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Objective(s): Free cholesterol in the diet can cause liver fibrosis by accumulating in Hepatic stellate cells (HSCs). The rate of mortality of this disease is high worldwide and there is no definite remedy for it, but might be treated by anti-fibrotic therapies. MSCs-derived exosomes are known as the new mechanism of cell-to-cell communication, showing that exosomes can be used as a new treatment. In this study, we investigated the ability of exosomes of WJ-MSCs as a new remedy to reduce cholesterol-induced liver fibrosis in the LX2 cell line. Materials and Methods: MSCs were isolated from Wharton’s jelly of the umbilical cord and the exosomes were extracted. The LX2 cell line was cultured in DMEM medium with 10% FBS, then cells were treated with 75 and 100 μM concentrations of cholesterol for 24 hr. The mRNA expression of TGF-β, αSMA, and collagen1α genes, and the level of Smad3 protein were measured to assess liver fibrosis. Results: Cholesterol increased the expression of TGF-β, αand -SMA, and collagen1α genes by increasing the phosphorylation of the Smad3 protein. Treatment with Exosomes significantly reduced the expression of TGF-β, α-SMA, and collagen1α genes (fibrosis genes). Treatment with exosomes prevented the activation of HSCs by inhibiting the phosphorylation of the Smad3 protein. Conclusion: The exosomes of WJ-MSCs can inhibit the TGFβ/Smad3 signaling pathway preventing further activation of HSCs and progression of liver fibrosis. So, the exosomes of WJ-MSCs s could be introduced as a treatment for liver failure. [ABSTRACT FROM AUTHOR]

Objective(s): Activated cells which are called star-shaped cells, are some of the key factors in the development of liver fibrosis. Activation of NADPH oxidase (NOX) is associated with increased HSCs activity and progression of hepatic fibrosis. In this study, the effects of human exosomes derived from WJ-MSCs on NOX1, NOX2, and NOX4 gene expression in TGF-ß-induced hepatic fibrosis were investigated. Materials and Methods: LX2 cell line was treated with 2 ng/ml TGF-ß for 24 hr, in order to induce liver fibrosis after starvation. In the next step, the cells were treated with several concentrations of the exosomes derived from WJ-MSCs (10, 20, 30, 40, and 50 µg/ml). Finally, Smad3C phosphorylated protein expression level and NOX1, NOX2, and NOX4 gene expression levels were measured. Results: The results demonstrated that the level of NOX1, NOX2, and NOX4 mRNA expressions decreased significantly during 24 hrs at concentrations of 40 and 50 µg/ml of WJ-MSCs exosomes in TGF-ß-induced-HSCs. The p-Smad3C proteins were significantly decreased (fold change: 1.83, P-value<0.05) after exposure to WJ-MSC-derived exosomes. Conclusion: Treatment with exosomes prevents further activation of HSCs by inhibiting the level of Smad3C phosphorylation. The experimental data of our study suggested that in liver fibrosis, the protection of HSCs activation against TGF-ß by inhibiting the NOX pathway via human exosomes of WJ-MSCs is extremely important. It needs further research as a treatment method. [ABSTRACT FROM AUTHOR]

Background: Hepatic stellate cells (HSCs) play a primary role in liver fibrogenesis. NOXs are the main origin of reactive oxygen species (ROS) in the liver. Among them, NOX1, NOX2, and NOX4 are expressed more in HSCs and are involved in the development of liver fibrosis. Quercetin, an abundant citrus flavonoid, is known to have beneficial effects on liver injury and hepatic fibrosis. Objectives: In this study, the effect of quercetin on NOX1, NOX2, and NOX4 expression and Smad3 phosphorylation induced by TGF-β in the human hepatic LX2 cell line was investigated. Methods: The cytotoxic effects of quercetin on the cells were determined by MTT assay. The cells were activated with 2 ng/mL of TGF-β for 24 h and then treated with different concentrations of Quercetin. Them RNA expression rates of NOX1, NOX2, NOX4, and phosphorylated Smad 3C (p-Smad3C) were analyzed using real-time polymerase chain reaction (PCR) and western blot assays. Results: TGF-β increased the mRNA expression of NOX1, NOX2, and NOX4 and the protein level of p-Smad3C in the LX2 cell line. Quercetin significantly decreased the mRNA expression of NOX1,NOX2, and NOX4 in the LX-2 cells. Moreover, quercetin significantly diminished the p-Smad3C level in the LX-2 cell line activated with TGF-β. Conclusions: Quercetin may be effective in improving hepatic fibrosis via the reduction of NOX1, NOX2, and NOX4 expression in activated HSCs. The main mechanism through which quercetin reduces the expression of these target genes may be related to the reduction of the p-Smad3C level. [ABSTRACT FROM AUTHOR]