Int J Stem Cells.  2020 Nov;13(3):404-413. 10.15283/ijsc20052.

Upregulation of C-Reactive Protein by Placenta-Derived Mesenchymal Stem Cells Promotes Angiogenesis in A Rat Model with Cirrhotic Liver

• Jun JH
• Jung J
• Kim JY
• Hwang SG
• Bae SH
• Kim GJ

• KMID: 2508914
• DOI: http://doi.org/10.15283/ijsc20052

Abstract

Background and Objectives
Liver cirrhosis is accompanied by abnormal vascular shunts. The Wnt pathway is essential for endothelial cell survival and proliferation. C-reactive protein (CRP), which is produced by hepatocyte, activates angiogenesis in cardiovascular diseases.
Methods and Results
The expression of CRP in CCl 4 -injured rat livers was detected using qRT-PCR and Western blotting after transplantation of placenta-derived mesenchymal stem cells (PD-MSCs) into rats. To determine whether CRP functions in hepatic regeneration by promoting angiogenesis through the Wnt pathway, we detected VEGF and β-catenin in liver tissues and BrdU and β-catenin in hepatocytes by immunofluorescence. The expression levels of CRP, Wnt pathway-related and angiogenic factors were increased in CCl 4 -injured and PD-MSCs transplanted rat livers. In vitro, the expression levels of Wnt signaling and angiogenic factors were decreased in siRNA-CRP-transfected rat hepatocytes.
Conclusions
CRP upregulation by PD-MSCs participates in vascular remodeling to promote liver regeneration via the Wnt signaling pathway during hepatic failure.

Keyword

Liver cirrhosis; C-reactive protein; Placenta-derived mesenchymal stem cells; Wnt pathway; Vascular remodeling

Figure

• 

  Fig. 1 PD-MSC transplantation attenuates inflammation in CCl4-injured rats. Expression of NF-κB was detected by immunohistochemistry (A). ×400, Scale bars; 20 μm. NF-κB-positive cells were quantified based on the total number of cells (B). Expression of IL-10 (C) and IL-6 (D) was analyzed in rat serum by ELISA. Data are represented as the triplicated mean±SD. p<0.05, *; NTX vs.; NTX and TTX refer to the non-transplanted group and the PD-MSC-transplanted group, respectively.

• 

  Fig. 2 PD-MSC transplantation improves the expression of CRP. Expre-ssion of CRP was analyzed by ELISA (A) and Western blotting from total proteins (C) and exosomes (B) and (D). Data are represented as the triplicated or duplicated mean±SD. SD; p<0.05, *; NTX vs.; NTX and TTX refer to the non-transplanted group and the PD-MSC-transplanted group, respectively.

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  Fig. 3 PD-MSC transplantation induces Wnt signaling and angiogene-sis in CCl4-injured rats. Angiogenic (A) and Wnt signaling (B) factors were detected by Western blotting. Localization and expression of VEGF and β-catenin visualized using immunofluorescence (C). ×630, Scale bars; 20 μm. Positive correlations were found between CRP and β-catenin (D) also, between β-catenin and VEGF (E). NTX and TTX refer to the non-transplanted group and the PD-MSC-transplanted group, respec-tively.

• 

  Fig. 4 CRP regulates angiogenesis and Wnt signaling in rat hepatocytes (WB-F344s). Protein levels of CRP (A), VEGF (B), β-catenin (C), ALB (D), HNF1α (E), and CyclinD1 (F) were evaluated in siRNA-CRP-transfected rat hepatocytes by western blot. Data are represented as the triplicated mean±SD of. *, p<0.05. Mock and si-CRP refer to non-transfected and siRNA-CRP-transfected rat hepatocytes, respectively.

• 

  Fig. 5 PD-MSCs promote the expression of CRP and proliferation of hepatocytes through Wnt signaling. Expression of CRP was analyzed by qRT-PCR (A) and ELISA (B) after CCl4 treatment and co-culture with PD-MSCs. Immunofluorescence staining shows the localization and expression of BrdU and β-catenin in rat hepatocytes treated with CCl4 and co-cultured with PD-MSCs (C). ×400, Scale bars; 20 μm. BrdU- or β-ca-tenin-(D) positive rat hepatocytes were quantified after immunofluore-scence staining. Protein levels of HNF1α, and CyclinD1 were detected by western blotting in rat hepatocytes treated with CCl4 and co-cultured with PD-MSCs (E). Intensity of HNF1α (F) and Cyclin D1 (G) protein bands was calculated. Tripli-cated data are represented as the mean±SD. *, p<0.05.

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  Fig. 6 PD-MSCs induce the expression of CRP and VEGF at the mRNA level. Schematic showing the experimental protocol (A). mRNA level of CRP (B), VEGF (D) was quantified in rat hepatocytes treated with CCl4 and co-cultured with HUVECs and PD-MSCs. mRNA level of CRP (C), VEGF (E) was assayed in HUVECs after CCl4 treatment and co-culture with rat hepatocytes and PD-MSCs. Duplicated data are represented as the mean±SD. *, p<0.05.

Reference

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