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Aging increases the susceptivity of MSCs to reactive oxygen species and impairs their therapeutic potency for myocardial infarction.

Li L, Guo Y, Zhai H, Yin Y, Zhang J, Chen H, Wang L, Li N, Liu R, Xia Y - PLoS ONE (2014)

Bottom Line: By exposing these MSCs to H2O2, we found that the adhesion of MSCs from old donors was damaged more severely.Specifically, decreased expression of integrin and reduced phosphorylation of focal adhesion kinase Src and FAK were observed.The low viability of engrafted old MSCs consequently impaired their therapeutic effectiveness, judging by the histology and function of heart.

View Article: PubMed Central - PubMed

Affiliation: First Department of Cadres, First Hospital Affiliated to General Hospital of People's Liberation Army, Beijing, China.

ABSTRACT
Myocardial infarction (MI) is one of the leading causes of death worldwide and Mesenchymal Stem Cells (MSCs) transplantation has been considered a promising therapy. Recently, it was reported that the therapeutic effectiveness of MSCs is dependent on the age of the donor, yet the underlying mechanism has not been thoroughly investigated. This study was designed to investigate whether this impaired therapeutic potency is caused by an increased susceptivity of MSCs from old donors to reactive oxygen species (ROS). The MSCs were isolated from the subcutaneous inguinal region of young (8-10 weeks) and old (18 months) Sprague-Dawley (SD) rats. By exposing these MSCs to H2O2, we found that the adhesion of MSCs from old donors was damaged more severely. Specifically, decreased expression of integrin and reduced phosphorylation of focal adhesion kinase Src and FAK were observed. Furthemore, H2O2 triggered an increased apoptosis of MSCs from old donors. To study the viability and therapeutic potency of MSCs from young and old donors in vivo, these MSCs were transplanted into acute MI model rats. We observed a more rapidly decreased survival rate of the old MSCs in the infarct region, which may be caused by their increased susceptivity to the micro-environmental ROS, as transplantation of the old MSCs with N-acetyl-L-cysteine (NAC), a ROS scavenger, protected them. The low viability of engrafted old MSCs consequently impaired their therapeutic effectiveness, judging by the histology and function of heart. Our study may help to understand the mechanism of MSCs-host interaction during MI, as well as shed light on the design of therapeutic strategy in clinic.

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Susceptibility of MSCs from young and old donors to ROS.A. Representative images of adhered cells. ‘a’ and ‘b’: Adhesion of MSCs from young and old donors without H2O2; c′ and ‘d’: Adhesion of MSCs from young and old donors under 50 µM H2O2. Bar = 100 µm. B. H2O2 influences the MSC adhesion in a dose-dependent manner. * P<0.05, **P<0.01. C. Immunofluorescence stain for expression of integrin β1 (red) in MSCs of above adhesion assay. Cell nucleus was stained with DAPI (blue). Arrows indicate the spread morphology of MSCs. Arrows indicate the spread morphology of cells. D and E. The relative expression level of integrin β1 and αV were analyzed by RT-PCR in MSCs from young and old donors with indicated treatment. The mRNA level of PBS treated MSCs from young donors was set as 100%. F-G. Phosphorylation of Src and FAK were analyzed by west-blot in MSCs from young and old donors with indicated treatment. The phosphorylation level of PBS treated young MSCs was set as 100%. ** p<0.01 compared with young and old MSCs without H2O2 treatment; ##** p<0.01 compared with young MSC with H2O2 treatment.
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pone-0111850-g002: Susceptibility of MSCs from young and old donors to ROS.A. Representative images of adhered cells. ‘a’ and ‘b’: Adhesion of MSCs from young and old donors without H2O2; c′ and ‘d’: Adhesion of MSCs from young and old donors under 50 µM H2O2. Bar = 100 µm. B. H2O2 influences the MSC adhesion in a dose-dependent manner. * P<0.05, **P<0.01. C. Immunofluorescence stain for expression of integrin β1 (red) in MSCs of above adhesion assay. Cell nucleus was stained with DAPI (blue). Arrows indicate the spread morphology of MSCs. Arrows indicate the spread morphology of cells. D and E. The relative expression level of integrin β1 and αV were analyzed by RT-PCR in MSCs from young and old donors with indicated treatment. The mRNA level of PBS treated MSCs from young donors was set as 100%. F-G. Phosphorylation of Src and FAK were analyzed by west-blot in MSCs from young and old donors with indicated treatment. The phosphorylation level of PBS treated young MSCs was set as 100%. ** p<0.01 compared with young and old MSCs without H2O2 treatment; ##** p<0.01 compared with young MSC with H2O2 treatment.

Mentions: ROS impairs adhesion of stem cells [16]. To test whether old MSCs were more susceptible to ROS, adhesion assays were performed with MSCs from young and old donors by treating them exogenously with H2O2. As shown in Figure 2A and 2B, H2O2 impaired the adhesion of MSCs in a dose dependent manner. Meanwhile, a greater decrease in the adhesion of the old MSCs, compared with that of the young MSCs at each H2O2 concentration, was observed. In addition, we evaluated the altered expressions of adhesion-related integrin molecules in H2O2-treated MSCs using immune-fluorescence (Figure 2C) and RT-PCR (Figure 2D and 2E). A significant change in cell morphology and down-regulation of integrin β1 (also termed CD29) and αV were observed in H2O2-treated old MSCs, compared with H2O2-treated young MSCs and non-treated MSCs. Furthermore, we examined the phosphorylation of kinases FAK and Src, as they play vital roles in cellular focal adhesion by regulating the actin cytoskeleton. As seen in Figure 2F-2H, H2O2 treatment significantly reduced the phosphorylation of both kinases, with old MSCs suffered more than the young ones.


Aging increases the susceptivity of MSCs to reactive oxygen species and impairs their therapeutic potency for myocardial infarction.

Li L, Guo Y, Zhai H, Yin Y, Zhang J, Chen H, Wang L, Li N, Liu R, Xia Y - PLoS ONE (2014)

Susceptibility of MSCs from young and old donors to ROS.A. Representative images of adhered cells. ‘a’ and ‘b’: Adhesion of MSCs from young and old donors without H2O2; c′ and ‘d’: Adhesion of MSCs from young and old donors under 50 µM H2O2. Bar = 100 µm. B. H2O2 influences the MSC adhesion in a dose-dependent manner. * P<0.05, **P<0.01. C. Immunofluorescence stain for expression of integrin β1 (red) in MSCs of above adhesion assay. Cell nucleus was stained with DAPI (blue). Arrows indicate the spread morphology of MSCs. Arrows indicate the spread morphology of cells. D and E. The relative expression level of integrin β1 and αV were analyzed by RT-PCR in MSCs from young and old donors with indicated treatment. The mRNA level of PBS treated MSCs from young donors was set as 100%. F-G. Phosphorylation of Src and FAK were analyzed by west-blot in MSCs from young and old donors with indicated treatment. The phosphorylation level of PBS treated young MSCs was set as 100%. ** p<0.01 compared with young and old MSCs without H2O2 treatment; ##** p<0.01 compared with young MSC with H2O2 treatment.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4230939&req=5

pone-0111850-g002: Susceptibility of MSCs from young and old donors to ROS.A. Representative images of adhered cells. ‘a’ and ‘b’: Adhesion of MSCs from young and old donors without H2O2; c′ and ‘d’: Adhesion of MSCs from young and old donors under 50 µM H2O2. Bar = 100 µm. B. H2O2 influences the MSC adhesion in a dose-dependent manner. * P<0.05, **P<0.01. C. Immunofluorescence stain for expression of integrin β1 (red) in MSCs of above adhesion assay. Cell nucleus was stained with DAPI (blue). Arrows indicate the spread morphology of MSCs. Arrows indicate the spread morphology of cells. D and E. The relative expression level of integrin β1 and αV were analyzed by RT-PCR in MSCs from young and old donors with indicated treatment. The mRNA level of PBS treated MSCs from young donors was set as 100%. F-G. Phosphorylation of Src and FAK were analyzed by west-blot in MSCs from young and old donors with indicated treatment. The phosphorylation level of PBS treated young MSCs was set as 100%. ** p<0.01 compared with young and old MSCs without H2O2 treatment; ##** p<0.01 compared with young MSC with H2O2 treatment.
Mentions: ROS impairs adhesion of stem cells [16]. To test whether old MSCs were more susceptible to ROS, adhesion assays were performed with MSCs from young and old donors by treating them exogenously with H2O2. As shown in Figure 2A and 2B, H2O2 impaired the adhesion of MSCs in a dose dependent manner. Meanwhile, a greater decrease in the adhesion of the old MSCs, compared with that of the young MSCs at each H2O2 concentration, was observed. In addition, we evaluated the altered expressions of adhesion-related integrin molecules in H2O2-treated MSCs using immune-fluorescence (Figure 2C) and RT-PCR (Figure 2D and 2E). A significant change in cell morphology and down-regulation of integrin β1 (also termed CD29) and αV were observed in H2O2-treated old MSCs, compared with H2O2-treated young MSCs and non-treated MSCs. Furthermore, we examined the phosphorylation of kinases FAK and Src, as they play vital roles in cellular focal adhesion by regulating the actin cytoskeleton. As seen in Figure 2F-2H, H2O2 treatment significantly reduced the phosphorylation of both kinases, with old MSCs suffered more than the young ones.

Bottom Line: By exposing these MSCs to H2O2, we found that the adhesion of MSCs from old donors was damaged more severely.Specifically, decreased expression of integrin and reduced phosphorylation of focal adhesion kinase Src and FAK were observed.The low viability of engrafted old MSCs consequently impaired their therapeutic effectiveness, judging by the histology and function of heart.

View Article: PubMed Central - PubMed

Affiliation: First Department of Cadres, First Hospital Affiliated to General Hospital of People's Liberation Army, Beijing, China.

ABSTRACT
Myocardial infarction (MI) is one of the leading causes of death worldwide and Mesenchymal Stem Cells (MSCs) transplantation has been considered a promising therapy. Recently, it was reported that the therapeutic effectiveness of MSCs is dependent on the age of the donor, yet the underlying mechanism has not been thoroughly investigated. This study was designed to investigate whether this impaired therapeutic potency is caused by an increased susceptivity of MSCs from old donors to reactive oxygen species (ROS). The MSCs were isolated from the subcutaneous inguinal region of young (8-10 weeks) and old (18 months) Sprague-Dawley (SD) rats. By exposing these MSCs to H2O2, we found that the adhesion of MSCs from old donors was damaged more severely. Specifically, decreased expression of integrin and reduced phosphorylation of focal adhesion kinase Src and FAK were observed. Furthemore, H2O2 triggered an increased apoptosis of MSCs from old donors. To study the viability and therapeutic potency of MSCs from young and old donors in vivo, these MSCs were transplanted into acute MI model rats. We observed a more rapidly decreased survival rate of the old MSCs in the infarct region, which may be caused by their increased susceptivity to the micro-environmental ROS, as transplantation of the old MSCs with N-acetyl-L-cysteine (NAC), a ROS scavenger, protected them. The low viability of engrafted old MSCs consequently impaired their therapeutic effectiveness, judging by the histology and function of heart. Our study may help to understand the mechanism of MSCs-host interaction during MI, as well as shed light on the design of therapeutic strategy in clinic.

Show MeSH
Related in: MedlinePlus