{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST000358","ANALYSIS_ID":"AN001447","VERSION":"1","CREATED_ON":"November 3, 2017, 11:25 am"},

"PROJECT":{"PROJECT_TITLE":"Metabolic Profiling of Anxiety Prone HSV-Latently Infected Obese Mice","PROJECT_TYPE":"Broad Spectrum LCMS","PROJECT_SUMMARY":"The biological factors that lead children from low socioeconomic backgrounds to be at greater risk for the development of anxiety and learning problems are not well understood. While it is clear that there are genetic components to the risk of developing mental health disorders, a role for environmental factors in inducing these problems has been suggested. Many of these factors that affect the brain likely involve exposures that occur early in life. Two such factors are the higher rates of herpes simplex virus (HSV)-1 seropositivity and prevalence of obesity among these children. HSV-1 infection has been associated with impaired cognition during childhood and mental health problems in adulthood. Additionally, obese adults are shown to have higher HSV-1 titers. Similarly, other studies have correlated a diet high in saturated fat and obesity with increased risk of mood disorders and anxiety. A mouse model of obese HSV-1 latent infection was developed. Broad spectrum metabolomics analysis was performed to better understand the metabolomic profile of hippocampus and to compare this metabolomics profile with that of the hypothalamus, microglia, and peripheral blood mononuclear cells. Brain tissue samples for metabolomics experiments were generated in the following manner: 3-week old mice were placed on a 10% low fat (LF) diet and acclimated for one week prior to intranasal HSV-1 infection or mock infection with PBS. Fourteen days post-infection mice were randomized to either a 45% high fat diet (HF) or remained on the LF diet. Eight weeks post-diet transition, mice were euthanized and brain tissue samples were collected and processed for metabolomics.","INSTITUTE":"University of North Carolina , Chapel Hill","DEPARTMENT":"Gillings School of Public Health","LABORATORY":"Department of Nutrition","LAST_NAME":"Sheridan","FIRST_NAME":"Patricia","ADDRESS":"2002 Hooker Research Center, CB#7461, Chapel Hill NC, 27599","EMAIL":"patricia_sheridan@med.unc.edu","PHONE":"919-843-6434","FUNDING_SOURCE":"NIH, Common Fund, Pilot and Feasibility"},

"STUDY":{"STUDY_TITLE":"Broad Spectrum MS analysis of mouse hippocampus from Anxiety Prone HSV-Latently Infected Obese Mice","STUDY_TYPE":"Broad Spectrum LCMS","STUDY_SUMMARY":"Brain tissue samples for metabolomics experiments were generated in the following manner: 3-week old mice were placed on a 10% low fat (LF) diet and acclimated for one week prior to intranasal HSV-1 infection or mock infection with PBS. Fourteen days post-infection mice were randomized to either a 45% high fat diet (HF) or remained on the LF diet. Eight weeks post-diet transition, mice were euthanized and brain tissue samples were collected and processed for metabolomics.","INSTITUTE":"RTI","DEPARTMENT":"Discovery Sciences","LABORATORY":"System and Translational Sciences","LAST_NAME":"Sumner","FIRST_NAME":"Susan","ADDRESS":"3040 Cornwallis Rd, RTP,NC","EMAIL":"ssumner@rti.org","PHONE":"919-541-7479","NUM_GROUPS":"4","TOTAL_SUBJECTS":"31","NUM_MALES":"31"},

"SUBJECT":{"SUBJECT_TYPE":"mouse","SUBJECT_SPECIES":"Mus musculus","TAXONOMY_ID":"10090","GENOTYPE_STRAIN":"C57Bl/6","AGE_OR_AGE_RANGE":"5.5 mo","WEIGHT_OR_WEIGHT_RANGE":"29.1 g-49.2 g","GENDER":"male","ANIMAL_ANIMAL_SUPPLIER":"JAX","ANIMAL_HOUSING":"4/cage","ANIMAL_FEED":"Purina chow"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"409",
"Sample ID":"Hipp409",
"Factors":{"HSV-1":"yes","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"37.1"}
},
{
"Subject ID":"394",
"Sample ID":"Hipp394",
"Factors":{"HSV-1":"yes","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"48.2"}
},
{
"Subject ID":"391",
"Sample ID":"Hipp391",
"Factors":{"HSV-1":"no","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"31.2"}
},
{
"Subject ID":"414",
"Sample ID":"Hipp414",
"Factors":{"HSV-1":"yes","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"36.6"}
},
{
"Subject ID":"398",
"Sample ID":"Hipp398",
"Factors":{"HSV-1":"yes","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"35.2"}
},
{
"Subject ID":"392",
"Sample ID":"Hipp392",
"Factors":{"HSV-1":"no","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"34"}
},
{
"Subject ID":"386",
"Sample ID":"Hipp386",
"Factors":{"HSV-1":"no","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"34.8"}
},
{
"Subject ID":"396",
"Sample ID":"Hipp396",
"Factors":{"HSV-1":"yes","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"48.4"}
},
{
"Subject ID":"388",
"Sample ID":"Hipp388",
"Factors":{"HSV-1":"no","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"44.1"}
},
{
"Subject ID":"412",
"Sample ID":"Hipp412",
"Factors":{"HSV-1":"yes","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"34.7"}
},
{
"Subject ID":"413",
"Sample ID":"Hipp413",
"Factors":{"HSV-1":"yes","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"36"}
},
{
"Subject ID":"389",
"Sample ID":"Hipp389",
"Factors":{"HSV-1":"no","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"34"}
},
{
"Subject ID":"411",
"Sample ID":"Hipp411",
"Factors":{"HSV-1":"yes","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"40.2"}
},
{
"Subject ID":"408",
"Sample ID":"Hipp408",
"Factors":{"HSV-1":"no","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"29.1"}
},
{
"Subject ID":"403",
"Sample ID":"Hipp403",
"Factors":{"HSV-1":"no","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"48.6"}
},
{
"Subject ID":"415",
"Sample ID":"Hipp415",
"Factors":{"HSV-1":"yes","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"34.8"}
},
{
"Subject ID":"401",
"Sample ID":"Hipp401",
"Factors":{"HSV-1":"no","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"49.2"}
},
{
"Subject ID":"405",
"Sample ID":"Hipp405",
"Factors":{"HSV-1":"no","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"30.8"}
},
{
"Subject ID":"395",
"Sample ID":"Hipp395",
"Factors":{"HSV-1":"yes","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"40.9"}
},
{
"Subject ID":"402",
"Sample ID":"Hipp402",
"Factors":{"HSV-1":"no","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"38.8"}
},
{
"Subject ID":"390",
"Sample ID":"Hipp390",
"Factors":{"HSV-1":"no","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"34.5"}
},
{
"Subject ID":"410",
"Sample ID":"Hipp410",
"Factors":{"HSV-1":"yes","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"37.8"}
},
{
"Subject ID":"385",
"Sample ID":"Hipp385",
"Factors":{"HSV-1":"no","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"41.6"}
},
{
"Subject ID":"397",
"Sample ID":"Hipp397",
"Factors":{"HSV-1":"yes","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"33.1"}
},
{
"Subject ID":"399",
"Sample ID":"Hipp399",
"Factors":{"HSV-1":"yes","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"33"}
},
{
"Subject ID":"393",
"Sample ID":"Hipp393",
"Factors":{"HSV-1":"yes","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"44.1"}
},
{
"Subject ID":"407",
"Sample ID":"Hipp407",
"Factors":{"HSV-1":"no","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"34.7"}
},
{
"Subject ID":"406",
"Sample ID":"Hipp406",
"Factors":{"HSV-1":"no","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"31.4"}
},
{
"Subject ID":"387",
"Sample ID":"Hipp387",
"Factors":{"HSV-1":"no","Diet at Week 2":"HF"},
"Additional sample data":{"Weight (g)":"31.9"}
},
{
"Subject ID":"400",
"Sample ID":"Hipp400",
"Factors":{"HSV-1":"yes","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"33.2"}
},
{
"Subject ID":"416",
"Sample ID":"Hipp416",
"Factors":{"HSV-1":"yes","Diet at Week 2":"LF"},
"Additional sample data":{"Weight (g)":"35.2"}
},
{
"Subject ID":"-",
"Sample ID":"Total_Pool_01",
"Factors":{"HSV-1":"-","Diet at Week 2":"-"},
"Additional sample data":{"Weight (g)":"-"}
},
{
"Subject ID":"-",
"Sample ID":"Total_Pool_02",
"Factors":{"HSV-1":"-","Diet at Week 2":"-"},
"Additional sample data":{"Weight (g)":"-"}
},
{
"Subject ID":"-",
"Sample ID":"Total_Pool_03",
"Factors":{"HSV-1":"-","Diet at Week 2":"-"},
"Additional sample data":{"Weight (g)":"-"}
},
{
"Subject ID":"-",
"Sample ID":"Total_Pool_04",
"Factors":{"HSV-1":"-","Diet at Week 2":"-"},
"Additional sample data":{"Weight (g)":"-"}
},
{
"Subject ID":"-",
"Sample ID":"Total_Pool_05",
"Factors":{"HSV-1":"-","Diet at Week 2":"-"},
"Additional sample data":{"Weight (g)":"-"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"Brain tissue samples were homogenized in 50:50 Acetonitrile:Water (10 µL to every 1 mg of tissue) with washed ceramic beads on a MagNA Lyser, using two 30 sec pulses at 2,000 rpm with a 1 min chilling step in between pulses. Samples were vortexed on a multi-tube vortexer for 2 min at 5,000 rpm and centrifuged at room temperature for 4 min at 16,000 rcf. A 50 µL aliquot of each experimental brain tissue homogenized supernatant was transferred to a labeled 2.0 mL Lo-Bind eppendorf tube. Analytical pooled samples were created by combining 15 µL aliquot from each experimental sample in a 2.0 mL Lo-Bind eppendorf tube. The QC pooled sample was vortexed for 30 sec and 50 µL aliquots were transferred to 5 labeled 2.0 mL Lo-Bind eppendorf tubes. Next, 350 µL of acetonitrile was added to study sample and QC pooled sample tubes. The samples were vortexed on a multi-tube vortexer for 2 min at 5,000 rpm, and centrifuged at room temperature for 4 min at 16,000 rcf. The supernatants were transferred to new labeled tubes and dried on a lyophilizer overnight. Dried samples were reconstituted in 50 µL of 95:5 Water:Methanol, mixed on an analog vortex mixer for 1 min with speed set at 10. Then, the samples were centrifuged at room temperature for 4 min at 16,000 rcf, and the supernatants were transferred to autosampler vials for data acquisition. UPLC-MS Methods: UPLC-MS spectra were collected for all samples. UPLC was performed on a Waters Acquity UPLC with an Acquity HSS T3 column (2.1x 100mm x 1.8 µm) at 50 °C using the reversed-phase separation. Water with 0.1% formic acid (mobile phase A) and methanol with 0.1% formic acid (mobile phase B) were used as mobile phases and the metabolites were chromatographically separated using a gradient separation: (see the 3. Sheridan-Mice-Hypothalamus_RP-Metadata and Analytical Metadata.xlsx file for the flow gradient). Mass spectroscopy analysis was performed using a Synapt G2 Si ESI-Q-TOF using a 10 µL injection volume. UPLC-MS data were collected over 70-1000 m/z in both positive and negative modes.","SAMPLE_TYPE":"brain homogenate"},

"TREATMENT":{"TREATMENT_SUMMARY":"3-week old mice were placed on a 10% low fat (LF) diet and acclimated for one week prior to intranasal HSV-1 infection or mock infection with PBS. Fourteen days post-infection mice were randomized to either a 45% high fat diet (HF) or remained on the LF diet. Eight weeks post-diet transition, mice were euthanized and brain tissue samples were collected and processed for metabolomics."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"Brain tissue samples were homogenized in 50:50 Acetonitrile:Water (10 µL to every 1 mg of tissue) with washed ceramic beads on a MagNA Lyser, using two 30 sec pulses at 2,000 rpm with a 1 min chilling step in between pulses. Samples were vortexed on a multi-tube vortexer for 2 min at 5,000 rpm and centrifuged at room temperature for 4 min at 16,000 rcf. A 50 µL aliquot of each experimental brain tissue homogenized supernatant was transferred to a labeled 2.0 mL Lo-Bind eppendorf tube. Analytical pooled samples were created by combining 15 µL aliquot from each experimental sample in a 2.0 mL Lo-Bind eppendorf tube. The QC pooled sample was vortexed for 30 sec and 50 µL aliquots were transferred to 5 labeled 2.0 mL Lo-Bind eppendorf tubes. Next, 350 µL of acetonitrile was added to study sample and QC pooled sample tubes. The samples were vortexed on a multi-tube vortexer for 2 min at 5,000 rpm, and centrifuged at room temperature for 4 min at 16,000 rcf. The supernatants were transferred to new labeled tubes and dried on a lyophilizer overnight. Dried samples were reconstituted in 50 µL of 95:5 Water:Methanol, mixed on an analog vortex mixer for 1 min with speed set at 10. Then, the samples were centrifuged at room temperature for 4 min at 16,000 rcf, and the supernatants were transferred to autosampler vials for data acquisition."},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_SUMMARY":"UPLC was performed on a Waters Acquity UPLC with an Acquity HSS T3 column (2.1x 100mm x 1.8 µm) at 50 °C using the reversed-phase separation. Water with 0.1% formic acid (mobile phase A) and methanol with 0.1% formic acid (mobile phase B) were used as mobile phases and the metabolites were chromatographically separated using a gradient separation:","CHROMATOGRAPHY_TYPE":"Reversed phase","INSTRUMENT_NAME":"Waters Acquity","COLUMN_NAME":"Acquity HSS T3","METHODS_FILENAME":"RTI-RCMRC-RP"},

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

"MS":{"MS_COMMENTS":"-","INSTRUMENT_NAME":"Waters Synapt G2 Si QTOF","INSTRUMENT_TYPE":"QTOF","MS_TYPE":"ESI","ION_MODE":"NEGATIVE","MS_RESULTS_FILE":"ST000358_AN001447_Results.txt UNITS:intensity"}

}