Summary of Study ST002805
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 PR001752. The data can be accessed directly via it's Project DOI: 10.21228/M87Q7Z This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
| Study ID | ST002805 |
| Study Title | IL-1β-mediated adaptive re-programming of endogenous human cardiac fibroblasts to cells with immune features during fibrotic remodeling |
| Study Summary | The source and roles of fibroblasts and CD4 helper T-cells during maladaptive remodeling and myocardial fibrosis in pulmonary arterial hypertension (PAH) have been long debated. We demonstrate, using single-cell mass cytometry, a sub-population of endogenous human cardiac fibroblasts expressing increased levels of CD4, a helper T-cell marker, in addition to myofibroblast markers distributed in human fibrotic RV tissue, interstitial/perivascular lesions of SUGEN/Hypoxia (SuHx) rats and fibroblasts labelled with pdgfrα CreERt2/+ in R26R-tdTomato mice. Recombinant IL-1β increases IL-1R, CCR2 receptor expression, modifies the secretome, and differentiates cardiac fibroblasts to form CD68 positive cell clusters. IL-1β also activates stemness markers such as NANOG and SOX2 and genes involved in de-differentiation, lymphoid cell function and metabolic reprogramming. IL-1β induction of lineage traced primary mouse cardiac fibroblasts causes these cells to lose their fibroblast identity and acquire an immune phenotype. Our results identify IL-1β induced immune-competency in human cardiac fibroblasts and suggest that fibroblast secretome modulation may constitute a therapeutic approach to PAH and other diseases typified by inflammation and fibrotic remodeling. |
| Institute | Brown University |
| Department | Molecular Pharmacology, Physiology and Biotechnology |
| Laboratory | Siamwala Lab |
| Last Name | Siamwala |
| First Name | Jamila |
| Address | 830 Chalkstone Avenue, Building 35 |
| jamila_siamwala@brown.edu | |
| Phone | 6192136223 |
| Submit Date | 2023-07-18 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | wiff |
| Analysis Type Detail | Other |
| Release Date | 2023-08-07 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR001752 |
| Project DOI: | doi: 10.21228/M87Q7Z |
| Project Title: | IL-1β-mediated adaptive re-programming of endogenous human cardiac fibroblasts to cells with immune features during fibrotic remodeling |
| Project Summary: | The source and roles of fibroblasts and CD4 helper T-cells during maladaptive remodeling and myocardial fibrosis in pulmonary arterial hypertension (PAH) have been long debated. We demonstrate, using single-cell mass cytometry, a sub-population of endogenous human cardiac fibroblasts expressing increased levels of CD4, a helper T-cell marker, in addition to myofibroblast markers distributed in human fibrotic RV tissue, interstitial/perivascular lesions of SUGEN/Hypoxia (SuHx) rats and fibroblasts labelled with pdgfrα CreERt2/+ in R26R-tdTomato mice. Recombinant IL-1β increases IL-1R, CCR2 receptor expression, modifies the secretome, and differentiates cardiac fibroblasts to form CD68 positive cell clusters. IL-1β also activates stemness markers such as NANOG and SOX2 and genes involved in de-differentiation, lymphoid cell function and metabolic reprogramming. IL-1β induction of lineage traced primary mouse cardiac fibroblasts causes these cells to lose their fibroblast identity and acquire an immune phenotype. Our results identify IL-1β induced immune-competency in human cardiac fibroblasts and suggest that fibroblast secretome modulation may constitute a therapeutic approach to PAH and other diseases typified by inflammation and fibrotic remodeling. |
| Institute: | Brown University |
| Department: | MPPB |
| Laboratory: | Siamwala Lab |
| Last Name: | Siamwala |
| First Name: | Jamila |
| Address: | 830 Chalkstone Avenue, Building 35 |
| Email: | jamila_siamwala@brown.edu |
| Phone: | 6192136223 |
Subject:
| Subject ID: | SU002912 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Age Or Age Range: | 63 - 73 |
| Gender: | Male and female |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Sex | Treatment |
|---|---|---|---|
| SA300847 | 534282_c | F | Control |
| SA300848 | 534282_t | F | IL-1B |
| SA300849 | 1281202_c | M | Control |
| SA300850 | 62122_c | M | Control |
| SA300851 | 62122_t | M | IL-1B |
| SA300852 | 1281202_t | M | IL-1B |
| Showing results 1 to 6 of 6 |
Collection:
| Collection ID: | CO002905 |
| Collection Summary: | Metabolomics of conditioned media was performed at the Beth Israel Deaconess Medical Center Mass Spectrometry Core according to published protocols (M. Yuan, S. B. Breitkopf, X. Yang, J. M. Asara, A positive/negative ion-switching, targeted mass spectrometry-based metabolomics platform for bodily fluids, cells, and fresh and fixed tissue. Nat Protoc 7, 872-881 (2012)). hVCF (2 x 106) were seeded and grown to confluence and treated with IL-1β (10 ng/ml) for 24h. Conditioned media (1ml) from the vehicle or IL-1β treated hVCF cells were used for the metabolomics analysis. |
| Sample Type: | Cultured fibroblasts |
Treatment:
| Treatment ID: | TR002921 |
| Treatment Summary: | hVCF (2 x 106) were seeded and grown to confluence and treated with IL-1β (10 ng/ml) for 24h. Conditioned media (1ml) from the vehicle or IL-1β treated hVCF cells were used for the metabolomics analysis. |
Sample Preparation:
| Sampleprep ID: | SP002918 |
| Sampleprep Summary: | 500 μL of chilled -80°C 80% methanol was added to 15ml tubes containing 1ml of conditioned media and evaporated using Speedvac to pellet the metabolites. SRM with polarity switching with a QTRAP 5500 mass spectrometer (AB/SCIEX) was used to assay 300 polar molecules, based on the previously published protocol. |
Combined analysis:
| Analysis ID | AN004561 |
|---|---|
| Analysis type | MS |
| Chromatography type | HILIC |
| Chromatography system | Shimadzu Prominence UFLC HPLC |
| Column | Waters XBridge Amide (100 x 4.6mm,3.5um) |
| MS Type | ESI |
| MS instrument type | Triple quadrupole |
| MS instrument name | ABI Sciex 6500 QTrap |
| Ion Mode | UNSPECIFIED |
| Units | Q3 Peak Area (area under curve) |
Chromatography:
| Chromatography ID: | CH003427 |
| Methods Filename: | nprot_2012_024_v2.pdf |
| Instrument Name: | Shimadzu Prominence UFLC HPLC |
| Column Name: | Waters XBridge Amide (100 x 4.6mm,3.5um) |
| Column Temperature: | 475 |
| Flow Gradient: | Gradients were run with a Prominence UFLC HPLC (Shimadzu) starting from 85% buffer B (HPLC grade acetonitrile) to 42% B from 0-5 minutes; 42% B to 0% B from 5-16 minutes; 0% B was held from 16-24 minutes; 0% B to 85% B from 24-25 minutes; 85% B was held for 7 minutes to re-equilibrate the column. |
| Flow Rate: | 400uL/min |
| Solvent A: | 20 mM ammonium hydroxide/20 mM ammonium acetate (pH=9.0) in 95:5 water:acetonitrile |
| Solvent B: | HPLC grade acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS004307 |
| Analysis ID: | AN004561 |
| Instrument Name: | ABI Sciex 6500 QTrap |
| Instrument Type: | Triple quadrupole |
| MS Type: | ESI |
| MS Comments: | Ion Mode Positive/Negative Switching: Metabolomics of conditioned media was performed at the Beth Israel Deaconess Medical Center Mass Spectrometry Core according to published protocols (M. Yuan, S. B. Breitkopf, X. Yang, J. M. Asara, A positive/negative ion-switching, targeted mass spectrometry-based metabolomics platform for bodily fluids, cells, and fresh and fixed tissue. Nat Protoc 7, 872-881 (2012)). hVCF (2 x 106) were seeded and grown to confluence and treated with IL-1β (10 ng/ml) for 24h. Conditioned media (1ml) from the vehicle or IL-1β treated hVCF cells were used for the metabolomics analysis. 500 μL of chilled -80°C 80% methanol was added to 15ml tubes containing 1ml of conditioned media and evaporated using Speedvac to pellet the metabolites. Samples were re-suspended using 20 mL HPLC grade water for mass spectrometry. 5-7 μL were injected and analyzed using a hybrid 6500 QTRAP triple quadrupole mass spectrometer (AB/SCIEX) coupled to a Prominence UFLC HPLC system (Shimadzu) via selected reaction monitoring (SRM) of a total of 283 endogenous water soluble metabolites for steady-state analyses of samples. Some metabolites were targeted in both positive and negative ion mode for a total of 304 SRM transitions using positive/negative ion polarity switching. ESI voltage was +4950V in positive ion mode and –4500V in negative ion mode. The dwell time was 3 ms per SRM transition and the total cycle time was 1.39 seconds. Approximately 10-14 data points were acquired per detected metabolite. Samples were delivered to the mass spectrometer via hydrophilic interaction chromatography (HILIC) using a 4.6 mm i.d x 10 cm Amide XBridge column (Waters) at 400 μL/min. Gradients were run with a Prominence UFLC HPLC (Shimadzu) starting from 85% buffer B (HPLC grade acetonitrile) to 42% B from 0-5 minutes; 42% B to 0% B from 5-16 minutes; 0% B was held from 16-24 minutes; 0% B to 85% B from 24-25 minutes; 85% B was held for 7 minutes to re-equilibrate the column. Buffer A was comprised of 20 mM ammonium hydroxide/20 mM ammonium acetate (pH=9.0) in 95:5 water:acetonitrile. Peak areas from the total ion current for each metabolite SRM transition were integrated using MultiQuant v3.0 software (AB/SCIEX) |
| Ion Mode: | UNSPECIFIED |
| Analysis Protocol File: | nprot_2012_024_v2.pdf |
