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PRC1 Promotes GLI1-dependent Osteopontin Expression in Association With the Wnt/β-catenin Signaling Pathway and Aggravates Liver Fibrosis

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PRC1 Promotes GLI1-dependent Osteopontin Expression in Association With the Wnt/β-catenin Signaling Pathway and Aggravates Liver Fibrosis

Shenzong Rao et al. Cell Biosci.

Abstract

Background: PRC1 (Protein regulator of cytokinesis 1) regulates microtubules organization and functions as a novel regulator in Wnt/β-catenin signaling pathway. Wnt/β-catenin is involved in development of liver fibrosis (LF). We aim to investigate effect and mechanism of PRC1 on liver fibrosis.

Methods: Carbon tetrachloride (CCl4)-induced mice LF model was established and in vitro cell model for LF was induced by mice primary hepatic stellate cell (HSC) under glucose treatment. The expression of PRC1 in mice and cell LF models was examined by qRT-PCR (quantitative real-time polymerase chain reaction), western blot and immunohistochemistry. MTT assay was used to detect cell viability, and western blot to determine the underlying mechanism. The effect of PRC1 on liver pathology was examined via measurement of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and hydroxyproline, as well as histopathological analysis.

Results: PRC1 was up-regulated in CCl4-induced mice LF model and activated HSC. Knockdown of PRC1 inhibited cell viability and promoted cell apoptosis of activated HSC. PRC1 expression was regulated by Wnt3a signaling, and PRC1 could regulate downstream β-catenin activation. Moreover, PRC1 could activate glioma-associated oncogene homolog 1 (GLI1)-dependent osteopontin expression to participate in LF. Adenovirus-mediated knockdown of PRC1 in liver attenuated LF and reduced collagen deposition.

Conclusions: PRC1 aggravated LF through regulating Wnt/β-catenin mediated GLI1-dependent osteopontin expression, providing a new potential therapeutic target for LF treatment.

Keywords: GLI; HSC; Liver fibrosis; Osteopontin; PRC1; Wnt/β-catenin.

Conflict of interest statement

Competing interestsThe authors declare that they have no competing interests, and all authors should confirm its accuracy.

Figures

Fig. 1
Fig. 1
PRC1 was up-regulated in CCl4-induced mice LF. a Plasma levels of AST and ATL in CCl4-induced mice (LF) and the mice without CCl4 treatment (Sham). **Represents LF vs. Sham, p < 0.01. b Masson staining of liver tissues from LF and Sham mice. c Quantitative analysis of liver Hyp content from LF and Sham mice. **Represents LF vs. Sham, p < 0.01. d Proteins expression of α-SMA and type I collagen of liver tissues from LF and Sham mice. **Represents LF vs. Sham, p < 0.01. e mRNA expression of PRC1 in liver tissues from LF and Sham mice. **Represents LF vs. Sham, p < 0.01. f Immunohistochemistry analysis of PRC1 in in liver tissues from LF and Sham mice
Fig. 2
Fig. 2
PRC1 was up-regulated in activated HSCs. a mRNA expression of PRC1 in quiescent and activated HSCs. **Represents quiescent vs. activated, p < 0.01. b Proteins expression of PRC1, α-SMA and type I collagen in quiescent and activated HSCs. **Represents quiescent vs. activated, p < 0.01
Fig. 3
Fig. 3
Knockdown of PRC1 suppressed cell proliferation and promoted cell apoptosis of activated HSCs. a Transfection efficiency of shPRC1 #1/#2 or pcDNA3.1-PRC1 in activated HSCs. **Represents shPRC1 #1 or #2 vs. shNC; pcDNA3.1-PRC1 (PRC1) vs. pcDNA3.1 (vector) p < 0.01. b The effects of shPRC1 or pcDNA3.1-PRC1 on cell viability of activated HSCs. **Represents shPRC1 vs. shNC; PRC1 vs. vector, p < 0.01. c The effects of shPRC1 or pcDNA3.1-PRC1 on proteins expression of Bcl-2, Bax, Cleaved Caspase-3, caspase-3, PARP, Cleaved PARP, cyto cytochrome C and mito cytochrome C in activated HSCs. **Represents shPRC1 vs. shNC, p < 0.01. ##Represents PRC1 vs. vector, p < 0.01
Fig. 4
Fig. 4
Knockdown of PRC1 attenuated LF progression. a The effect of tail vein injection of Ad-shPRC1 on plasma levels of AST and ATL in mice liver tissues. **Represents LF + Ad-shNC vs. Sham, p < 0.01. ##LF + Ad-shPRC1 vs. LF + Ad-shNC, p < 0.01. b Masson staining of liver tissues from Sham mice and LF mice injected with Ad-shNC or Ad-shPRC1. c Quantitative analysis of liver Hyp content from Sham mice and LF mice injected with Ad-shNC or Ad-shPRC1. **Represents LF + Ad-shNC vs. Sham, p < 0.01. ##LF + Ad-shPRC1 vs. LF + Ad-shNC, p < 0.01. d mRNA expression of PRC1 in liver tissues from Sham mice and LF mice injected with Ad-shNC or Ad-shPRC1. **Represents LF + Ad-shNC vs. Sham, p < 0.01. ##LF + Ad-shPRC1 vs. LF + Ad-shNC, p < 0.01. e Immunohistochemistry analysis of PRC1 in in liver tissues from Sham mice and LF mice injected with Ad-shNC or Ad-shPRC1. f Proteins expression of α-SMA, type I collagen, PRC1, GLI1 and osteopontin of liver tissues from Sham mice and LF mice injected with Ad-shNC or Ad-shPRC1. **Represents LF + Ad-shNC vs. Sham, p < 0.01. ##LF + Ad-shPRC1 vs. LF + Ad-shNC, p < 0.01
Fig. 5
Fig. 5
PRC1 regulated GLI1 expression in association with the Wnt/β-catenin signaling pathway. a The effect of PRC1 and Wnt3a on mRNAs expression of survivin, MYC and JUN in activated HSCs. **Represents Wnt3a (−) + shPRC1 vs. Wnt3a (−) + shNC, p < 0.01. ##Represents Wnt3a (+) + shPRC1 vs. Wnt3a (+) + shNC, p < 0.01. b The effect of PRC1 and Wnt3a on proteins expression of PRC1, β-catenin, active β-catenin and GLI1 in activated HSCs. **Represents Wnt3a (−) + shPRC1 vs. Wnt3a (−) + shNC, p < 0.01. ##Represents Wnt3a (+) + shPRC1 vs. Wnt3a (+) + shNC, p < 0.01
Fig. 6
Fig. 6
PRC1 promoted GLI1-dependent osteopontin expression. The effect of PRC1 and GLI1 on proteins expression of PRC1, GLI1 and osteopontin in activated HSCs. **Represents pcDNA3.1-GLI1 + shNC vs. pcDNA3.1-NC + shNC, p < 0.01. ##pcDNA3.1-GLI1 + shPRC1 vs. pcDNA3.1-NC + shPRC1, p < 0.01

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