Dataset Information

The Role of CTGF in Liver Fibrosis Induced in 3D Human Liver Spheroids.

ABSTRACT: Connective tissue growth factor (CTGF) is involved in the regulation of extracellular matrix (ECM) production. Elevated levels of CTGF can be found in plasma from patients with liver fibrosis and in experimental animal models of liver fibrosis, but the exact role of CTGF in, e.g., diet-induced human liver fibrosis is not entirely known. To address this question, we utilized a 3D human liver co-culture spheroid model composed of hepatocytes and non-parenchymal cells, in which fibrosis is induced by TGF-β1, CTGF or free fatty acids (FFA). Treatment of the spheroids with TGF-β1 or FFA increased COL1A1 deposition as well as the expression of TGF-β1 and CTGF. Recombinant CTGF, as well as angiotensin II, caused increased expression and/or production of CTGF, TGF-β1, COL1A1, LOX, and IL-6. In addition, silencing of CTGF reduced both TGF-β1- and FFA-induced COL1A1 deposition. Furthermore, we found that IL-6 induced CTGF, COL1A1 and TGF-β1 production, suggesting that IL-6 is a mediator in the pathway of CTGF-induced fibrosis. Taken together, our data indicate a specific role for CTGF and CTGF downstream signaling pathways for the development of liver inflammation and fibrosis in the human 3D liver spheroid model.

SUBMITTER: Trampuz SR

PROVIDER: S-EPMC9857203 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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The Role of CTGF in Liver Fibrosis Induced in 3D Human Liver Spheroids.

Trampuž Sara Redenšek SR van Riet Sander S Nordling Åsa Å Ingelman-Sundberg Magnus M

Cells 20230113 2

Connective tissue growth factor (CTGF) is involved in the regulation of extracellular matrix (ECM) production. Elevated levels of CTGF can be found in plasma from patients with liver fibrosis and in experimental animal models of liver fibrosis, but the exact role of CTGF in, e.g., diet-induced human liver fibrosis is not entirely known. To address this question, we utilized a 3D human liver co-culture spheroid model composed of hepatocytes and non-parenchymal cells, in which fibrosis is induced ...[more]

PMID: 36672237

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Project description:Liver fibrosis is the common outcome of many chronic liver diseases, resulting from altered cell-cell and cell-matrix interactions that promote hepatic stellate cell (HSC) activation and excessive matrix production. This study aimed to investigate functions of cellular communication network factor 2 (CCN2)/Connective tissue growth factor (CTGF), an extracellular signaling modulator of the CYR61/CTGF/Nov (CCN) family, in liver fibrosis. Tamoxifen-inducible conditional knockouts in mice and hepatocyte-specific deletion of this gene in rats were generated using the Cre-lox system. These animals were subjected to peri-central hepatocyte damage caused by carbon tetrachloride. Potential crosstalk of this molecule with a new profibrotic pathway mediated by the Slit2 ligand and Roundabout (Robo) receptors was also examined. We found that Ccn2/Ctgf was highly upregulated in periportal hepatocytes during carbon tetrachloride-induced hepatocyte damage, liver fibrosis and cirrhosis in mice and rats. Overexpression of this molecule was observed in human hepatocellular carcinoma (HCC) that were surrounded with fibrotic cords. Deletion of the Ccn2/Ctgf gene significantly reduced expression of fibrosis-related genes including Slit2, a smooth muscle actin (SMA) and Collagen type I during carbon tetrachloride-induced liver fibrosis in mice and rats. In addition, Ccn2/Ctgf and its truncated mutant carrying the first three domains were able to interact with the 7th -9th epidermal growth factor (EGF) repeats and the C-terminal cysteine knot (CT) motif of Slit2 protein in cultured HSC and fibrotic murine livers. Ectopic expression of Ccn2/Ctgf protein upregulated Slit2, promoted HSC activation, and potentiated fibrotic responses following chronic intoxication by carbon tetrachloride. Moreover, Ccn2/Ctgf and Slit2 synergistically enhanced activation of phosphatidylinositol 3-kinase (PI3K) and AKT in primary HSC, whereas soluble Robo1-Fc chimera protein could inhibit these activities. These observations demonstrate conserved cross-species functions of Ccn2/Ctgf protein in rodent livers. This protein can be induced in hepatocytes and contribute to liver fibrosis. Its novel connection with the Slit2/Robo signaling may have therapeutic implications against fibrosis in chronic liver disease.

Project description:Adenosine deaminase acting on RNA 2 (ADAR2), an RNA editing enzyme is involved in a site-selective modification of adenosine (A) to inosine (I) in double-stranded RNA (dsRNA). Its role in the lungs is unknown. The phenotypic characterization of Adarb1 mice that lacked ADAR2 auto-regulation due to the deletion of editing complementary sequence (ΔECS mice) determined the functional role of ADAR2 in the lungs. ADAR2 protein expression increased in the ΔECS mice. These mice display immune cell infiltration and alveolar disorganization. The lung wet by dry ratio indicates there is no lung edema in ΔECS mice. Bronchoalveolar lavage (BAL) analysis of ΔECS mice reveals a significant increase in neutrophils. Interestingly, ΔECS mice spontaneously develop lung fibrosis as indicated by Sirius red staining of collagen fibers in the lung sections and a significant increase in hydroxyproline level in their lungs. ADAR2 expression increased significantly in a bleomycin mouse model, implicating a role of ADAR2 in lung fibrosis. Furthermore, there is a likely possibility that the genetically modified ΔECS mice does not model the physiological or pathophysiological process of lung fibrosis. Nevertheless, this model is useful in interrogating the role of ADAR2 in the lungs. The Ctgf mRNA and connective tissue growth factor (CTGF) protein significantly increased in ΔECS lungs and occurs in bronchial epithelial cells. There is a significant increase in Human antigen R (ELAVL1; HuR) protein levels in ΔECS lungs and suggests a role in stabilizing Ctgf mRNA. Lung mechanics such as total respiratory resistance, Newtonian resistance and tissue damping were increased, whereas inspiratory capacity was decreased in the ΔECS mice. Taken together, these data indicate that overexpression of ADAR2 causes spontaneous lung fibrosis via HuR-mediated CTGF signaling and implicate a role for ADAR2 auto-regulation in lung homeostasis. The identification of ADAR2 target genes in ΔECS mice would facilitate a mechanistic understanding of the role of ADAR2 in the lungs and provide a therapeutic strategy for lung fibrosis.

Dataset Information

The Role of CTGF in Liver Fibrosis Induced in 3D Human Liver Spheroids.

Publications

The Role of CTGF in Liver Fibrosis Induced in 3D Human Liver Spheroids.

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