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Microvesicles derived from human umbilical cord mesenchymal stem cells facilitate tubular epithelial cell dedifferentiation and growth via hepatocyte growth factor induction.

Ju GQ, Cheng J, Zhong L, Wu S, Zou XY, Zhang GY, Gu D, Miao S, Zhu YJ, Sun J, Du T - PLoS ONE (2015)

Bottom Line: In a rat model of unilateral AKI, the administration of MVs promoted kidney recovery.RNase treatment abrogated all MV effects.Intriguingly, these effects were completely abrogated by either c-Met inhibitor or MEK inhibitor, suggesting that HGF induction is a crucial contributor to the acceleration of cell dedifferentiation and growth.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Shanghai Children's Medical Center, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.

ABSTRACT
During acute kidney injury (AKI), tubular cell dedifferentiation initiates cell regeneration; hepatocyte growth factor (HGF) is involved in modulating cell dedifferentiation. Mesenchymal stem cell (MSC)-derived microvesicles (MVs) deliver RNA into injured tubular cells and alter their gene expression, thus regenerating these cells. We boldly speculated that MVs might induce HGF synthesis via RNA transfer, thereby facilitating tubular cell dedifferentiation and regeneration. In a rat model of unilateral AKI, the administration of MVs promoted kidney recovery. One of the mechanisms of action is the acceleration of tubular cell dedifferentiation and growth. Both in vivo and in vitro, rat HGF expression in damaged rat tubular cells was greatly enhanced by MV treatment. In addition, human HGF mRNA present in MVs was delivered into rat tubular cells and translated into the HGF protein as another mechanism of HGF induction. RNase treatment abrogated all MV effects. In the in vitro experimental setting, the conditioned medium of MV-treated injured tubular cells, which contains a higher concentration of HGF, strongly stimulated cell dedifferentiation and growth, as well as Erk1/2 signaling activation. Intriguingly, these effects were completely abrogated by either c-Met inhibitor or MEK inhibitor, suggesting that HGF induction is a crucial contributor to the acceleration of cell dedifferentiation and growth. All these findings indicate that MV-induced HGF synthesis in damaged tubular cells via RNA transfer facilitates cell dedifferentiation and growth, which are important regenerative mechanisms.

No MeSH data available.


Related in: MedlinePlus

At 48 h post-injury, kidney HGF gene and protein expression is substantially enhanced by MV administration.(A)-(D) HGF gene expression in injured kidney tissues. MV administration led to a significant up-regulation of kidney HGF gene expression. The examination of rat HGF expression in kidney tissues using species-specific primers also indicated a similar result. As negative controls, no rat HGF mRNA was identified in MVs or in the cells of origin (hUC-MSCs). RNase pretreatment abolished the effect of MVs. By contrast, EGF, IGF-1 or TGFβ1 gene expression was not altered by MV administration. Gene expression levels in sham-treated samples were regarded as the baseline levels (dotted line). The relative expression levels of each gene were calculated using the 2−ΔΔCt method. The data were collected from 6 rats for each experimental condition. *P<0.05, AKI+MVs vs. AKI+VEHICLE; #P<0.05, AKI+RNase-MVs vs. AKI+MVs. (E) Densitometric analysis of kidney HGF protein expression. At 48 h, MV administration also resulted in a prominent increase in kidney HGF protein expression. This effect was abrogated by RNase pre-treatment. The values in the graph are expressed as densitometric ratios of HGF/GAPDH as fold changes compared with the control (sham-operated samples) (dotted line). *P<0.05, AKI+MVs vs. AKI+VEHICLE; #P<0.05, AKI+RNase-MVs vs. AKI+MVs; xP<0.01, AKI+VEHICLE vs. SHAM. (F) Representative gel photograph of kidney HGF protein expression. (G) HGF staining on kidney sections. Most of the positive staining was observed in damaged tubular cells. HGF staining of injured tubular cells was remarkably intensified in MV-treated animals at 48 h post-injury. Magnification, ×40.
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pone.0121534.g003: At 48 h post-injury, kidney HGF gene and protein expression is substantially enhanced by MV administration.(A)-(D) HGF gene expression in injured kidney tissues. MV administration led to a significant up-regulation of kidney HGF gene expression. The examination of rat HGF expression in kidney tissues using species-specific primers also indicated a similar result. As negative controls, no rat HGF mRNA was identified in MVs or in the cells of origin (hUC-MSCs). RNase pretreatment abolished the effect of MVs. By contrast, EGF, IGF-1 or TGFβ1 gene expression was not altered by MV administration. Gene expression levels in sham-treated samples were regarded as the baseline levels (dotted line). The relative expression levels of each gene were calculated using the 2−ΔΔCt method. The data were collected from 6 rats for each experimental condition. *P<0.05, AKI+MVs vs. AKI+VEHICLE; #P<0.05, AKI+RNase-MVs vs. AKI+MVs. (E) Densitometric analysis of kidney HGF protein expression. At 48 h, MV administration also resulted in a prominent increase in kidney HGF protein expression. This effect was abrogated by RNase pre-treatment. The values in the graph are expressed as densitometric ratios of HGF/GAPDH as fold changes compared with the control (sham-operated samples) (dotted line). *P<0.05, AKI+MVs vs. AKI+VEHICLE; #P<0.05, AKI+RNase-MVs vs. AKI+MVs; xP<0.01, AKI+VEHICLE vs. SHAM. (F) Representative gel photograph of kidney HGF protein expression. (G) HGF staining on kidney sections. Most of the positive staining was observed in damaged tubular cells. HGF staining of injured tubular cells was remarkably intensified in MV-treated animals at 48 h post-injury. Magnification, ×40.

Mentions: Considering the role of growth factors produced by the tubular epithelium in tubular cell dedifferentiation and growth, we examined the effect of MVs on growth factor expression. In AKI animals, kidney HGF expression was greatly enhanced by MV administration at 48 h post-injury, as demonstrated by RT-PCR and by western blot analysis (P<0.05, Fig. 3D, 3F-G). Immunohistochemistry staining also revealed that a substantial intensification of HGF staining was observed in damaged tubular cells (P<0.05, Fig. 3H). By contrast, TGF-β1, IGF-1 or EGF expression was not significantly altered by MV administration (Fig. 3A-3C). Using species-specific (rat) primers, we also screened for rat HGF mRNA in damaged kidney tissues. We found that MV administration also markedly induced rat HGF expression (Fig. 3E). As a negative control, no rat HGF mRNA was detected in MVs or in their cells of origin (hUC-MSCs) (P<0.05, data not shown).


Microvesicles derived from human umbilical cord mesenchymal stem cells facilitate tubular epithelial cell dedifferentiation and growth via hepatocyte growth factor induction.

Ju GQ, Cheng J, Zhong L, Wu S, Zou XY, Zhang GY, Gu D, Miao S, Zhu YJ, Sun J, Du T - PLoS ONE (2015)

At 48 h post-injury, kidney HGF gene and protein expression is substantially enhanced by MV administration.(A)-(D) HGF gene expression in injured kidney tissues. MV administration led to a significant up-regulation of kidney HGF gene expression. The examination of rat HGF expression in kidney tissues using species-specific primers also indicated a similar result. As negative controls, no rat HGF mRNA was identified in MVs or in the cells of origin (hUC-MSCs). RNase pretreatment abolished the effect of MVs. By contrast, EGF, IGF-1 or TGFβ1 gene expression was not altered by MV administration. Gene expression levels in sham-treated samples were regarded as the baseline levels (dotted line). The relative expression levels of each gene were calculated using the 2−ΔΔCt method. The data were collected from 6 rats for each experimental condition. *P<0.05, AKI+MVs vs. AKI+VEHICLE; #P<0.05, AKI+RNase-MVs vs. AKI+MVs. (E) Densitometric analysis of kidney HGF protein expression. At 48 h, MV administration also resulted in a prominent increase in kidney HGF protein expression. This effect was abrogated by RNase pre-treatment. The values in the graph are expressed as densitometric ratios of HGF/GAPDH as fold changes compared with the control (sham-operated samples) (dotted line). *P<0.05, AKI+MVs vs. AKI+VEHICLE; #P<0.05, AKI+RNase-MVs vs. AKI+MVs; xP<0.01, AKI+VEHICLE vs. SHAM. (F) Representative gel photograph of kidney HGF protein expression. (G) HGF staining on kidney sections. Most of the positive staining was observed in damaged tubular cells. HGF staining of injured tubular cells was remarkably intensified in MV-treated animals at 48 h post-injury. Magnification, ×40.
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Related In: Results  -  Collection

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pone.0121534.g003: At 48 h post-injury, kidney HGF gene and protein expression is substantially enhanced by MV administration.(A)-(D) HGF gene expression in injured kidney tissues. MV administration led to a significant up-regulation of kidney HGF gene expression. The examination of rat HGF expression in kidney tissues using species-specific primers also indicated a similar result. As negative controls, no rat HGF mRNA was identified in MVs or in the cells of origin (hUC-MSCs). RNase pretreatment abolished the effect of MVs. By contrast, EGF, IGF-1 or TGFβ1 gene expression was not altered by MV administration. Gene expression levels in sham-treated samples were regarded as the baseline levels (dotted line). The relative expression levels of each gene were calculated using the 2−ΔΔCt method. The data were collected from 6 rats for each experimental condition. *P<0.05, AKI+MVs vs. AKI+VEHICLE; #P<0.05, AKI+RNase-MVs vs. AKI+MVs. (E) Densitometric analysis of kidney HGF protein expression. At 48 h, MV administration also resulted in a prominent increase in kidney HGF protein expression. This effect was abrogated by RNase pre-treatment. The values in the graph are expressed as densitometric ratios of HGF/GAPDH as fold changes compared with the control (sham-operated samples) (dotted line). *P<0.05, AKI+MVs vs. AKI+VEHICLE; #P<0.05, AKI+RNase-MVs vs. AKI+MVs; xP<0.01, AKI+VEHICLE vs. SHAM. (F) Representative gel photograph of kidney HGF protein expression. (G) HGF staining on kidney sections. Most of the positive staining was observed in damaged tubular cells. HGF staining of injured tubular cells was remarkably intensified in MV-treated animals at 48 h post-injury. Magnification, ×40.
Mentions: Considering the role of growth factors produced by the tubular epithelium in tubular cell dedifferentiation and growth, we examined the effect of MVs on growth factor expression. In AKI animals, kidney HGF expression was greatly enhanced by MV administration at 48 h post-injury, as demonstrated by RT-PCR and by western blot analysis (P<0.05, Fig. 3D, 3F-G). Immunohistochemistry staining also revealed that a substantial intensification of HGF staining was observed in damaged tubular cells (P<0.05, Fig. 3H). By contrast, TGF-β1, IGF-1 or EGF expression was not significantly altered by MV administration (Fig. 3A-3C). Using species-specific (rat) primers, we also screened for rat HGF mRNA in damaged kidney tissues. We found that MV administration also markedly induced rat HGF expression (Fig. 3E). As a negative control, no rat HGF mRNA was detected in MVs or in their cells of origin (hUC-MSCs) (P<0.05, data not shown).

Bottom Line: In a rat model of unilateral AKI, the administration of MVs promoted kidney recovery.RNase treatment abrogated all MV effects.Intriguingly, these effects were completely abrogated by either c-Met inhibitor or MEK inhibitor, suggesting that HGF induction is a crucial contributor to the acceleration of cell dedifferentiation and growth.

View Article: PubMed Central - PubMed

Affiliation: Department of Urology, Shanghai Children's Medical Center, School of Medicine, Shanghai Jiao Tong University, Shanghai, China.

ABSTRACT
During acute kidney injury (AKI), tubular cell dedifferentiation initiates cell regeneration; hepatocyte growth factor (HGF) is involved in modulating cell dedifferentiation. Mesenchymal stem cell (MSC)-derived microvesicles (MVs) deliver RNA into injured tubular cells and alter their gene expression, thus regenerating these cells. We boldly speculated that MVs might induce HGF synthesis via RNA transfer, thereby facilitating tubular cell dedifferentiation and regeneration. In a rat model of unilateral AKI, the administration of MVs promoted kidney recovery. One of the mechanisms of action is the acceleration of tubular cell dedifferentiation and growth. Both in vivo and in vitro, rat HGF expression in damaged rat tubular cells was greatly enhanced by MV treatment. In addition, human HGF mRNA present in MVs was delivered into rat tubular cells and translated into the HGF protein as another mechanism of HGF induction. RNase treatment abrogated all MV effects. In the in vitro experimental setting, the conditioned medium of MV-treated injured tubular cells, which contains a higher concentration of HGF, strongly stimulated cell dedifferentiation and growth, as well as Erk1/2 signaling activation. Intriguingly, these effects were completely abrogated by either c-Met inhibitor or MEK inhibitor, suggesting that HGF induction is a crucial contributor to the acceleration of cell dedifferentiation and growth. All these findings indicate that MV-induced HGF synthesis in damaged tubular cells via RNA transfer facilitates cell dedifferentiation and growth, which are important regenerative mechanisms.

No MeSH data available.


Related in: MedlinePlus