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Spatially-targeted optical microproteomics timecourse of rat myocardial infarction

ABSTRACT: Myocardial infarction (MI) results from occlusion of blood supply to the heart muscle causing death of cardiac muscle cells. Following myocardial infarction (MI), scar formation acts to mechanically stabilise the injured heart to prevent rupture and death. While fibroblasts and macrophages are implicated in post-MI scar formation and maturation, their exact contributions to this process are poorly characterised, especially in the long-term (i.e., beyond 1-week post-MI). Here, we employ state-of-the-art spatially-targetted optical micro-proteomics (STOMP) to isolate proteomes of tissue fractions enriched for fibroblasts (sma+) and macrophages (cd68+) over a 6-week post-MI timecourse and compare them to whole-scar proteome. We illustrate dynamic, specific changes in sma- and cd68-enriched fraction protein composition over 1-6 weeks post-MI with some of these changes reflected in whole-infarct preparations. These results link specific cell populations to specific protein changes in whole infarct.

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Rattus Norvegicus (rat)

TISSUE(S): Heart

DISEASE(S): Myocardial Infarction

SUBMITTER: Venkatesh Mallikarjun

LAB HEAD: Sarah Ewald

PROVIDER: PXD034413 | Pride | 2023-10-31

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	STOMP3protein-peptidesLFQ_50pcmissingremoved_PEAKSoutput.csv	Csv
	STOMP3protein-peptidesLFQ_50pcmissingremoved_normalisation_logObsMedian.csv	Csv
	STOMP3protein-peptidesPEAKSDB_PEAKSoutput.csv	Csv
	STOMP3protein-peptidesPEAKSPTM_PEAKSoutput.csv	Csv
	STOMP3protein-peptidesSPIDER_PEAKSoutput.csv	Csv

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Project description:Background—Mesenchymal stem cells (MSCs) have shown therapeutic potency for treating cardiovascular diseases, but their therapeutic efficacy shows significant heterogeneity depending on their tissue of origin. We herein sought to identify the optimal source of MSCs for cardiovascular disease therapy. Methods—Heart- and bone marrow (BM)-derived GFP+ cells positive for the MSCs marker, Nestin, were flow cytometrically sorted from 7-day-postnatal Nestin-GFP transgenic mice. To study their biological characteristics in vitro, we characterized their self-renewal capacity, multi-lineage differentiation ability, and surface markers. To investigate their therapeutic potential in vivo, we intramyocardially injected Nestin+ cells (3×105) into the infarct border zone of mice subjected to acute myocardial infarction (MI), and performed echocardiography and Masson’s-Trichrome staining at 1 and 3 weeks post-MI. We characterized gene profiles with RNA-sequencing analysis, and analyzed the putative reparative mechanism, that of Periostin-mediated M2 macrophages polarization, by immunofluorescence staining, qPCR, ELISA, and flow cytometry in vivo and in vitro. Results—In vitro, the heart- and BM-derived Nestin+ cells (Nes+cMSCs and Nes+bmMSCs, respectively) were found to possess self-renewal ability, show similar tri-lineage differentiation potentials, and express some MSCs-related surface markers. Importantly, Nes+cMSCs significantly improved cardiac function, attenuated left ventricular (LV) remodeling, and decreased infarct size in the mouse MI model, compared with Nes+bmMSCs- or saline-treated MI controls. Nes+cMSC treatment notably reduced the total number of CD68+ pan-macrophages while inducing the polarization of macrophages toward an anti-inflammatory M2 phenotype in ischemic myocardium, compared with the saline-treated MI control. Periostin, which was highly expressed in Nes+cMSCs, could promote the polarization of M2-subtype macrophages in vitro and in vivo. Finally, Periostin knockdown remarkably reduced the therapeutic effects of Nes+cMSCs, inhibiting the survival and LV remodeling of MI model mice and significantly decreasing the number of M2 macrophages at lesion sites. Conclusions—Nestin+ cMSCs have greater efficacy than Nestin+ bmMSCs for cardiac repair following acute MI, using a reparative mechanism that acts at least partly through Periostin-mediated M2 macrophage polarization.

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Spatially-targeted optical microproteomics timecourse of rat myocardial infarction

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