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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

Absent of miR-223 impairs MSC-induced protection against sepsis-triggered cardiac injury.(A–H) Characterization of MSCs derived from bone marrow of WT mice (A–D) and miR-223 KO mice (E–H). (I) Total RNA was isolated from MSCs and performed RT-PCR analysis, and showed that miR-223-5p and -3p both are absent in KO-MSCs. U6 snRNA was used as internal control. (J) Injection of miR-223-KO MSCs (n = 14) at 1 h post-CLP did not improve animal survival, whereas the survival rate was significantly improved in WT-MSC-treated mice (n = 12), compared with PBS controls (n = 10, *p < 0.05). (K–M) CLP-induced cardiac depression, measured by echocardiography (K), was significantly attenuated in WT-MSC-treated mice, but not in KO-MSC-treated mice. *p < 0.05, vs. sham group; #p < 0.05, vs. CLP+PBS control group; n = 6 for sham group, n = 10 for CLP + PBS, n = 12 for WT- and KO-MSC-treated groups. (N) Representative TUNEL staining for detection of apoptotic cardiomyocytes (green dots indicated by arrows, upper row). Cardiomyocytes were stained with α-actin (Red) and nuclei were stained with DAPI (blue), which were merged together, indicated in the lower row. (O) Quantitative results of TUNEL staining, which was further confirmed by ELISA measurement of histone-associated DNA fragmentation (P). (n = 5 hearts, *p < 0.05, vs. shams; #p < 0.05 vs. CLP+PBS control group).
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f1: Absent of miR-223 impairs MSC-induced protection against sepsis-triggered cardiac injury.(A–H) Characterization of MSCs derived from bone marrow of WT mice (A–D) and miR-223 KO mice (E–H). (I) Total RNA was isolated from MSCs and performed RT-PCR analysis, and showed that miR-223-5p and -3p both are absent in KO-MSCs. U6 snRNA was used as internal control. (J) Injection of miR-223-KO MSCs (n = 14) at 1 h post-CLP did not improve animal survival, whereas the survival rate was significantly improved in WT-MSC-treated mice (n = 12), compared with PBS controls (n = 10, *p < 0.05). (K–M) CLP-induced cardiac depression, measured by echocardiography (K), was significantly attenuated in WT-MSC-treated mice, but not in KO-MSC-treated mice. *p < 0.05, vs. sham group; #p < 0.05, vs. CLP+PBS control group; n = 6 for sham group, n = 10 for CLP + PBS, n = 12 for WT- and KO-MSC-treated groups. (N) Representative TUNEL staining for detection of apoptotic cardiomyocytes (green dots indicated by arrows, upper row). Cardiomyocytes were stained with α-actin (Red) and nuclei were stained with DAPI (blue), which were merged together, indicated in the lower row. (O) Quantitative results of TUNEL staining, which was further confirmed by ELISA measurement of histone-associated DNA fragmentation (P). (n = 5 hearts, *p < 0.05, vs. shams; #p < 0.05 vs. CLP+PBS control group).

Mentions: To address whether miR-223 contributes to MSC-induced protection in sepsis, we harvested MSCs from bone marrow of female miR-223 KO (miR-223−/−) mice. MSCs derived from female wild-type (WT) bone marrow were used as controls. MiR-223-KO MSCs were phenotypically similar to WTs, and expressed mesenchymal markers (i.e. CD29 and Sca-1), but not hematopoietic markers (i.e. CD34) (Fig. 1A–H). We also validated that the expression of both strands (miR-223-5p and miR-223-3p) was deficient in MSCs collected from miR-223-KO mice (Fig. 1I). To understand the functional significance of miR-223 in MSC-mediated protection against sepsis injury, miR-223-KO MSCs and controls were administered intravenously to male WT mice at 1 h post-CLP surgery. We observed that survival rate was significantly improved in CLP-mice treated with WT-MSCs, as evidenced by 58% of the mice (n = 12) survived until the end of day 3, when only 20% of PBS-treated mice survived (n = 10) (Fig. 1J). These results are consistent with previous reports3. However, CLP-mice treated with miR-223- MSCs (n = 14) did not exhibit an improved survival, compared to PBS-treated controls (Fig. 1J). This suggests that miR-223 may be essential for MSC-elicited beneficial effects on survival during polymicrobial sepsis.


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)

Absent of miR-223 impairs MSC-induced protection against sepsis-triggered cardiac injury.(A–H) Characterization of MSCs derived from bone marrow of WT mice (A–D) and miR-223 KO mice (E–H). (I) Total RNA was isolated from MSCs and performed RT-PCR analysis, and showed that miR-223-5p and -3p both are absent in KO-MSCs. U6 snRNA was used as internal control. (J) Injection of miR-223-KO MSCs (n = 14) at 1 h post-CLP did not improve animal survival, whereas the survival rate was significantly improved in WT-MSC-treated mice (n = 12), compared with PBS controls (n = 10, *p < 0.05). (K–M) CLP-induced cardiac depression, measured by echocardiography (K), was significantly attenuated in WT-MSC-treated mice, but not in KO-MSC-treated mice. *p < 0.05, vs. sham group; #p < 0.05, vs. CLP+PBS control group; n = 6 for sham group, n = 10 for CLP + PBS, n = 12 for WT- and KO-MSC-treated groups. (N) Representative TUNEL staining for detection of apoptotic cardiomyocytes (green dots indicated by arrows, upper row). Cardiomyocytes were stained with α-actin (Red) and nuclei were stained with DAPI (blue), which were merged together, indicated in the lower row. (O) Quantitative results of TUNEL staining, which was further confirmed by ELISA measurement of histone-associated DNA fragmentation (P). (n = 5 hearts, *p < 0.05, vs. shams; #p < 0.05 vs. CLP+PBS control group).
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f1: Absent of miR-223 impairs MSC-induced protection against sepsis-triggered cardiac injury.(A–H) Characterization of MSCs derived from bone marrow of WT mice (A–D) and miR-223 KO mice (E–H). (I) Total RNA was isolated from MSCs and performed RT-PCR analysis, and showed that miR-223-5p and -3p both are absent in KO-MSCs. U6 snRNA was used as internal control. (J) Injection of miR-223-KO MSCs (n = 14) at 1 h post-CLP did not improve animal survival, whereas the survival rate was significantly improved in WT-MSC-treated mice (n = 12), compared with PBS controls (n = 10, *p < 0.05). (K–M) CLP-induced cardiac depression, measured by echocardiography (K), was significantly attenuated in WT-MSC-treated mice, but not in KO-MSC-treated mice. *p < 0.05, vs. sham group; #p < 0.05, vs. CLP+PBS control group; n = 6 for sham group, n = 10 for CLP + PBS, n = 12 for WT- and KO-MSC-treated groups. (N) Representative TUNEL staining for detection of apoptotic cardiomyocytes (green dots indicated by arrows, upper row). Cardiomyocytes were stained with α-actin (Red) and nuclei were stained with DAPI (blue), which were merged together, indicated in the lower row. (O) Quantitative results of TUNEL staining, which was further confirmed by ELISA measurement of histone-associated DNA fragmentation (P). (n = 5 hearts, *p < 0.05, vs. shams; #p < 0.05 vs. CLP+PBS control group).
Mentions: To address whether miR-223 contributes to MSC-induced protection in sepsis, we harvested MSCs from bone marrow of female miR-223 KO (miR-223−/−) mice. MSCs derived from female wild-type (WT) bone marrow were used as controls. MiR-223-KO MSCs were phenotypically similar to WTs, and expressed mesenchymal markers (i.e. CD29 and Sca-1), but not hematopoietic markers (i.e. CD34) (Fig. 1A–H). We also validated that the expression of both strands (miR-223-5p and miR-223-3p) was deficient in MSCs collected from miR-223-KO mice (Fig. 1I). To understand the functional significance of miR-223 in MSC-mediated protection against sepsis injury, miR-223-KO MSCs and controls were administered intravenously to male WT mice at 1 h post-CLP surgery. We observed that survival rate was significantly improved in CLP-mice treated with WT-MSCs, as evidenced by 58% of the mice (n = 12) survived until the end of day 3, when only 20% of PBS-treated mice survived (n = 10) (Fig. 1J). These results are consistent with previous reports3. However, CLP-mice treated with miR-223- MSCs (n = 14) did not exhibit an improved survival, compared to PBS-treated controls (Fig. 1J). This suggests that miR-223 may be essential for MSC-elicited beneficial effects on survival during polymicrobial sepsis.

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