Summary of Study ST002056
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001301. The data can be accessed directly via it's Project DOI: 10.21228/M8JH7P This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST002056 |
| Study Title | Integrated Multilayer Omics Reveals the Genomic, Proteomic and Metabolic Influences of the Histidyl Dipeptides on Heart |
| Study Type | Triomics |
| Study Summary | Histidyl dipeptides, such as carnosine, present in a micro-millimolar ranges in the heart, are synthesized via the enzyme carnosine synthase (Carns). These dipeptides facilitate glycolysis by proton buffering, form conjugates with reactive aldehydes, such as acrolein, and attenuate ischemia and reperfusion injury. While these dipeptides exhibit multifunctional properties, a composite understanding of their roles in myocardium is lacking. To identify the landscape of histidyl dipeptide mediated responses in the heart, we used a triomics approach of genome wide RNA sequencing, global proteomics and unbiased metabolomics in the cardio specific Carns transgenic (Tg) mice and integrated the three data sets. Our result show higher myocardial levels of histidyl dipeptides lead to extensive changes in several microRNAs, which could target the expression of contractile proteins, beta-fatty acid oxidation and citric acid cycle (TCA) enzymes. Global proteomics shows, expression of contractile proteins, enzymes of beta-fatty acid oxidation and TCA cycle, were enriched in the CarnsTg heart. Under aerobic conditions, the CarnsTg hearts had lower levels of short and long-chain fatty acids and TCA cycle intermediate-succinic acid, whereas, under ischemic conditions the accumulation of fatty acids and TCA cycle intermediates were significantly attenuated in the CarnsTg heart. Integration of multiple data sets suggests that beta-fatty acid oxidation and TCA cycle pathways exhibited correlative changes in the CarnsTg hearts at all three levels. Our triomics approach shows histidyl dipeptides are critical regulators of myocardial structure, function and energetics. |
| Institute | University of Louisville |
| Last Name | Baba |
| First Name | Shahid |
| Address | 580 S. Preston St |
| spbaba01@louisville.edu | |
| Phone | 5028524274 |
| Submit Date | 2021-12-15 |
| Num Groups | 4 |
| Analysis Type Detail | GC-MS |
| Release Date | 2022-01-31 |
| Release Version | 1 |
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Project:
| Project ID: | PR001301 |
| Project DOI: | doi: 10.21228/M8JH7P |
| Project Title: | Integrated multilayer omics reveals the genomic, proteomic and metabolic influences of the histidyl dipeptides on heart |
| Project Type: | Triomics of WT and Carnosin synthase transgenic heart MSTFA dervitization |
| Project Summary: | Histidyl dipeptides, such as carnosine, present in a micro-millimolar ranges in the heart, are synthesized via the enzyme carnosine synthase (Carns). These dipeptides facilitate glycolysis by proton buffering, form conjugates with reactive aldehydes, such as acrolein, and attenuate ischemia and reperfusion injury. While these dipeptides exhibit multifunctional properties, a composite understanding of their roles in myocardium is lacking. To identify the landscape of histidyl dipeptide mediated responses in the heart, we used a triomics approach of genome wide RNA sequencing, global proteomics and unbiased metabolomics in the cardio specific Carns transgenic (Tg) mice and integrated the three data sets. Our result show higher myocardial levels of histidyl dipeptides lead to extensive changes in several microRNAs, which could target the expression of contractile proteins, beta-fatty acid oxidation and citric acid cycle (TCA) enzymes. Global proteomics shows, expression of contractile proteins, enzymes of beta-fatty acid oxidation and TCA cycle, were enriched in the CarnsTg heart. Under aerobic conditions, the CarnsTg hearts had lower levels of short and long-chain fatty acids and TCA cycle intermediate-succinic acid, whereas, under ischemic conditions the accumulation of fatty acids and TCA cycle intermediates were significantly attenuated in the CarnsTg heart. Integration of multiple data sets suggests that beta-fatty acid oxidation and TCA cycle pathways exhibited correlative changes in the CarnsTg hearts at all three levels. Our triomics approach shows histidyl dipeptides are critical regulators of myocardial structure, function and energetics. |
| Institute: | University of Louisville |
| Department: | Medicine |
| Laboratory: | Diabetes and Obesity Center |
| Last Name: | Baba |
| First Name: | Shahid |
| Address: | 580 S. Preston St |
| Email: | spbaba01@louisville.edu |
| Phone: | 5028524274 |
| Funding Source: | NIH |
Subject:
| Subject ID: | SU002138 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Age Or Age Range: | 8-12 weeks |
| Gender: | Male |
Factors:
Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)
| mb_sample_id | local_sample_id | Genotype |
|---|---|---|
| SA193790 | CarnsTG1 | CarnsTG |
| SA193791 | CarnsTG4 | CarnsTG |
| SA193792 | CarnsTG3 | CarnsTG |
| SA193793 | CarnsTG2 | CarnsTG |
| SA193794 | WT4 | Wild-type |
| SA193795 | WT1 | Wild-type |
| SA193796 | WT2 | Wild-type |
| SA193797 | WT3 | Wild-type |
| Showing results 1 to 8 of 8 |
Collection:
| Collection ID: | CO002131 |
| Collection Summary: | To determine the effects of the Carns overexpression on the global cardio metabolomic profile under the basal conditions, hearts were collected from the WT and CarnsTg mice upon sacrifice and snap-frozen in liquid nitrogen. |
| Collection Protocol ID: | 18404 |
| Collection Protocol Filename: | Metabolomics |
| Collection Protocol Comments: | Effect of Carns overexpression |
| Sample Type: | Heart |
| Collection Location: | University of Louisville |
| Storage Conditions: | -80℃ |
| Collection Tube Temp: | -80 |
Treatment:
| Treatment ID: | TR002150 |
| Treatment Summary: | To determine the changes in the cardio metabolomic profile associated with the ischemic injury and whether Carns overexpression influences the cardiac metabolism, isolated hearts from the WT and CarnsTg mice hearts were perfused in the Langendorff mode for 35 min, perfused for 20 min followed by 5 min of ischemia and perfused for 20 min followed by 15 min of ischemia as described previousl |
Sample Preparation:
| Sampleprep ID: | SP002144 |
| Sampleprep Summary: | To extract polar metabolites, 800 µL of methanol was added to 200 µL of homogenized heart sample. Mixture was vortexed for 2 min and then placed on ice for 10 min, followed by another 2 min of vortex mixing, and centrifuged at 15,000 rpm for 20 min at 4 °C. Supernatant was transferred into a glass vial and dried in a SpeedVac evaporator to remove methanol, followed by lyophilization to remove water. The dried metabolite extract was dissolved with 30 µL of 20 mg/mL methoxyamine hydrochloride pyridine solution followed by vigorous vortex mixing for 1 min. Methoxylation was performed by sonicating the sample for 20 min and incubation at 60 °C for 1 h. Derivatization was performed by adding 20 µL of N-trimethylsilyl-N-methyl trifluoroacetamide (MSTFA) or N-Methyl-N-tert-butyldimethysilyltrifuoroacetamide (MTBSTFA) with 1% trimethylchlorosilane to the glass vial. Samples were incubated for 1 h at 60 °C and the mixture was transferred to a GC vial for analysis. Pooled samples were prepared by mixing 30 µL of derivatized metabolite extract from each sample to monitor the instrumental variations during the course of GC×GC-MS analysis. |
Combined analysis:
| Analysis ID | AN003348 |
|---|---|
| Analysis type | MS |
| Chromatography type | GC |
| Chromatography system | Agilent 6890N |
| Column | Agilent DB5-MS (60m × 0.25mm, 0.25um) |
| MS Type | EI |
| MS instrument type | GC-TOF |
| MS instrument name | Leco Pegasus III GC TOF |
| Ion Mode | UNSPECIFIED |
| Units | intensity |
Chromatography:
| Chromatography ID: | CH002479 |
| Chromatography Summary: | The extracted and derivatized samples were analyzed on a LECO Pegasus GC×GC-TOF MS instrument (LECO Corp., St. Joseph, MI, USA) coupled to an Agilent 6890 gas chromatography and a Gerstel MPS2 autosampler (GERSTEL Inc., Linthicum, MD, USA), featuring a LECO two-stage cryogenic modulator and secondary oven. The primary column was a 60 m × 0.25 mm 1dc × 0.25 µm 1df DB-5 ms GC capillary column (phenyl arylene polymer virtually equivalent to (5%-phenyl)-methylpolysiloxane). The secondary GC column 1 m × 0.25 mm 2dc × 0.25 µm 2df, DB-17 ms ((50% phenyl)-methylpolysiloxane) was placed inside the secondary GC oven following the thermal modulator. Both columns were obtained from Agilent Technologies (Agilent Technologies J&W, Santa Clara, CA, USA) and were connected through a press fit connector. The helium carrier gas (99.999% purity) flow rate was set to 2.0 mL/min at a corrected constant flow via pressure ramps. The inlet temperature was set at 280 °C. The primary column temperature was programmed with an initial temperature of 60 °C for 0.5 min, then ramped at 5°C/min to 270 °C, and maintained for 15 min. The secondary column temperature program was set to an initial temperature of 70 °C for 0.5 min and then ramped at the same temperature gradient employed in the first column to 280 °C, accordingly. The thermal modulator was set to 15 °C relative to the primary oven and a modulation time was 2s. The mass range was set as 29-800 m/z with an acquisition rate of 200 mass spectra per second. The ion source chamber was 230°C with the transfer line temperature of 280°C, and the detector voltage was 1440 V with electron energy of 70 eV. The acceleration voltage was turned on after a solvent delay of 544 s, the split ratio was set at 10:1. |
| Instrument Name: | Agilent 6890N |
| Column Name: | Agilent DB5-MS (60m × 0.25mm, 0.25um) |
| Chromatography Type: | GC |
MS:
| MS ID: | MS003117 |
| Analysis ID: | AN003348 |
| Instrument Name: | Leco Pegasus III GC TOF |
| Instrument Type: | GC-TOF |
| MS Type: | EI |
| MS Comments: | None |
| Ion Mode: | UNSPECIFIED |
