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ftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14245/m_MTBLS14245_LC-MS_alternating_hilic_v2_maf.tsvftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14245/a_MTBLS14245_LC-MS_alternating_hilic.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14245/i_Investigation.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14245/s_MTBLS14245.txtprimaryOK200ftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14245Metabolite identification was performed using the MWDB database constructed from authentic standards. Quantification was conducted by MRM on a triple quadrupole MS, where Q1 selects precursor ions, the collision cell generates fragments, and Q3 filters characteristic product ions for specific and reproducible quantification. Calibration curves were used for absolute quantification.Quantification was performed by substituting sample peak areas into the linear standard curve equation. The actual metabolite content (ng/tube) was calculated using the formula: c × v / (n × 1000), where c represents the concentration from the standard curve (ng/mL), v is the extraction volume (μL), and n is the sample amount (tube). The formula includes integrated unit conversion.MetaboLightsPublicLiquid Chromatography MS - alternating - hilicThe sample extracts were analyzed using an LC-ESI-MS/MS system ((Waters ACQUITY H-ClassD, https://www.waters.com/nextgen/cn/zh.html; MS, QTRAP 6500+ System, https://sciex.com /). The analytical conditions were as follows.Amide method: HPLC: column, ACQUITY UPLC BEH Amide (i.d.2.1×100 mm, 1.7 μm); solvent system, water with 10mM Ammonium acetate and 0.3% Ammonium hydroxide (A), 90% acetonitrile/water (V/V)(B); The gradient was started at 95% B (0-1.2 min), decreased to 70% B (8 min),50% B (9-11 min), finaly ramped back to 95% B (11.1-15 min); flow rate, 0.4 mL/min; temperature, 40°C; injection volume: 2 μL.Cross-Species Mitochondrial Rescue by Gastrodia elata-Derived Extracellular Vesicles Protects the Stressed Mammalian Heart.Ziyi YinHuazhong Agricultural UniversityHeartplantmass spectrometry assayThe sample was thawed on ice, then mixed with methanol/water (precooled at -20°C) and vortexed for 2 min under the condition of 2500 r/min. The sample was frozenin liquid nitrogen for 5 min, removed on ice for 5 min, after that, the sample was vortexed for 2 min.The previous step was repeated for 3 times. The sample was centrifuged at 12000 r/min for 10 min at 4°C. Take 300 μL of supernatant into a new centrifuge tube and place the supernatant in -20°C refrigerator for 30 min.Then the supernatant was centrifuged at 12000 r/min for 10 min at 4°C. After centrifugation, transfer supernatant through Protein Precipitation Plate for further LC-MS analysis.Mus musculusGastrodia elataRattus norvegicushttps://www.ebi.ac.uk/metabolights/MTBLS14245Ziyi Yin. College of Biomedicine and Health, College of Life science and Technology, Huazhong Agricultural University. 2411612041@qq.com.Min Zhang. College of Biomedicine and Health, College of Life science and Technology, Huazhong Agricultural University. minzhang@mail.hzau.edu.cn.Data acquisitions were performed using Analyst 1.6.3 software (Sciex). Multiquant 3.0.3 software (Sciex) was used to quantify all metabolites. Mass spectrometer parameters including the declustering potentials (DP) and collision energies (CE) for individual MRM transitions were done with further DP and CE optimization. A specific set of MRM transitions were monitored for each period according to the metabolites eluted within this period.Group2411612041@qq.com2025.07.06: Fresh Gastrodia elata was washed, peeled, and then used as the source of plant-derived extracellular vesicles. The juice was filtered after extraction using a household high-speed blender. Briefly, the filtered juice of Gastrodia elata was sequentially centrifuged using an Allegra X-30R centrifuge (Beckman Coulter, Inc., Brea, CA, USA) at 1000 × g for 10 min, 3000 × g for 20 min, 10,000 × g for 60 min at 4 C to remove large particles and cellular debris. The supernatant was subjected to ultracentrifugation using a Beckman Optima XE-100 ultracentrifuge (Beckman Coulter, Inc., Brea, CA, USA) at 150,000 × g for 90 min at 4 C. Then, the supernatant was removed, and the pellet was resuspended in 1× PBS. The samples were concentrated using 100 kDa molecular weight cutoff ultrafiltration tubes. The resulting pellet was filtered through a 0.22 µm membrane and aliquoted for storage under sterile conditions at least 1 h at 4 C. Final samples of Gastrodia elata-derived extracellular vesicles (GE-EVs) was aliquoted, snap frozen in liquid nitrogen, and stored at 80 C until analysis.2025.07.18: H9c2 cells were obtained from China Center for Type Culture Collection (CCTCC), and cultured in DMEM (Gibco) supplemented with 10% fetal bovine serum (FBS) (NEWZERUM), 100 units/ml penicillin-streptomycin (Gibco, 15140-122). Cells were cultivated at 37 °C in 5% CO2 humidity. Gastrodia elata-derived extracellular vesicles (GE-EVs) were pre-incubated with cardiomyocytes for 48 h, followed by exposure to hypoxic conditions (0.5% O2, 2 h). Cells were then digested, trypsin and culture medium were discarded, and cell pellets were snap-frozen in liquid nitrogen and stored at -80°C until analysis.2025.11: A/A Mice were pre-treated with epinephrine and subjected to 21 days of social isolation (SI) or group housing (GH). GE-EVs or PBS (Ctrl) were administered every 3 days. Fresh hearts were excised, snap-frozen in liquid nitrogen, and stored at -80°C.MetabolomicsHeartMetabolomicsMus musculusHeart tissue70-1000Waters ACQUITY UPLC H-Class Systemtargeted analysisRattus norvegicusmzminetargeted metabolite profilingcellcardiovascular disordervesiclesAB SCIEX QTRAP 6500+Gastrodia elataplantHeartMetabolomicsHeart tissueMus musculus70-1000Waters ACQUITY UPLC H-Class Systemtargeted analysisRattus norvegicusmzminetargeted metabolite profilingcellcardiovascular disordervesiclesAB SCIEX QTRAP 6500+Gastrodia elataplantLinear ion trap (LIT) and triple quadrupole (QQQ) scans were acquired on a triple quadrupole-linear ion trap mass spectrometer (QTRAP), QTRAP 6500+ LC-MS/MS System, equipped with an ESI Turbo Ion-Spray interface, operating in both positive and negative ion mode and controlled by Analyst 1.6.3 software (Sciex). The ESI source operation parameters were as follows: ion source, ESI+/-; source temperature 550 ; ion spray voltage (IS) 5500 V(Positive), -4500 V(Negative); curtain gas (CUR) was set at 35 psi, respectively. Energy and its metabolites were analyzed using scheduled multiple reaction monitoring (MRM).6-Phosphogluconic-acidSuccinyl-CoASuccinic-AcidLactateGuanosineFructose-1,6-bisphosphatec-di-AMPOxaloacetateDL-Glyceric-AcidGlucuronic-acidADPNicotinaMide-adenine-dinucleotide(NAD)GluconatePhosphoenolpyruvic-acidL-AlanineD-Ribose-5-phosphate-disodiumItaconic-acid2-Phospho-D-glycerateAlpha-Ketoglutaric-Acid2-Oxoadipic-acidTyrosineL-LeucineThreonineAMPL-CystineFlavin-mononucleotideD-Fructose-6-phosphateD-Erythrose-4-phosphate3-phenyllactic-acidGlyceraldehyde-3-phosphateIMPD-Ribulose-5-phosphateArgininosuccinic-acidPhosphorylethanolamineLysineL-Glutamic-acidFumaric-acidUMPD-Glucose-1-phosphateOrnithinedTMPTrehalose-6-phosphateCitric-acidSerineD-Glucose-6-phosphateL-2-Hydroxyglutaric-acid-disodiumCyclic-AMPL-AsparagineDihydroxyacetone-phosphateUDP-GlcNAcMalic-acidD(+)-GlucoseL-citrullineGlycerol-3-phosphateArginineD-ribulose-1,5-bisphosphate2,3-DPGIsocitric-acidGlycolic-acidPyruvic-acidSedoheptulose-7-phosphatecis-Aconitic-acidAdenineAcetyl-CoAD-Mannose-6-phosphatedCMPdAMPCysteic-acidXylulose-5-phosphateTriphosphate-guanosine3-phosphoglycerateInosinedUMPDihydronicotinamide-adenine-dinucleotide-phosphate(NADPH)L-AspartateUreidopropionateUracilATPGuanosine-diphosphateGlutaminefalseCross-Species Mitochondrial Rescue by Gastrodia elata-Derived Extracellular Vesicles Protects the Stressed Mammalian HeartIn this study, we developed Gastrodia elata (G. elata)-derived extracellular vesicles (GE-EVs) to achieve functional rescue of the stressed mammalian heart. Using RBM24 phosphorylation-deficient (S181A, A/A) mice, we demonstrated that GE-EVs have a profound cross-species rescue effect on mitochondria. GE-EVs therapy restores mitochondrial integrity and ATP synthesis through mitochondrial quality control, thereby reversing myocardial hypoxia and attenuating ROS production.2026-04-092026-04-09MTBLS14245