{ "METABOLOMICS WORKBENCH":{"STUDY_ID":"ST001150","ANALYSIS_ID":"AN001897","VERSION":"1","CREATED_ON":"March 19, 2019, 12:08 pm"}, "PROJECT":{"PROJECT_TITLE":"Metabolomics of a Mouse Model for Retinitis Pigmentosa","PROJECT_SUMMARY":"Retinitis pigmentosa (RP) is a degenerative disease of the retina that affects approximately 1 million people worldwide. There are multiple genetic causes of this disease, for which, at present, there are no effective therapeutic strategies. We utilized broad spectrum metabolomics to identify perturbations in the metabolism of the rd10 mouse, a genetic model for RP. C57BL/6J and rd10 mice were raised in cyclic light followed by either light or dark adaptation at postnatal day (P) 18, an early stage in the degeneration process. Mice raised entirely in the dark until P18 were also evaluated. After euthanasia, retinas were removed and extracted for analysis by ultra-performance liquid chromatography-time of flight-mass spectrometry (UPLC-QTOF-MS).","INSTITUTE":"University of North Carolina at Chapel Hill","LABORATORY":"ERCMRC","LAST_NAME":"Sumner","FIRST_NAME":"Susan","ADDRESS":"NRI, 500 Laureate Way, Kannapolis, NC, 28081, USA","EMAIL":"susan_sumner@unc.edu","PHONE":"704-250-5066","FUNDING_SOURCE":"NIDDK U24DK097193 (Sumner, PI); Supplement to NEI R01EY12224 (Weiss, PI)"}, "STUDY":{"STUDY_TITLE":"Metabolomics of a Mouse Model for Retinitis Pigmentosa","STUDY_SUMMARY":"Retinitis pigmentosa (RP) is a degenerative disease of the retina that affects approximately 1 million people worldwide. There are multiple genetic causes of this disease, for which, at present, there are no effective therapeutic strategies. We utilized broad spectrum metabolomics to identify perturbations in the metabolism of the rd10 mouse, a genetic model for RP. C57BL/6J and rd10 mice were raised in cyclic light followed by either light or dark adaptation at postnatal day (P) 18, an early stage in the degeneration process. Mice raised entirely in the dark until P18 were also evaluated. After euthanasia, retinas were removed and extracted for analysis by ultra-performance liquid chromatography-time of flight-mass spectrometry (UPLC-QTOF-MS).","INSTITUTE":"University of North Carolina at Chapel Hill","DEPARTMENT":"Department of Cell Biology & Physiology","LAST_NAME":"Weiss","FIRST_NAME":"Ellen","ADDRESS":"CB# 7545, 5340B MBRB, 111 Mason Farm Rd. Chapel Hill, NC 27599-7090","EMAIL":"erweiss@med.unc.edu","PHONE":"919-966-7683"}, "SUBJECT":{"SUBJECT_TYPE":"Mammal","SUBJECT_SPECIES":"Mus musculus","TAXONOMY_ID":"10090","GENOTYPE_STRAIN":"rd10 and C57BL/6J","AGE_OR_AGE_RANGE":"P18","ANIMAL_ANIMAL_SUPPLIER":"Jackson Laboratory"}, "SUBJECT_SAMPLE_FACTORS":[ { "Subject ID":"-", "Sample ID":"All_pool_QC1", "Factors":{"Condition":"QC Pool"} }, { "Subject ID":"-", "Sample ID":"All_pool_QC2", "Factors":{"Condition":"QC Pool"} }, { "Subject ID":"-", "Sample ID":"All_pool_QC3", "Factors":{"Condition":"QC Pool"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-L-100913-RT-1", "Factors":{"Condition":"RD10-RCL-L"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-L-110113-RT-1", "Factors":{"Condition":"RD10-RCL-L"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-L-110113-RT-2", "Factors":{"Condition":"RD10-RCL-L"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-L-110913-RT-1", "Factors":{"Condition":"RD10-RCL-L"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-L-110913-RT-2", "Factors":{"Condition":"RD10-RCL-L"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-L-111013-RT-1", "Factors":{"Condition":"RD10-RCL-L"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-D-100913-RT-2", "Factors":{"Condition":"RD10-RCL-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-D-110113-RT-3", "Factors":{"Condition":"RD10-RCL-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-D-110913-RT-3", "Factors":{"Condition":"RD10-RCL-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-D-111013-RT-2", "Factors":{"Condition":"RD10-RCL-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RCL-D-111013-RT-3", "Factors":{"Condition":"RD10-RCL-D"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-L-101513-RT-1", "Factors":{"Condition":"WT-RCL-L"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-L-101513-RT-2", "Factors":{"Condition":"WT-RCL-L"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-L-111013-RT-1", "Factors":{"Condition":"WT-RCL-L"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-L-111013-RT-2", "Factors":{"Condition":"WT-RCL-L"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-L-111113-RT-1", "Factors":{"Condition":"WT-RCL-L"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-L-111713-RT-1", "Factors":{"Condition":"WT-RCL-L"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-D-101513-RT-3", "Factors":{"Condition":"WT-RCL-D"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-D-111013-RT-3", "Factors":{"Condition":"WT-RCL-D"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-D-111113-RT-2", "Factors":{"Condition":"WT-RCL-D"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-D-111113-RT-3", "Factors":{"Condition":"WT-RCL-D"} }, { "Subject ID":"-", "Sample ID":"WT-RCL-D-111713-RT-2", "Factors":{"Condition":"WT-RCL-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RD-D-120513-RT-1", "Factors":{"Condition":"RD10-RD-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RD-D-120513-RT-2", "Factors":{"Condition":"RD10-RD-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RD-D-120813-RT-1", "Factors":{"Condition":"RD10-RD-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RD-D-120813-RT-2", "Factors":{"Condition":"RD10-RD-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RD-D-120813-RT-3", "Factors":{"Condition":"RD10-RD-D"} }, { "Subject ID":"-", "Sample ID":"RD10-RD-D-120813-RT-4", "Factors":{"Condition":"RD10-RD-D"} }, { "Subject ID":"-", "Sample ID":"WT-RD-D-011614-RT-1", "Factors":{"Condition":"WT-RD-D"} }, { "Subject ID":"-", "Sample ID":"WT-RD-D-011614-RT-2", "Factors":{"Condition":"WT-RD-D"} }, { "Subject ID":"-", "Sample ID":"WT-RD-D-120513-RT-1", "Factors":{"Condition":"WT-RD-D"} }, { "Subject ID":"-", "Sample ID":"WT-RD-D-120513-RT-2", "Factors":{"Condition":"WT-RD-D"} }, { "Subject ID":"-", "Sample ID":"WT-RD-D-121213-RT-1", "Factors":{"Condition":"WT-RD-D"} }, { "Subject ID":"-", "Sample ID":"WT-RD-D-121213-RT-2", "Factors":{"Condition":"WT-RD-D"} } ], "COLLECTION":{"COLLECTION_SUMMARY":"C57BL/6J and rd10 mice were raised under varying light conditions in standard housing, temperature and access to food and water. Some mice were raised under normal light/dark cycle at ambient light levels of approximately 80 lux. At 18 days after birth, mice were exposed to 250 lux for 3 h (light-adapted) followed by euthanasia using cervical dislocation. The eyes were enucleated and the retinas dissected in PBS in the light under a microscope, followed by rapid freezing by dropping them individually into a container filled with liquid nitrogen. Each set of samples were pooled in liquid nitrogen. Other mice raised in cyclic light were adapted overnight at P17 in complete darkness (dark-adapted) before euthanasia at P18. Using infrared goggles and an infrared microscope, these retinas were processed similarly as light-adapted samples except that all procedures were performed in the dark. Mice raised in total darkness were euthanized at P18 and dissected in the dark using infrared goggles and an infrared microscope.","SAMPLE_TYPE":"Retina"}, "TREATMENT":{"TREATMENT_SUMMARY":"C57BL/6J and rd10 mice were raised under varying light conditions in standard housing, temperature and access to food and water. Some mice were raised under normal light/dark cycle at ambient light levels of approximately 80 lux. At 18 days after birth, mice were exposed to 250 lux for 3 h (light-adapted) followed by euthanasia. Other mice raised in cyclic light were adapted overnight at P17 in complete darkness (dark-adapted) before euthanasia at P18. Mice raised in total darkness were euthanized at P18."}, "SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"Each sample was homogenized in buffer containing 50:50 acetonitrile:water at a ratio of 1mg:25 µl tissue:buffer. A 320 µl aliquot of homogenate (12.8 mg tissue equivalent) was transferred to a tube and centrifuged. The supernatant was dried and re-suspended in 100 µl 95:5 H2O:methanol for UPLC-TOF-MS analysis."}, "CHROMATOGRAPHY":{"CHROMATOGRAPHY_SUMMARY":"Reversed-Phase Gradient Seperation","CHROMATOGRAPHY_TYPE":"Reversed phase","INSTRUMENT_NAME":"Waters Acquity","COLUMN_NAME":"Waters Acquity BEH HSS T3 (100 x 2.1mm, 1.8um)","FLOW_GRADIENT":"Time(min) Flow Rate(mL/min) %A %B Curve 1. Initial 0.400 99.0 1.0 6 2. 1.00 0.400 99.0 1.0 6 3. 16.00 0.400 1.0 99.0 6 4. 20.00 0.400 1.0 99.0 6 5. 20.50 0.400 99.0 1.0 6 6. 22.00 0.400 99.0 1.0 6","FLOW_RATE":"0.4 mL/min","COLUMN_TEMPERATURE":"50 °C","COLUMN_PRESSURE":"6,000-10,000 psi","INJECTION_TEMPERATURE":"8 °C","INTERNAL_STANDARD":"L-Tryptophan-d5"}, "ANALYSIS":{"ANALYSIS_TYPE":"MS"}, "MS":{"INSTRUMENT_NAME":"Waters Synapt G2 Si QTOF","INSTRUMENT_TYPE":"QTOF","MS_TYPE":"ESI","ION_MODE":"NEGATIVE","MS_COMMENTS":"MassLynx 4.1","CAPILLARY_TEMPERATURE":"110 °C","CAPILLARY_VOLTAGE":"1.0 kV","COLLISION_ENERGY":"4","FRAGMENTATION_METHOD":"CID","HELIUM_FLOW":"180","IONIZATION":"ES-","MASS_ACCURACY":"5 ppm","SOURCE_TEMPERATURE":"110 °C","DATAFORMAT":"Continuum","DESOLVATION_GAS_FLOW":"800 L/Hr","DESOLVATION_TEMPERATURE":"400 °C","RESOLUTION_SETTING":"20000","SCAN_RANGE_MOVERZ":"50-1000 m/z","SCANNING_CYCLE":"1.0 s","TUBE_LENS_VOLTAGE":"75","MS_RESULTS_FILE":"ST001150_AN001897_Results.txt UNITS:m/z_retention time Has m/z:Yes Has RT:Yes RT units:Minutes"} }