MetaboLightsapplication/xmlftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/m_MTBLS15144_LC-MS_positive_reversed-phase-chromatography_v2_maf.tsvftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/m_MTBLS15144_LC-MS_negative_reversed-phase-chromatography_v2_maf.tsvftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/s_MTBLS15144.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/i_Investigation.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/a_MTBLS15144_LC-MS_negative_reversed-phase-chromatography.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/a_MTBLS15144_LC-MS_positive_reversed-phase-chromatography.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/CAP3.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/CAP3.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/control2.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/EAU5.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/EAU5.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/control3.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/EAU4.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/control3.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/CAP2.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/CAP2.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/EAU2.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/CAP6.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/EAU4.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/EAU3.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/control1.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/control6.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/control4.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/CAP5.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/CAP5.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/EAU3.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/CAP1.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/control5.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/control2.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/EAU2.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/CAP1.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/CAP6.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/control5.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/EAU6.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/CAP4.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/CAP4.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/EAU6.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/control1.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/neg/control4.rawftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144/FILES/RAW_FILES/pos/control6.rawprimaryOK200ftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS15144<p>Metabolite identification was performed based on the PSNGM (PerSonalbio Next-Generation Metabolomics Database) developed by Personalbio. The database integrates multiple spectral libraries, including an in-house standard compound library, mzCloud (https://www.mzcloud.org/), LIPID MAPS (https://www.lipidmaps.org/), Human Metabolome Database (HMDB) (https://hmdb.ca/), MoNA (https://mona.fiehnlab.ucdavis.edu/), NIST 2020 MS/MS library, and an AI-predicted MS/MS spectral library.</p><p>Metabolite annotation was performed by spectral matching using the following parameters: MS1 tolerance for identification: 0.01 Da; MS2 tolerance for identification: 0.05 Da; smoothing level: 3; minimum peak height: 10,000; minimum peak width: 5; mass slice width: 0.05 Da; identification score cutoff: 60/70.</p>MetaboLightsPublicLiquid Chromatography MS - negative - reversed-phase-chromatographyLiquid Chromatography MS - positive - reversed-phase-chromatography<p>Mass spectrometric data acquisition was performed using a Thermo Q Exactive HF-X mass spectrometer (Thermo Fisher Scientific) controlled by Xcalibur software (version 4.7, Thermo Fisher Scientific). Data were collected in both positive and negative ionization modes using a data-dependent acquisition (DDA) strategy.</p><p>The instrument was equipped with a heated electrospray ionization (HESI) source. The spray voltage was set at 3.5 kV, with sheath gas and auxiliary gas flow rates of 40 and 10 arbitrary units (arb), respectively. The capillary temperature was maintained at 320°C, and the auxiliary gas heater temperature was set at 300°C.</p><p>Full-scan MS spectra were acquired at a resolution of 60,000 over an m/z range of 70–1000. The automatic gain control (AGC) target was set to Standard, with a maximum injection time (Max IT) of 100 ms. The top 10 most intense precursor ions were selected for MS/MS fragmentation. The dynamic exclusion time was set to 4 s. MS/MS spectra were acquired at a resolution of 15,000 using higher-energy collisional dissociation (HCD) with a normalized collision energy (NCE) of 30%. The AGC target was set to Standard, and the maximum injection time was set to Auto.</p><p>All study samples and quality control (QC) samples were analyzed under the same LC–MS conditions. Before sample acquisition, 2–4 QC injections were performed to equilibrate and stabilize the LC–MS system. During the analytical sequence, QC samples were injected every 6–12 study samples to monitor instrument performance and data quality. For datasets containing 11 or fewer samples, QC samples were not prepared.</p>High-dose capsaicin exacerbates uveitis by disrupting intestinal ho-meostasis and upregulating IL-17.Jilin Universityliu taofecesmass spectrometry assay<p>Approximately 20 mg of fecal sample was weighed and transferred into a 2 mL centrifuge tube.</p><p>A total of 300 μL of pre-chilled methanol containing 5 ppm 2-chlorophenylalanine (internal standard) was added to each sample, followed by the addition of two steel beads. The samples were vortexed for 30 s.</p><p>Samples were homogenized using a high-throughput tissue grinder at 55 Hz for 60 s. This homogenization step was repeated once.</p><p>The samples were then sonicated in an ultrasonic cleaning bath for 10 min.</p><p>Samples were incubated at −20°C for 30 min to facilitate protein precipitation.</p><p>The extracts were centrifuged at 12,000 rpm for 10 min at 4°C. The supernatants were collected and filtered through a 0.22 μm membrane filter. The filtered extracts were transferred into sample vials for LC–MS analysis.</p><p>Quality control (QC) samples were prepared by mixing 10–20 μL of each filtered sample extract. For studies containing 11 or fewer samples, QC samples were not prepared. QC samples were used to evaluate instrument stability and data reliability during metabolomic analysis.</p>Mus musculushttps://www.ebi.ac.uk/metabolights/MTBLS15144tao liu. Jilin University. 1224791633@qq.com.xiaoli liu. jilin University. Department of Ophthalmology, The Second Hospital of Jilin University 218 Ziqiang Street, Changchun, Jilin 130041, China. lpw_lxl@126.com.<p>Raw LC–MS data files in .raw format were imported into MS-DIAL software (version 4.9.221218) for data processing. Peak detection, peak alignment, peak filtering, and metabolite annotation were performed using MS-DIAL. Features with missing values exceeding 50% within each experimental group were removed. Missing values of undetected features were imputed, followed by data normalization for subsequent quantitative analysis.</p>Disease1224791633@qq.com<p>Fecal samples were collected from experimental mice on day 21 after induction of experimental autoimmune uveitis (EAU). Mice were randomly assigned to different experimental groups, including Control, EAU, EAU treated with capsaicin, and EAU treated with capsaicin combined with kynurenic acid groups. Fresh fecal pellets were collected individually from each mouse under sterile conditions and immediately frozen in liquid nitrogen. Samples were stored at −80°C until further metabolomic analysis. All samples were collected at the same time point to minimize circadian and environmental variations.</p>MetabolomicsMetabolomicsProteoWizard msconvertMus musculusControlQ Exactive HF-Xuntargeted analysisThermo Scientific Vanquish Flex UHPLC SystemUveitisEAUexperimental blankfecesMetabolomicsProteoWizard msconvertControlMus musculusQ Exactive HF-Xuntargeted analysisThermo Scientific Vanquish Flex UHPLC SystemUveitisEAUexperimental blankfeces<p>Mass spectrometric data acquisition was performed using a Thermo Q Exactive HF-X mass spectrometer (Thermo Fisher Scientific) controlled by Xcalibur software (version 4.7, Thermo Fisher Scientific). Data were acquired in both positive and negative electrospray ionization (ESI) modes using a data-dependent acquisition (DDA) strategy.</p><p>The mass spectrometer was equipped with a heated electrospray ionization (HESI) source. The spray voltage was set at 3.5 kV, with sheath gas and auxiliary gas flow rates of 40 and 10 arbitrary units (arb), respectively. The capillary temperature was maintained at 320°C, and the auxiliary gas heater temperature was set at 300°C.</p><p>For full-scan MS acquisition, the resolution was set to 60,000, with a scan range of m/z 70–1000. The automatic gain control (AGC) target was set to Standard, and the maximum injection time (Max IT) was set to 100 ms. The top 10 most intense ions were selected for MS/MS fragmentation. The dynamic exclusion time was set to 4 s. MS/MS spectra were acquired at a resolution of 15,000 using higher-energy collisional dissociation (HCD) with a normalized collision energy (NCE) of 30%. The AGC target was set to Standard, and the maximum injection time was set to Auto.</p><p>All experimental samples and quality control (QC) samples were analyzed using the same chromatographic and mass spectrometric conditions described above. Prior to sample acquisition, 2–4 QC injections were performed to equilibrate and stabilize the LC–MS system. During the analytical sequence, a QC sample was injected every 6–12 experimental samples to monitor instrument stability and data quality. For studies containing 11 or fewer samples, QC samples were not prepared. QC samples were used for subsequent data evaluation and quality control.</p>falseHigh-dose capsaicin exacerbates uveitis by disrupting intestinal ho-meostasis and upregulating IL-17Purpose: To determine whether capsaicin (CAP) exacerbates experimental autoimmune uveitis (EAU) and whether this effect is associated with impaired intestinal homeostasis, gut microbiota re-modeling, and altered metabolic profiles. Methods: An EAU mouse model was used to evaluate the dose-dependent effects of CAP on ocular inflammation and intestinal homeostasis. EAU severity was assessed by fundus examination, clinical scoring, and retinal histopathology. Ileal and colonic morphology, colonic ZO-1 and MUC2 expression, and IL-17A levels in the colon and spleen were evaluated using hematox-ylin and eosin staining, immunofluorescence, and enzyme-linked immunosorbent assays. 16S rRNA gene sequencing, untargeted fecal metabolomics, and colonic RNA sequencing were performed to characterize CAP-associated changes in the gut microbiota, metabolome, and transcriptome. Kynurenic acid (KYNA) was subsequently administered to determine whether supplementation attenuated CAP-induced exacerbation of EAU and intestinal barrier injury. Results: CAP exacerbated EAU in a dose-dependent manner. At 80 mg/kg, CAP aggravated retinal inflammation and tissue injury, increased clinical and histopathologic scores, and elevated IL-17A levels in the colon and spleen. High-dose CAP was also associated with ileal archi-tectural abnormalities, reduced colonic mucosal thickness, and decreased colonic ZO-1 and MUC2 expression. 16S rRNA gene sequencing revealed alterations in gut microbial diversity and community structure. CAP-associated differential metabolites were enriched in the tryp-tophan metabolism pathway, and fecal KYNA levels were significantly reduced. Colonic RNA sequencing showed enrichment of the IL-17 signaling pathway, upregulation of Il17a, Il17f, and Rorc, and altered expression of genes involved in tryptophan metabolism. KYNA supplementation reduced clinical and histopathologic scores, decreased IL-17A levels in the colon and spleen, and partially restored colonic ZO-1 and MUC2 expression. Conclusion: Under these experimental conditions, high-dose CAP exacerbated EAU and was associat-ed with intestinal barrier disruption, gut microbiota remodeling, altered tryptophan metabo-lism, and enhanced IL-17-related inflammation. The partial attenuation observed after KYNA supplementation supports further investigation of KYNA as a candidate metabolic factor in high-dose CAP-treated EAU mice.2026-07-232026-07-23MTBLS15144