eluted using 90/10 (v/v) acetonitrile/methanol and CO:COLLECTION_SUMMARY collected The eluate was dried and redissolved before injection for LC-MS CO:COLLECTION_SUMMARY analysis. CO:SAMPLE_TYPE Blood (serum) #TREATMENT TR:TREATMENT_SUMMARY Serum samples were obtained immediately after mice were subjected to 48 hours of TR:TREATMENT_SUMMARY continuous sleep deprivation. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY A total of 100 μL of serum sample was collected and mixed by adding 100 μL of SP:SAMPLEPREP_SUMMARY methanol, and then 100 μL of water was added and mixed at 4 ° C The samples SP:SAMPLEPREP_SUMMARY were centrifuged at 12000 rpm for 15 min. After that, the supernatant was SP:SAMPLEPREP_SUMMARY removed and slowly passed through the Oasis PRiME HLB column. make The columella SP:SAMPLEPREP_SUMMARY were washed with 5% methanol. Finally, the animal hormones were eluted using SP:SAMPLEPREP_SUMMARY 90/10 (v/v) acetonitrile/methanol and collected The eluate was dried and SP:SAMPLEPREP_SUMMARY redissolved before injection for LC-MS analysis. #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME AB Sciex QTRAP 6500+ CH:COLUMN_NAME Kinetex C18 column 2.1×100mm (2.6μm) CH:SOLVENT_A 0.1% formic acid in 5mM ammonium acetate CH:SOLVENT_B 0.1% formic acid in acetonitrile CH:FLOW_GRADIENT initial 5% B, 2min; 5-40% B, 4min; 40-70% B, 7 min; 70-100% B, 7.5min; 100% B, CH:FLOW_GRADIENT 9min; 100-5% B, 9.1min; 5% B, 11min. CH:FLOW_RATE 0.30 mL/min CH:COLUMN_TEMPERATURE 45°C #ANALYSIS AN:ANALYSIS_TYPE MS #MS MS:INSTRUMENT_NAME Agilent 6520 QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:MS_COMMENTS Curtain Gas：35psi； Collision Gas：Medium；IonSpray Voltage：4500V； MS:MS_COMMENTS Temperature：550℃； Ion Source Gas 1：60； Ion Source Gas 2：60 The MS:MS_COMMENTS characteristic ion of each substance is screened by the triple quadrupole, and MS:MS_COMMENTS the signal of the characteristic ion is obtained in the detector No. Intensity MS:MS_COMMENTS (CPS), open the sample mass spectrometry file with SCIEX OS V1.4 software, and MS:MS_COMMENTS perform chromatographic peaks Integration and calibration work, the peak Area MS:MS_COMMENTS (Area) of each chromatographic peak represents the relative quantitative value MS:MS_COMMENTS of the corresponding substance, the most All chromatographic peak area integral MS:MS_COMMENTS data were then exported #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS ng/ml MS_METABOLITE_DATA_START Samples SD0d_1 SD0d_2 SD0d_3 SD0d_4 SD0d_5 SD2d_1 SD2d_2 SD2d_3 SD2d_4 SD2d_5 Factors Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:Ctrl Sample source:serum | Treatment:SD2d Sample source:serum | Treatment:SD2d Sample source:serum | Treatment:SD2d Sample source:serum | Treatment:SD2d Sample source:serum | Treatment:SD2d 11-deoxycortisol 5333000 3859000 2084000 3445000 1545000 908800 436500 210800 1126000 1573000 Pregnenolone 12710 5135 9559 9371 10870 6658 12010 7656 9167 23080 Progesterone 275700 853400 190800 196600 182700 89270 92730 63990 90320 2889000 Dihydrotestosterone 29120 28280 11990 17510 18850 4981 8140 Androstenedione 6659 5349 4089 4569 3195 5502 5411 3576 4954 13730 Melatonin 121900 121000 128600 111900 119800 115900 114100 109200 105800 125700 17α-hydroxypregnenolone 1881 4450 2527 9410 37090 15250 28410 9938 39290 Corticosterone 13430000 9735000 5221000 8677000 3958000 2308000 1090000 529100 2801000 3958000 Cortisol 1229000 1138000 757300 1098000 587600 579900 249900 66680 268900 615800 Dehydroepiandrosterone 52890 82330 34820 46780 53950 39110 68970 45550 56170 88760 Aldosterone 19760 11200 5400 5662 2726 3525 1520 1067 2153 3463 Testosterone 12590 19000 18790 22940 42250 9493 15490 29850 21780 29890 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name 11-deoxycortisol Pregnenolone Progesterone Dihydrotestosterone Androstenedione Melatonin 17α-hydroxypregnenolone Corticosterone Cortisol Dehydroepiandrosterone Aldosterone Testosterone METABOLITES_END #END