Role of GDC-0449 in a rat model of liver fibrosis induced by carbon tetrachloride combined with 2-acetylaminofluorene

Yonghong HU; Zhun XIAO; Yadong FU; Yue LIANG; Linzhang ZHANG; Wei LIU; Yongping MU; Chenghai LIU; Ping LIU; Jiamei CHEN

doi:10.3969/j.issn.1001-5256.2022.02.016

Volume 38 Issue 2

Feb. 2022

Turn off MathJax

Article Contents

Article Navigation > Journal of Clinical Hepatology > 2022 > 38(2): 334-341

PDF( 6040 KB)

Role of GDC-0449 in a rat model of liver fibrosis induced by carbon tetrachloride combined with 2-acetylaminofluorene

DOI: 10.3969/j.issn.1001-5256.2022.02.016

Yonghong HU^{1, 2},
Zhun XIAO^{1, 3},
Yadong FU^{1, 3},
Yue LIANG^{1, 2},
Linzhang ZHANG^{1, 3},
Wei LIU^{1, 2},
Yongping MU^{1, 2},
Chenghai LIU^{1, 2},
Ping LIU^{1, 2, 3
,
,},
Jiamei CHEN^{1, 2
,
,}

1.
Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine; Institute of Liver Diseases, Shanghai Institute of Traditional Chinese Medicine, Shanghai 201203, China
2.
Key Laboratory of Liver and Kidney Diseases, Ministry of Education; Shanghai Key Laboratory of Traditional Chinese Clinical Medicine, Shanghai 201203, China
3.
Institute of Interdisciplinary Integrative Medicine Research, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China

Research funding:

National Natural Science Foundation of China (81973613);

National Natural Science Foundation of China (81603465);

Morning Star Project of Shanghai Sci & Tech (19QA1408900);

Wang Baoen Liver Fibrosis Foundation (CFHPC2020010);

Siming Youth Fund Project of Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine (SGKJ-202004)

More Information

Corresponding author: LIU Ping, liuliver@vip.sina.com; CHEN Jiamei, cjm0102@126.com
Received Date: 2021-10-07
Accepted Date: 2021-11-30
Published Date: 2022-02-20

Abstract

Abstract

Objective To investigate the intervention effect of GDC-0449, a hedgehog signaling pathway inhibitor, on rats with liver fibrosis induced by carbon tetrachloride (CCl₄) combined with 2-acetylaminofluorene (2-AAF). Methods A total of 18 female Fisher344 rats were randomly divided into normal group, CCl₄/2-AAF group, and GDC-0449 group, with 6 rats in each group. The rats in the CCl₄/2-AAF group and the GDC-0449 group were given subcutaneously injected 30% CCl₄-olive oil solution at a dose of 2 mL/kg twice a week for 6 weeks to induce liver fibrosis; since week 7, in addition to the injection of CCl₄-olive oil solution, the rats in these two groups were given 2-AAF (100 mg/kg/d) by gavage, and the rats in the GDC-0449 group were given GDC-0449 (25 mg/kg/d) by gavage, while those in the normal group were given an equal volume of olive oil solution by injection and normal saline by gavage. All rats were sacrificed at the end of week 9, and related samples were collected. HE staining and sirius red (SR) staining were used to observe the changes in liver histopathology and collagen deposition, and the semi-quantitative analysis of SR-positive area and Ishak score were used to evaluate fibrosis degree; the alkaline hydrolysis method was used to measure the level of hydroxyproline (Hyp) in liver tissue; immunohistochemistry, Western blot, and qRT-PCR were used to measure the expression of α-smooth muscle actin (α-SMA), type Ⅰ collagen (Col-Ⅰ), type Ⅳ collagen (Col-Ⅳ), cytokeratin 19 (CK19), cytokeratin 7 (CK7), the epithelial cell adhesion molecule Epcam, and the hedgehog signaling pathway in liver tissue; double immunofluorescence staining was used to observe the colocalization of CK19 and the oval cell marker OV6. A one-way analysis of variance was used for comparison of continuous data between multiple groups, and the least significant difference t-test was used for further comparison between two groups. Results Compared with the normal group, the CCl₄/2-AAF group had marked inflammatory cell aggregation and collagen deposition in liver tissue, with the formation of a pseudolobular structure, as well as significant increases in Hyp level and collagen positive area ratio in liver tissue (P < 0.05), Ishak score (P < 0.05), and the expression of α-SMA, Col-Ⅰ, Col-Ⅳ, Epcam, CK19, CK7, the transmembrane transporter Smoothened (Smo), Hedgehog ligand Desert Hedgehog (Dhh), the Indian Hedgehog membrane-binding receptor Patched (Ptch2), and glioma-related oncogenes Gli1, Gli2, and Gli3 (all P < 0.05); double immunofluorescence staining showed that CK19-positive cells also expressed OV6 in the liver tissue of rats in the CCl₄/2-AAF group, with a significant increase compared with the normal group. Compared with the CCl₄/2-AAF group, the GDC-0449 group had significant reductions in inflammatory cell aggregation and collagen deposition in liver tissue, Hyp level and collagen positive area ratio in liver tissue (P < 0.05), Ishak score (P < 0.05), and the expression of α-SMA, Epcam, CK19, CK7, Smo, Ptch2, Gli1, Gli2, and Gli3 (all P < 0.05); double immunofluorescence staining showed a significant reduction in the number of cells with co-expression of OV6 and CK19 in liver tissue. Conclusion The Hedgehog signaling pathway inhibitor GDC-0449 can significantly inhibit the progression of liver fibrosis induced by CCl₄/2-AAF in rats, possibly by inhibiting hepatic stellate cell activation, collagen deposition, activation and proliferation of hepatic progenitor cells, and differentiation of hepatic progenitor cells into biliary epithelial cells.
- Liver Cirrhosis,
- Hedgehog Signaling Pathway,
- Rats, Inbred F344

FullText(HTML)

References(22)

References

[1]	LLOVET JM, ZUCMAN-ROSSI J, PIKARSKY E, et al. Hepatocellular carcinoma[J]. Nat Rev Dis Primers, 2016, 2: 16018. DOI: 10.1038/nrdp.2016.18.
[2]	D'AMICO G, MORABITO A, D'AMICO M, et al. New concepts on the clinical course and stratification of compensated and decompensated cirrhosis[J]. Hepatol Int, 2018, 12(Suppl 1): 34-43. DOI: 10.1007/s12072-017-9808-z.
[3]	PAROLA M, PINZANI M. Liver fibrosis: Pathophysiology, pathogenetic targets and clinical issues[J]. Mol Aspects Med, 2019, 65: 37-55. DOI: 10.1016/j.mam.2018.09.002.
[4]	ASRANI SK, DEVARBHAVI H, EATON J, et al. Burden of liver diseases in the world[J]. J Hepatol, 2019, 70(1): 151-171. DOI: 10.1016/j.jhep.2018.09.014.
[5]	HERNANDEZ-GEA V, FRIEDMAN SL. Pathogenesis of liver fibrosis[J]. Annu Rev Pathol, 2011, 6: 425-456. DOI: 10.1146/annurev-pathol-011110-130246.
[6]	KISSELEVA T, BRENNER D. Molecular and cellular mechanisms of liver fibrosis and its regression[J]. Nat Rev Gastroenterol Hepatol, 2021, 18(3): 151-166. DOI: 10.1038/s41575-020-00372-7.
[7]	MARCELLIN P, GANE E, BUTI M, et al. Regression of cirrhosis during treatment with tenofovir disoproxil fumarate for chronic hepatitis B: A 5-year open-label follow-up study[J]. Lancet, 2013, 381(9865): 468-475. DOI: 10.1016/S0140-6736(12)61425-1.
[8]	D'AMBROSIO R, AGHEMO A, RUMI MG, et al. A morphometric and immunohistochemical study to assess the benefit of a sustained virological response in hepatitis C virus patients with cirrhosis[J]. Hepatology, 2012, 56(2): 532-543. DOI: 10.1002/hep.25606.
[9]	VILAR-GOMEZ E, MARTINEZ-PEREZ Y, CALZADILLA-BERTOT L, et al. Weight loss through lifestyle modification significantly reduces features of nonalcoholic steatohepatitis[J]. Gastroenterology, 2015, 149(2): 367-378. DOI: 10.1053/j.gastro.2015.04.005.
[10]	MACHADO MV, DIEHL AM. Hedgehog signalling in liver pathophysiology[J]. J Hepatol, 2018, 68(3): 550-562. DOI: 10.1016/j.jhep.2017.10.017.
[11]	SHACKEL NA, MCGUINNESS PH, ABBOTT CA, et al. Identification of novel molecules and pathogenic pathways in primary biliary cirrhosis: cDNA array analysis of intrahepatic differential gene expression[J]. Gut, 2001, 49(4): 565-576. DOI: 10.1136/gut.49.4.565.
[12]	ISHAK K, BAPTISTA A, BIANCHI L, et al. Histological grading and staging of chronic hepatitis[J]. J Hepatol, 1995, 22(6): 696-699. DOI: 10.1016/0168-8278(95)80226-6.
[13]	LO RC, KIM H. Histopathological evaluation of liver fibrosis and cirrhosis regression[J]. Clin Mol Hepatol, 2017, 23(4): 302-307. DOI: 10.3350/cmh.2017.0078.
[14]	OMENETTI A, CHOI S, MICHELOTTI G, et al. Hedgehog signaling in the liver[J]. J Hepatol, 2011, 54(2): 366-373. DOI: 10.1016/j.jhep.2010.10.003.
[15]	FARGNOLI MC, PELLEGRINI C, PICCERILLO A, et al. Clinical determinants of complete response to vismodegib in locally advanced basal cell carcinoma: A multicentre experience[J]. J Eur Acad Dermatol Venereol, 2021, 35(12): e923-e926. DOI: 10.1111/jdv.17588.
[16]	PRATAP A, SINGH S, MUNDRA V, et al. Attenuation of early liver fibrosis by pharmacological inhibition of smoothened receptor signaling[J]. J Drug Target, 2012, 20(9): 770-782. DOI: 10.3109/1061186X.2012.719900.
[17]	SICKLICK JK, LI YX, CHOI SS, et al. Role for hedgehog signaling in hepatic stellate cell activation and viability[J]. Lab Invest, 2005, 85(11): 1368-1380. DOI: 10.1038/labinvest.3700349.
[18]	KUMAR V, DONG Y, KUMAR V, et al. The use of micelles to deliver potential hedgehog pathway inhibitor for the treatment of liver fibrosis[J]. Theranostics, 2019, 9(25): 7537-7555. DOI: 10.7150/thno.38913.
[19]	SICKLICK JK, LI YX, MELHEM A, et al. Hedgehog signaling maintains resident hepatic progenitors throughout life[J]. Am J Physiol Gastrointest Liver Physiol, 2006, 290(5): g859-g870. DOI: 10.1152/ajpgi.00456.2005.
[20]	OMENETTI A, SYN WK, JUNG Y, et al. Repair-related activation of hedgehog signaling promotes cholangiocyte chemokine production[J]. Hepatology, 2009, 50(2): 518-527. DOI: 10.1002/hep.23019.
[21]	OMENETTI A, POPOV Y, JUNG Y, et al. The hedgehog pathway regulates remodelling responses to biliary obstruction in rats[J]. Gut, 2008, 57(9): 1275-1282. DOI: 10.1136/gut.2008.148619.
[22]	OMENETTI A, YANG L, LI YX, et al. Hedgehog-mediated mesenchymal-epithelial interactions modulate hepatic response to bile duct ligation[J]. Lab Invest, 2007, 87(5): 499-514. DOI: 10.1038/labinvest.3700537.

Relative Articles

Supplements(0)

Cited By

Proportional views

Proportional views

通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

Figures(5) / Tables(4)

Get Citation

PDF

XML

Article Metrics

Article views (528) PDF downloads(38)

Role of GDC-0449 in a rat model of liver fibrosis induced by carbon tetrachloride combined with 2-acetylaminofluorene

DOI: 10.3969/j.issn.1001-5256.2022.02.016

Abstract

References

Proportional views

Catalog

通讯作者: 陈斌, bchen63@163.com

Article Metrics

Proportional views

Related

Role of GDC-0449 in a rat model of liver fibrosis induced by carbon tetrachloride combined with 2-acetylaminofluorene

DOI: 10.3969/j.issn.1001-5256.2022.02.016

Abstract

References

Proportional views

Catalog

通讯作者: 陈斌, bchen63@163.com

Article Metrics

Proportional views

Related

About Journal

Submission Guideline

Journal Online

Hepatobiliary College

Guidelines

Export File

Citation

Format

Content