{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST000848","ANALYSIS_ID":"AN001372","VERSION":"1","CREATED_ON":"August 10, 2017, 3:11 pm"},

"PROJECT":{"PROJECT_TITLE":"Mayo Pilot and Feasiblity: Targeting Myelin Metabolism of Kallikrein 6 Signals through PAR1 and PAR2 to Enchance Recovery of Function after SCI","PROJECT_SUMMARY":"CNS trauma generates a proteolytic imbalance contributing to secondary injury, including axonopathy and neuron degeneration. Kallikrein 6 (Klk6) is a serine protease implicated in neurodegeneration and here we investigate the role of protease activated receptors 1 (PAR1) and PAR2 in mediating these effects in mice with spinal cord injury (SCI). This project conducts untargeted Profiling (unbiased metabolomics assay) and targeted lipid analysis. The lipid assays are 1) free fatty acid composition of lipids; 2) free fatty acid panel; 3) cholesterol concentration (free and bound); 4) Ceramides, including galactosyl and glucosyl; 5) sphingomyelin.","INSTITUTE":"Mayo Clinic","LAST_NAME":"Scarisbrick","FIRST_NAME":"Isobel","ADDRESS":"200 First St. SW, Rochester, Minnesota, 55905, USA","EMAIL":"scarisbrick.isobel@mayo.edu","PHONE":"507-284-0124"},

"STUDY":{"STUDY_TITLE":"Targeting Myelin NEFA of Kallikrein 6 Signals through PAR1 and PAR2 to Enchance Recovery of Function after SCI","STUDY_SUMMARY":"Targeting mouse myelin NEFA of Kallikrein 6 Signals through PAR1 and PAR2 to Enchance Recovery of Function after SCI. The samples submitted are purified myelin preparations from the postnatal day 21, 60, or 90 mouse spinal cord (SC). There are 9 samples total in Project 1, n=3 for each genotype (control, PAR1 or PAR2) at 21 days. There are 9 samples total in Project 2, n=3 for each genotype (WT, PAR1-/- or PAR2-/-) at 60 days. There are 12 samples total in Project 3, n=3 for K6+/+ or K6-/- at either P21 or P90.","INSTITUTE":"Mayo Clinic","LAST_NAME":"Scarisbrick","FIRST_NAME":"Isobel","ADDRESS":"200 First St. SW, Rochester, Minnesota, 55905, USA","EMAIL":"scarisbrick.isobel@mayo.edu","PHONE":"507-284-0124"},

"SUBJECT":{"SUBJECT_TYPE":"Mouse","SUBJECT_SPECIES":"Mus musculus","TAXONOMY_ID":"10090"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"-",
"Sample ID":"ms5959-1",
"Factors":{"time pt":"P21d","grouping":"CTRL"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.81"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-2",
"Factors":{"time pt":"P21d","grouping":"CTRL"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.57"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-3",
"Factors":{"time pt":"P21d","grouping":"CTRL"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.99"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-4",
"Factors":{"time pt":"P21d","grouping":"PAR1"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.42"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-5",
"Factors":{"time pt":"P21d","grouping":"PAR1"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"2.15"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-6",
"Factors":{"time pt":"P21d","grouping":"PAR1"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.87"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-7",
"Factors":{"time pt":"P21d","grouping":"PAR2"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.83"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-8",
"Factors":{"time pt":"P21d","grouping":"PAR2"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.21"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-9",
"Factors":{"time pt":"P21d","grouping":"PAR2"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"2.20"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-10",
"Factors":{"time pt":"P60d","grouping":"Wt"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.55"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-11",
"Factors":{"time pt":"P60d","grouping":"Wt"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.73"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-12",
"Factors":{"time pt":"P60d","grouping":"Wt"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"2.49"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-13",
"Factors":{"time pt":"P60d","grouping":"PAR2-/-"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.86"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-14",
"Factors":{"time pt":"P60d","grouping":"PAR2-/-"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"1.48"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-15",
"Factors":{"time pt":"P60d","grouping":"PAR2-/-"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"2.70"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-16",
"Factors":{"time pt":"P60d","grouping":"PAR1-/-"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"2.94"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-17",
"Factors":{"time pt":"P60d","grouping":"PAR1-/-"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"2.61"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-18",
"Factors":{"time pt":"P60d","grouping":"PAR1-/-"},
"Additional sample data":{"gender":"Female","protein (ug/ul)":"2.52"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-19",
"Factors":{"time pt":"P21d","grouping":"K6+/+"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"2.61"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-20",
"Factors":{"time pt":"P21d","grouping":"K6+/+"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"2.48"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-21",
"Factors":{"time pt":"P21d","grouping":"K6+/+"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"1.93"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-22",
"Factors":{"time pt":"P21d","grouping":"K6-/-"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"1.72"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-23",
"Factors":{"time pt":"P21d","grouping":"K6-/-"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"2.16"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-24",
"Factors":{"time pt":"P21d","grouping":"K6-/-"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"2.47"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-25",
"Factors":{"time pt":"P90d","grouping":"K6+/+"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"2.70"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-26",
"Factors":{"time pt":"P90d","grouping":"K6+/+"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"2.57"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-27",
"Factors":{"time pt":"P90d","grouping":"K6+/+"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"2.93"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-28",
"Factors":{"time pt":"P90d","grouping":"K6-/-"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"3.16"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-29",
"Factors":{"time pt":"P90d","grouping":"K6-/-"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"2.96"}
},
{
"Subject ID":"-",
"Sample ID":"ms5959-30",
"Factors":{"time pt":"P90d","grouping":"K6-/-"},
"Additional sample data":{"gender":"Male","protein (ug/ul)":"3.88"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"The samples submitted are purified myelin preparations from the postnatal day 21 or 90 mouse spinal cord (SC). There are two projects but these can be run together since the down stream assays are identical. There are 9 samples total in Project 1, n=3 for each genotype (WT, PAR1-/- or PAR2-/-). There are 12 samples total in Project 2, n=3 for K6+/+ or K6-/- at either P21 or P90. We are submitting these samples for analysis of 1) free fatty acid panel; 2) free fatty acid composition of lipids; 3) cholesterol concentration (free and bound); 4) Ceramides, including galactosyl and glucosyl; 5) sphingomyelin. A total of approximately 130 to 150 ul for each sample is submitted with the protein concentrations already measured by BCA assay and indicated below. Concentrations are provided in ug/ul and the total volume is also indicated. One tube for each sample is submitted but this can be shared across the assays."},

"TREATMENT":{"TREATMENT_SUMMARY":"A 3g Clip produces moderate SCI including demyelination and clinical impairment and we recently published a detailed methodology. At 1 week after injury, the 3g injured mice are expected to have an average Basso Mouse Scale score (BMS)=5 on a 9 point scale such that they have frequent plantar stepping with no or some coordination. This level of impairment was chosen to provide a sufficient window to observe recovery."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"NEFA of mouse spinal cord Lipids will be quantified in myelin isolated in high yield and purity by subcellular fractionation from the lumbosacral spinal cord. While there are no absolutely ‘myelin-specific’ lipids, galactocerebroside is the most typical of myelin in the adult nervous system being directly proportional to the amount of myelin. Sulfatide is another galactolipid enriched in myelin. Together with cholesterol, these form 78% of the total amount of lipid in the myelin membrane and each will be quantified using LC/MS/MS. A highly sensitive assay for galactocerebroside was recently established by the Mayo Metabolomics Core and can be implemented immediately. The LC/MS/MS panel for free fatty acids, including the very long chain fatty acids found in myelin is also routinely performed by the Core. Cholesterol will be quantified using an NMR-based approach by the Mayo Dept. of Laboratory Medicine Clinical Core. Additionally, we have a plan in place with the Metabolomics Core to develop LC/MS/MS assays for sulfatide and sphingomyelin during the Pilot proposal. Having quantitative assays for each of these key myelin lipids will facilitate our goal to comprehensively profile myelin lipid metabolism and will form foundational assays for a future NIH grant focused on myelin metabolism."},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_TYPE":"Reversed phase","INSTRUMENT_NAME":"Agilent 1290 Infinity","COLUMN_NAME":"Waters Acquity BEH C18 (150 x 2.1mm, 1.7um)"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS"},

"MS":{"INSTRUMENT_NAME":"Agilent 6460 QQQ","INSTRUMENT_TYPE":"Triple quadrupole","MS_TYPE":"ESI","ION_MODE":"NEGATIVE"},

"MS_METABOLITE_DATA":{
"Units":"ng/ug of protein",

"Data":[{"Metabolite":"EPA","ms5959-1":"0.17","ms5959-2":"0.17","ms5959-3":"0.18","ms5959-4":"0.23","ms5959-5":"0.20","ms5959-6":"0.18","ms5959-7":"0.16","ms5959-8":"0.13","ms5959-9":"0.16","ms5959-10":"0.13","ms5959-11":"0.11","ms5959-12":"0.07","ms5959-13":"0.07","ms5959-14":"0.08","ms5959-15":"0.06","ms5959-16":"0.08","ms5959-17":"0.09","ms5959-18":"0.11","ms5959-19":"0.10","ms5959-20":"0.13","ms5959-21":"0.12","ms5959-22":"0.16","ms5959-23":"0.15","ms5959-24":"0.17","ms5959-25":"0.06","ms5959-26":"0.07","ms5959-27":"0.06","ms5959-28":"0.05","ms5959-29":"0.06","ms5959-30":"0.08"},{"Metabolite":"linolenic","ms5959-1":"0.02","ms5959-2":"0.03","ms5959-3":"0.03","ms5959-4":"0.03","ms5959-5":"0.19","ms5959-6":"0.02","ms5959-7":"0.01","ms5959-8":"0.02","ms5959-9":"0.03","ms5959-10":"0.03","ms5959-11":"0.09","ms5959-12":"0.02","ms5959-13":"0.02","ms5959-14":"0.03","ms5959-15":"0.01","ms5959-16":"0.03","ms5959-17":"0.02","ms5959-18":"0.03","ms5959-19":"0.01","ms5959-20":"0.02","ms5959-21":"0.02","ms5959-22":"0.03","ms5959-23":"0.04","ms5959-24":"0.04","ms5959-25":"0.02","ms5959-26":"0.03","ms5959-27":"0.02","ms5959-28":"0.02","ms5959-29":"0.03","ms5959-30":"0.03"},{"Metabolite":"DHA","ms5959-1":"0.63","ms5959-2":"0.63","ms5959-3":"0.73","ms5959-4":"0.84","ms5959-5":"0.71","ms5959-6":"0.56","ms5959-7":"0.53","ms5959-8":"0.70","ms5959-9":"0.75","ms5959-10":"0.53","ms5959-11":"0.76","ms5959-12":"0.51","ms5959-13":"0.50","ms5959-14":"0.61","ms5959-15":"0.64","ms5959-16":"0.44","ms5959-17":"0.59","ms5959-18":"0.61","ms5959-19":"0.71","ms5959-20":"0.67","ms5959-21":"0.79","ms5959-22":"0.96","ms5959-23":"0.80","ms5959-24":"0.94","ms5959-25":"0.32","ms5959-26":"0.43","ms5959-27":"0.38","ms5959-28":"0.38","ms5959-29":"0.47","ms5959-30":"0.53"},{"Metabolite":"myristic","ms5959-1":"0.25","ms5959-2":"0.18","ms5959-3":"0.21","ms5959-4":"0.31","ms5959-5":"0.51","ms5959-6":"0.17","ms5959-7":"0.20","ms5959-8":"0.31","ms5959-9":"0.27","ms5959-10":"0.29","ms5959-11":"0.33","ms5959-12":"0.11","ms5959-13":"0.12","ms5959-14":"0.13","ms5959-15":"0.12","ms5959-16":"0.12","ms5959-17":"0.15","ms5959-18":"0.16","ms5959-19":"0.14","ms5959-20":"0.20","ms5959-21":"0.25","ms5959-22":"0.18","ms5959-23":"0.15","ms5959-24":"0.29","ms5959-25":"0.14","ms5959-26":"0.33","ms5959-27":"0.07","ms5959-28":"0.08","ms5959-29":"0.10","ms5959-30":"0.16"},{"Metabolite":"palmitoleic","ms5959-1":"0.24","ms5959-2":"0.23","ms5959-3":"0.27","ms5959-4":"0.33","ms5959-5":"0.29","ms5959-6":"0.24","ms5959-7":"0.19","ms5959-8":"0.24","ms5959-9":"0.26","ms5959-10":"0.16","ms5959-11":"0.22","ms5959-12":"0.15","ms5959-13":"0.14","ms5959-14":"0.17","ms5959-15":"0.14","ms5959-16":"0.11","ms5959-17":"0.15","ms5959-18":"0.12","ms5959-19":"0.19","ms5959-20":"0.22","ms5959-21":"0.27","ms5959-22":"0.31","ms5959-23":"0.25","ms5959-24":"0.34","ms5959-25":"0.07","ms5959-26":"0.12","ms5959-27":"0.09","ms5959-28":"0.07","ms5959-29":"0.08","ms5959-30":"0.19"},{"Metabolite":"arachidonic","ms5959-1":"5.16","ms5959-2":"4.05","ms5959-3":"4.03","ms5959-4":"6.46","ms5959-5":"4.39","ms5959-6":"5.68","ms5959-7":"3.54","ms5959-8":"3.91","ms5959-9":"4.57","ms5959-10":"3.46","ms5959-11":"3.65","ms5959-12":"3.02","ms5959-13":"2.56","ms5959-14":"2.83","ms5959-15":"2.58","ms5959-16":"3.14","ms5959-17":"3.85","ms5959-18":"3.15","ms5959-19":"3.34","ms5959-20":"3.82","ms5959-21":"4.16","ms5959-22":"4.87","ms5959-23":"4.08","ms5959-24":"5.96","ms5959-25":"2.06","ms5959-26":"2.13","ms5959-27":"2.15","ms5959-28":"1.54","ms5959-29":"1.53","ms5959-30":"2.56"},{"Metabolite":"palmitelaidic","ms5959-1":"0.00","ms5959-2":"0.00","ms5959-3":"0.00","ms5959-4":"0.00","ms5959-5":"0.00","ms5959-6":"0.00","ms5959-7":"0.00","ms5959-8":"0.00","ms5959-9":"0.00","ms5959-10":"0.00","ms5959-11":"0.00","ms5959-12":"0.00","ms5959-13":"0.00","ms5959-14":"0.00","ms5959-15":"0.00","ms5959-16":"0.00","ms5959-17":"0.00","ms5959-18":"0.00","ms5959-19":"0.00","ms5959-20":"0.00","ms5959-21":"0.00","ms5959-22":"0.00","ms5959-23":"0.00","ms5959-24":"0.00","ms5959-25":"0.00","ms5959-26":"0.00","ms5959-27":"0.00","ms5959-28":"0.00","ms5959-29":"0.00","ms5959-30":"0.00"},{"Metabolite":"linoleic","ms5959-1":"0.75","ms5959-2":"0.76","ms5959-3":"0.80","ms5959-4":"1.06","ms5959-5":"0.79","ms5959-6":"0.85","ms5959-7":"0.68","ms5959-8":"0.86","ms5959-9":"0.87","ms5959-10":"0.63","ms5959-11":"1.31","ms5959-12":"0.65","ms5959-13":"0.56","ms5959-14":"0.60","ms5959-15":"0.50","ms5959-16":"0.60","ms5959-17":"0.62","ms5959-18":"0.58","ms5959-19":"0.68","ms5959-20":"0.79","ms5959-21":"0.82","ms5959-22":"0.94","ms5959-23":"0.78","ms5959-24":"1.06","ms5959-25":"0.45","ms5959-26":"0.66","ms5959-27":"0.52","ms5959-28":"0.49","ms5959-29":"0.54","ms5959-30":"0.57"},{"Metabolite":"palmitic","ms5959-1":"4.75","ms5959-2":"5.18","ms5959-3":"4.42","ms5959-4":"6.68","ms5959-5":"6.03","ms5959-6":"5.22","ms5959-7":"4.35","ms5959-8":"5.81","ms5959-9":"4.65","ms5959-10":"7.06","ms5959-11":"7.22","ms5959-12":"5.19","ms5959-13":"5.48","ms5959-14":"6.77","ms5959-15":"5.23","ms5959-16":"3.87","ms5959-17":"4.35","ms5959-18":"3.58","ms5959-19":"4.11","ms5959-20":"4.59","ms5959-21":"5.72","ms5959-22":"6.12","ms5959-23":"4.53","ms5959-24":"7.33","ms5959-25":"3.21","ms5959-26":"4.14","ms5959-27":"3.95","ms5959-28":"3.10","ms5959-29":"3.31","ms5959-30":"4.03"},{"Metabolite":"oleic","ms5959-1":"7.14","ms5959-2":"7.16","ms5959-3":"6.67","ms5959-4":"9.91","ms5959-5":"6.27","ms5959-6":"8.21","ms5959-7":"6.10","ms5959-8":"8.13","ms5959-9":"6.57","ms5959-10":"7.81","ms5959-11":"8.26","ms5959-12":"6.98","ms5959-13":"6.71","ms5959-14":"8.16","ms5959-15":"6.17","ms5959-16":"5.34","ms5959-17":"6.43","ms5959-18":"5.21","ms5959-19":"5.66","ms5959-20":"6.05","ms5959-21":"7.14","ms5959-22":"8.88","ms5959-23":"7.07","ms5959-24":"8.81","ms5959-25":"4.04","ms5959-26":"4.53","ms5959-27":"4.65","ms5959-28":"3.48","ms5959-29":"3.60","ms5959-30":"5.47"},{"Metabolite":"elaidic","ms5959-1":"0.00","ms5959-2":"0.00","ms5959-3":"0.00","ms5959-4":"0.00","ms5959-5":"0.03","ms5959-6":"0.00","ms5959-7":"0.00","ms5959-8":"0.00","ms5959-9":"0.00","ms5959-10":"0.00","ms5959-11":"0.00","ms5959-12":"0.00","ms5959-13":"0.00","ms5959-14":"0.00","ms5959-15":"0.00","ms5959-16":"0.00","ms5959-17":"0.00","ms5959-18":"0.00","ms5959-19":"0.00","ms5959-20":"0.00","ms5959-21":"0.00","ms5959-22":"0.00","ms5959-23":"0.00","ms5959-24":"0.03","ms5959-25":"0.01","ms5959-26":"0.07","ms5959-27":"0.00","ms5959-28":"0.00","ms5959-29":"0.00","ms5959-30":"0.04"},{"Metabolite":"stearic","ms5959-1":"4.88","ms5959-2":"5.53","ms5959-3":"3.82","ms5959-4":"6.79","ms5959-5":"6.48","ms5959-6":"6.16","ms5959-7":"5.22","ms5959-8":"5.70","ms5959-9":"4.22","ms5959-10":"6.89","ms5959-11":"5.94","ms5959-12":"4.46","ms5959-13":"4.91","ms5959-14":"5.78","ms5959-15":"4.14","ms5959-16":"3.63","ms5959-17":"3.61","ms5959-18":"2.25","ms5959-19":"3.99","ms5959-20":"4.39","ms5959-21":"6.04","ms5959-22":"6.17","ms5959-23":"4.28","ms5959-24":"7.66","ms5959-25":"3.12","ms5959-26":"3.75","ms5959-27":"4.02","ms5959-28":"2.83","ms5959-29":"2.77","ms5959-30":"3.41"}],

"Metabolites":[{"metabolite_name":"EPA"},{"metabolite_name":"linolenic"},{"metabolite_name":"DHA"},{"metabolite_name":"myristic"},{"metabolite_name":"palmitoleic"},{"metabolite_name":"arachidonic"},{"metabolite_name":"palmitelaidic"},{"metabolite_name":"linoleic"},{"metabolite_name":"palmitic"},{"metabolite_name":"oleic"},{"metabolite_name":"elaidic"},{"metabolite_name":"stearic"}]
}

}