Background: Mesenchymal stem cells (MSCs) are the most promising tools for cell treatment and human tissue regeneration, e.g., in liver fibrosis. Mesenchymal stem cells repair tissue damage through paracrine mediators such as exosomes. Types and concentrations of inflammatory mediators, including transforming growth factor-beta (TGFβ1), in MSCs microenvironment can affect MSCs’ function and therapeutic potency. Objectives: This experimental study aimed to explore the effects of Wharton jelly MSCs (WJ-MSCs) exosomes on fibrotic gene expression and Smad2/3 phosphorylation (phospho-Smad2/3 (p-Smad2/3)). Moreover, we further investigated whether WJ-MSCs pretreatment with different concentrations of TGFβ1 changes the anti-fibrotic properties of their exosomes. Methods: After isolation from the umbilical cord, WJ-MSCs were characterized by observing differentiation and measuring surface biomarkers using flowcytometry. The WJ-MSC-derived exosomes were extracted and identified using transmission electron microscopy (TEM), dynamic light scattering (DLS), and western blotting. Real-time PCR and western blot for extracellular matrix (ECM) and p-Smad2/3 expression detection were used to investigate the effect of exosomes from untreated and TGFβ1-pretreated WJ-MSCs on activated hepatic stellate cells (HSCs). Results: Phospho-Smad2/3, α-smooth muscle actin (α-SMA), and collagen1α1 levels were enhanced following treatment with TGFβ1, whereas E-cadherin was decreased. However, the outcomes were reversed after treatment with WJ-MSC-derived exosomes. Exosomes from TGFβ1-pretreated WJ-MSCs induced a significant decrease in p-Smad2/3 levels in activated HSCs, accompanied by the upregulation of E-cadherin gene expression and downregulation of α-SMA and collagen1α1 when compared to untreated WJ-MSC-derived exosomes. The p-Smad2/3 proteins were significantly decreased (fold change: 0.23, P-value < 0.0001) after exposure to low-dose TGFβ1-pretreated WJ-MSC-derived exosomes (0.1 ng/mL), showing the best effect on activated HSCs. Conclusions: Exosomes derived from untreated WJ-MSCs could regress TGFβ-Smad2/3 signaling and the expression of fibrotic markers in activated LX-2 cells. However, these effects were significantly profound with applying exosomes derived from 0.1 ng/mL TGFβ-pretreated WJ-MSCs. We also observed the dose-response effects of TGFβ on WJ-MSCs-derived exosomes. Therefore, exosomes derived from TGFβ-pretreated WJ-MSCs may be critical in improving fibrosis and benefit liver fibrosis patients.

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. The mRNA 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.

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Background: Advanced glycation end-products (AGEs) cause proinflammatory responses and macromolecular damages. Advanced oxidation protein products (AOPPs) are protein biomarkers for oxidative stress. Levels of AGEs and AOPPs increase with the progression of chronic renal dysfunction. Objectives: In this study, we aimed to measure these species in patients with renal transplantation and to analyze their correlation with the measured glomerular filtration rate (GFR) and renal function parameters. Patients and Methods: Eighty renal transplant patients and normal subjects were recruited. GFR was measured by the two-sample plasma method with technetium-99m-labeled diethylenetriaminepentaacetic acid (TC99m-DTPA) clearance. Biochemical measurements included creatinine, cystatin C, urea, total protein, and pentosidine. Serum AGEs were determined using a fluorometric assay and AOPPs were estimated spectrophotometrically. Results: The measured GFR found to be significantly decreased in renal transplant patients compared to the control subjects (P< 0.001). Levels of AGEs, AOPPs, serum creatinine, and cystatin C were increased in renal transplant patients with lower values of measured GFR (mGFR). A significant association between the levels of AGEs species (serum fluorescence and pentosidine) and mGFR when adjusted for creatinine and other risk factors in multiple linear regression model analysis was found (P=0.05 and P=0.001, respectively). Conclusions: This study demonstrated increased levels of pentosidine and AGEs in transplant recipients were associated with decreased mGFR. Their accumulation can be predictive for the progression of chronic allograft loss of function.

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