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Exosomal miR-223 Contributes to Mesenchymal Stem Cell-Elicited Cardioprotection in Polymicrobial Sepsis.

Wang X, Gu H, Qin D, Yang L, Huang W, Essandoh K, Wang Y, Caldwell CC, Peng T, Zingarelli B, Fan GC - Sci Rep (2015)

Bottom Line: However, WT-MSCs were able to provide protection which was associated with exosome release.Conversely, WT-MSC-derived-exosomes displayed protective effects.By contrast, WT-exosomes encased higher levels of miR-223, which could be delivered to cardiomyocytes, resulting in down-regulation of Sema3A and Stat3.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, Cincinnati, OH, USA.

ABSTRACT
Mesenchymal stem cells (MSCs) have been shown to elicit cardio-protective effects in sepsis. However, the underlying mechanism remains obscure. While recent studies have indicated that miR-223 is highly enriched in MSC-derived exosomes, whether exosomal miR-223 contributes to MSC-mediated cardio-protection in sepsis is unknown. In this study, loss-of-function approach was utilized, and sepsis was induced by cecal ligation and puncture (CLP). We observed that injection of miR-223-KO MSCs at 1 h post-CLP did not confer protection against CLP-triggered cardiac dysfunction, apoptosis and inflammatory response. However, WT-MSCs were able to provide protection which was associated with exosome release. Next, treatment of CLP mice with exosomes released from miR-223-KO MSCs significantly exaggerated sepsis-induced injury. Conversely, WT-MSC-derived-exosomes displayed protective effects. Mechanistically, we identified that miR-223-KO exosomes contained higher levels of Sema3A and Stat3, two known targets of miR-223 (5p &3p), than WT-exosomes. Accordingly, these exosomal proteins were transferred to cardiomyocytes, leading to increased inflammation and cell death. By contrast, WT-exosomes encased higher levels of miR-223, which could be delivered to cardiomyocytes, resulting in down-regulation of Sema3A and Stat3. These data for the first time indicate that exosomal miR-223 plays an essential role for MSC-induced cardio-protection in sepsis.

No MeSH data available.


Related in: MedlinePlus

Blockade of exosome release negates the inhibitory effects of WT-MSCs on LPS-triggered inflammatory cytokine production in macrophages and LPS-induced cardiomyocyte death.WT-MSCs significantly suppressed the production of TNF-α (A), IL-1β (B), and IL-6 (C) in co-cultured RAW264.7 cells upon LPS challenge. Such inhibitory effects were offset by addition of GW4869 (20 μM). n = 3 wells, *p < 0.05, vs. RAW264.7 cultured alone. Similar results were observed in three additional, independent experiments. (D) A diagram of cell co-culture system in which cardiomyocytes were cultured in the lower chamber of a 12-well plate pre-coated with laminin (10 μg/ml) and MSCs were cultured in the upper chamber of a 12-well insert. (E) LPS exposure significantly decreased cardiomyocyte survival, whereas it is greatly improved by co-culturing with WT-MSCs. n = 3 wells, *p < 0.05, vs. cardiomyocytes cultured alone. Addition of GW4869 (20 μM) offset WT-MSC-elicited protective effects on LPS-triggered myocyte death. Similar results were observed in two additional, independent experiments.
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f4: Blockade of exosome release negates the inhibitory effects of WT-MSCs on LPS-triggered inflammatory cytokine production in macrophages and LPS-induced cardiomyocyte death.WT-MSCs significantly suppressed the production of TNF-α (A), IL-1β (B), and IL-6 (C) in co-cultured RAW264.7 cells upon LPS challenge. Such inhibitory effects were offset by addition of GW4869 (20 μM). n = 3 wells, *p < 0.05, vs. RAW264.7 cultured alone. Similar results were observed in three additional, independent experiments. (D) A diagram of cell co-culture system in which cardiomyocytes were cultured in the lower chamber of a 12-well plate pre-coated with laminin (10 μg/ml) and MSCs were cultured in the upper chamber of a 12-well insert. (E) LPS exposure significantly decreased cardiomyocyte survival, whereas it is greatly improved by co-culturing with WT-MSCs. n = 3 wells, *p < 0.05, vs. cardiomyocytes cultured alone. Addition of GW4869 (20 μM) offset WT-MSC-elicited protective effects on LPS-triggered myocyte death. Similar results were observed in two additional, independent experiments.

Mentions: In recent years, exosomes have emerged as one of the most important players for both local and long-distance cell-cell communication1011. To examine whether WT-MSC-induced effects are associated with their released exosomes, we pre-treated WT-MSCs with GW4869 (20 μM), an inhibitor of exosome biogenesis and release13192627, then co-cultured with macrophages, followed by LPS stimulation for 12 h. In this study, we observed that the TNF-α concentration was reduced by 80% in co-culture supernatants, compared with macrophages cultured alone (p < 0.05) (Fig. 4A). However, the concentration of TNF-α in co-culture supernatants was similar between groups upon addition of GW4869 (Fig. 4A). Likewise, the secretion of IL-1β and IL-6 from macrophages was remarkably inhibited by WT-MSCs, but such a suppression was able to escape from pre-treatment with GW4869 (Fig. 4B,C). These results indicate that blockade of exosome generation with GW4869 impairs MSC-elicited anti-inflammatory effects.


Exosomal miR-223 Contributes to Mesenchymal Stem Cell-Elicited Cardioprotection in Polymicrobial Sepsis.

Wang X, Gu H, Qin D, Yang L, Huang W, Essandoh K, Wang Y, Caldwell CC, Peng T, Zingarelli B, Fan GC - Sci Rep (2015)

Blockade of exosome release negates the inhibitory effects of WT-MSCs on LPS-triggered inflammatory cytokine production in macrophages and LPS-induced cardiomyocyte death.WT-MSCs significantly suppressed the production of TNF-α (A), IL-1β (B), and IL-6 (C) in co-cultured RAW264.7 cells upon LPS challenge. Such inhibitory effects were offset by addition of GW4869 (20 μM). n = 3 wells, *p < 0.05, vs. RAW264.7 cultured alone. Similar results were observed in three additional, independent experiments. (D) A diagram of cell co-culture system in which cardiomyocytes were cultured in the lower chamber of a 12-well plate pre-coated with laminin (10 μg/ml) and MSCs were cultured in the upper chamber of a 12-well insert. (E) LPS exposure significantly decreased cardiomyocyte survival, whereas it is greatly improved by co-culturing with WT-MSCs. n = 3 wells, *p < 0.05, vs. cardiomyocytes cultured alone. Addition of GW4869 (20 μM) offset WT-MSC-elicited protective effects on LPS-triggered myocyte death. Similar results were observed in two additional, independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4562230&req=5

f4: Blockade of exosome release negates the inhibitory effects of WT-MSCs on LPS-triggered inflammatory cytokine production in macrophages and LPS-induced cardiomyocyte death.WT-MSCs significantly suppressed the production of TNF-α (A), IL-1β (B), and IL-6 (C) in co-cultured RAW264.7 cells upon LPS challenge. Such inhibitory effects were offset by addition of GW4869 (20 μM). n = 3 wells, *p < 0.05, vs. RAW264.7 cultured alone. Similar results were observed in three additional, independent experiments. (D) A diagram of cell co-culture system in which cardiomyocytes were cultured in the lower chamber of a 12-well plate pre-coated with laminin (10 μg/ml) and MSCs were cultured in the upper chamber of a 12-well insert. (E) LPS exposure significantly decreased cardiomyocyte survival, whereas it is greatly improved by co-culturing with WT-MSCs. n = 3 wells, *p < 0.05, vs. cardiomyocytes cultured alone. Addition of GW4869 (20 μM) offset WT-MSC-elicited protective effects on LPS-triggered myocyte death. Similar results were observed in two additional, independent experiments.
Mentions: In recent years, exosomes have emerged as one of the most important players for both local and long-distance cell-cell communication1011. To examine whether WT-MSC-induced effects are associated with their released exosomes, we pre-treated WT-MSCs with GW4869 (20 μM), an inhibitor of exosome biogenesis and release13192627, then co-cultured with macrophages, followed by LPS stimulation for 12 h. In this study, we observed that the TNF-α concentration was reduced by 80% in co-culture supernatants, compared with macrophages cultured alone (p < 0.05) (Fig. 4A). However, the concentration of TNF-α in co-culture supernatants was similar between groups upon addition of GW4869 (Fig. 4A). Likewise, the secretion of IL-1β and IL-6 from macrophages was remarkably inhibited by WT-MSCs, but such a suppression was able to escape from pre-treatment with GW4869 (Fig. 4B,C). These results indicate that blockade of exosome generation with GW4869 impairs MSC-elicited anti-inflammatory effects.

Bottom Line: However, WT-MSCs were able to provide protection which was associated with exosome release.Conversely, WT-MSC-derived-exosomes displayed protective effects.By contrast, WT-exosomes encased higher levels of miR-223, which could be delivered to cardiomyocytes, resulting in down-regulation of Sema3A and Stat3.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, Cincinnati, OH, USA.

ABSTRACT
Mesenchymal stem cells (MSCs) have been shown to elicit cardio-protective effects in sepsis. However, the underlying mechanism remains obscure. While recent studies have indicated that miR-223 is highly enriched in MSC-derived exosomes, whether exosomal miR-223 contributes to MSC-mediated cardio-protection in sepsis is unknown. In this study, loss-of-function approach was utilized, and sepsis was induced by cecal ligation and puncture (CLP). We observed that injection of miR-223-KO MSCs at 1 h post-CLP did not confer protection against CLP-triggered cardiac dysfunction, apoptosis and inflammatory response. However, WT-MSCs were able to provide protection which was associated with exosome release. Next, treatment of CLP mice with exosomes released from miR-223-KO MSCs significantly exaggerated sepsis-induced injury. Conversely, WT-MSC-derived-exosomes displayed protective effects. Mechanistically, we identified that miR-223-KO exosomes contained higher levels of Sema3A and Stat3, two known targets of miR-223 (5p &3p), than WT-exosomes. Accordingly, these exosomal proteins were transferred to cardiomyocytes, leading to increased inflammation and cell death. By contrast, WT-exosomes encased higher levels of miR-223, which could be delivered to cardiomyocytes, resulting in down-regulation of Sema3A and Stat3. These data for the first time indicate that exosomal miR-223 plays an essential role for MSC-induced cardio-protection in sepsis.

No MeSH data available.


Related in: MedlinePlus