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Involvement of hepatic macrophages in the antifibrotic effect of IGF-I-overexpressing mesenchymal stromal cells

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ABSTRACT

Background: Cirrhosis is a major health problem worldwide and new therapies are needed. Hepatic macrophages (hMø) have a pivotal role in liver fibrosis, being able to act in both its promotion and its resolution. It is well-known that mesenchymal stromal cells (MSCs) can modulate the immune/inflammatory cells. However, the effects of MSCs over hMø in the context of liver fibrosis remain unclear. We previously described evidence of the antifibrotic effects of in vivo applying MSCs, which were enhanced by forced overexpression of insulin-like growth factor 1 (AdIGF-I-MSCs). The aim of this work was to analyze the effect of MSCs on hMø behavior in the context of liver fibrosis resolution.

Methods: Fibrosis was induced in BALB/c mice by chronic administration of thioacetamide (8 weeks). In vivo gene expression analyses, in vitro experiments using hMø isolated from the nonparenchymal liver cells fraction, and in vivo experiments with depletion of Mø were performed.

Results: One day after treatment, hMø from fibrotic livers of MSCs-treated animals showed reduced pro-inflammatory and pro-fibrogenic gene expression profiles. These shifts were more pronounced in AdIGF-I-MSCs condition. This group showed a significant upregulation in the expression of arginase-1 and a higher downregulation of iNOS expression thus suggesting decreased levels of oxidative stress. An upregulation in IGF-I and HGF expression was observed in hMø from AdIGF-I-MSCs-treated mice suggesting a restorative phenotype in these cells. Factors secreted by hMø, preconditioned with MSCs supernatant, caused a reduction in the expression levels of hepatic stellate cells pro-fibrogenic and activation markers. Interestingly, hMø depletion abrogated the therapeutic effect achieved with AdIGF-I-MSCs therapy. Expression profile analyses for cell cycle markers were performed on fibrotic livers after treatment with AdIGF-I-MSCs and showed a significant regulation in genes related to DNA synthesis and repair quality control, cell cycle progression, and DNA damage/cellular stress compatible with early induction of pro-regenerative and hepatoprotective mechanisms. Moreover, depletion of hMø abrogated such effects on the expression of the most highly regulated genes.

Conclusions: Our results indicate that AdIGF-I-MSCs are able to induce a pro-fibrotic to resolutive phenotype shift on hepatic macrophages, which is a key early event driving liver fibrosis amelioration.

Electronic supplementary material: The online version of this article (doi:10.1186/s13287-016-0424-y) contains supplementary material, which is available to authorized users.

No MeSH data available.


Incidence of macrophages in fibrotic liver tissue after MSC treatments. a Representative microphotographs of liver sections stained for F4/80, at 1 day after saline, AdGFP-MSCs or AdIGF-I-MSCs administration. b Quantitative graph of F4/80-positive cells in liver sections. Bars represent the average of positive cells/field ± SEM from ten representative visual fields. ANOVA Tukey’s post-test, **p < 0.01 and ***p < 0.001 vs. saline-treated mice
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Fig1: Incidence of macrophages in fibrotic liver tissue after MSC treatments. a Representative microphotographs of liver sections stained for F4/80, at 1 day after saline, AdGFP-MSCs or AdIGF-I-MSCs administration. b Quantitative graph of F4/80-positive cells in liver sections. Bars represent the average of positive cells/field ± SEM from ten representative visual fields. ANOVA Tukey’s post-test, **p < 0.01 and ***p < 0.001 vs. saline-treated mice

Mentions: We previously showed that MSCs can be efficiently transduced with recombinant adenovirus with sustained significant levels of transgene expression up to 8 days post-infection. No significant changes in MSCs immunological properties were found after their genetic manipulation. Moreover, systemic administration of AdIGF-I-MSCs further reduced the collagen deposition and the degree of HeSCs activation. Considering the well-known immune-modulatory properties of MSCs, we asked whether they might induce changes in numbers of hMø in a TAA chronic model of hepatic fibrosis. Indeed, increased numbers of F4/80+ macrophages were found in liver sections from mice treated with AdGFP-MSCs and AdIGF-I-MSCs when compared to vehicle, at 1 day after treatment (Fig. 1a,b).Fig. 1


Involvement of hepatic macrophages in the antifibrotic effect of IGF-I-overexpressing mesenchymal stromal cells
Incidence of macrophages in fibrotic liver tissue after MSC treatments. a Representative microphotographs of liver sections stained for F4/80, at 1 day after saline, AdGFP-MSCs or AdIGF-I-MSCs administration. b Quantitative graph of F4/80-positive cells in liver sections. Bars represent the average of positive cells/field ± SEM from ten representative visual fields. ANOVA Tukey’s post-test, **p < 0.01 and ***p < 0.001 vs. saline-treated mice
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5120504&req=5

Fig1: Incidence of macrophages in fibrotic liver tissue after MSC treatments. a Representative microphotographs of liver sections stained for F4/80, at 1 day after saline, AdGFP-MSCs or AdIGF-I-MSCs administration. b Quantitative graph of F4/80-positive cells in liver sections. Bars represent the average of positive cells/field ± SEM from ten representative visual fields. ANOVA Tukey’s post-test, **p < 0.01 and ***p < 0.001 vs. saline-treated mice
Mentions: We previously showed that MSCs can be efficiently transduced with recombinant adenovirus with sustained significant levels of transgene expression up to 8 days post-infection. No significant changes in MSCs immunological properties were found after their genetic manipulation. Moreover, systemic administration of AdIGF-I-MSCs further reduced the collagen deposition and the degree of HeSCs activation. Considering the well-known immune-modulatory properties of MSCs, we asked whether they might induce changes in numbers of hMø in a TAA chronic model of hepatic fibrosis. Indeed, increased numbers of F4/80+ macrophages were found in liver sections from mice treated with AdGFP-MSCs and AdIGF-I-MSCs when compared to vehicle, at 1 day after treatment (Fig. 1a,b).Fig. 1

View Article: PubMed Central - PubMed

ABSTRACT

Background: Cirrhosis is a major health problem worldwide and new therapies are needed. Hepatic macrophages (hM&oslash;) have a pivotal role in liver fibrosis, being able to act in both its promotion and its resolution. It is well-known that mesenchymal stromal cells (MSCs) can modulate the immune/inflammatory cells. However, the effects of MSCs over hM&oslash; in the context of liver fibrosis remain unclear. We previously described evidence of the antifibrotic effects of in vivo applying MSCs, which were enhanced by forced overexpression of insulin-like growth factor 1 (AdIGF-I-MSCs). The aim of this work was to analyze the effect of MSCs on hM&oslash; behavior in the context of liver fibrosis resolution.

Methods: Fibrosis was induced in BALB/c mice by chronic administration of thioacetamide (8&nbsp;weeks). In vivo gene expression analyses, in vitro experiments using hM&oslash; isolated from the nonparenchymal liver cells fraction, and in vivo experiments with depletion of M&oslash; were performed.

Results: One day after treatment, hM&oslash; from fibrotic livers of MSCs-treated animals showed reduced pro-inflammatory and pro-fibrogenic gene expression profiles. These shifts were more pronounced in AdIGF-I-MSCs condition. This group showed a significant upregulation in the expression of arginase-1 and a higher downregulation of iNOS expression thus suggesting decreased levels of oxidative stress. An upregulation in IGF-I and HGF expression was observed in hM&oslash; from AdIGF-I-MSCs-treated mice suggesting a restorative phenotype in these cells. Factors secreted by hM&oslash;, preconditioned with MSCs supernatant, caused a reduction in the expression levels of hepatic stellate cells pro-fibrogenic and activation markers. Interestingly, hM&oslash; depletion abrogated the therapeutic effect achieved with AdIGF-I-MSCs therapy. Expression profile analyses for cell cycle markers were performed on fibrotic livers after treatment with AdIGF-I-MSCs and showed a significant regulation in genes related to DNA synthesis and repair quality control, cell cycle progression, and DNA damage/cellular stress compatible with early induction of pro-regenerative and hepatoprotective mechanisms. Moreover, depletion of hM&oslash; abrogated such effects on the expression of the most highly regulated genes.

Conclusions: Our results indicate that AdIGF-I-MSCs are able to induce a pro-fibrotic to resolutive phenotype shift on hepatic macrophages, which is a key early event driving liver fibrosis amelioration.

Electronic supplementary material: The online version of this article (doi:10.1186/s13287-016-0424-y) contains supplementary material, which is available to authorized users.

No MeSH data available.