Smad7-overexpressing rat BMSCs inhibit the fibrosis of hepatic stellate cells by regulating the TGF-β1/Smad signaling pathway

Abstract

Mesenchymal stem cells (MSCs) are able to differentiate into hepatocytes, promote the regeneration of hepatic cells and inhibit the progression of hepatic fibrosis. Transforming growth factor (TGF)-β1 is one of the key factors in the development of liver fibrosis, which also promotes extracellular matrix (ECM) formation. Drosophila mothers against decapentaplegic 7 (Smad7) is an essential negative regulator in the TGF-β1/Smad signaling pathway. In the present study, bone mesenchymal stem cells (BMSCs) were isolated from rat bone marrow and transfected with lentiviral vectors carrying the Smad7 gene. Smad7-enhanced green fluorescent protein (EGFP)-BMSCs stably expressing Smad7 were subsequently co-cultured with hepatic stellate cells (HSCs) for 48 h. Smad7 and TGF-β1 levels in the culture medium were detected using ELISA, and the levels of collagen (Col) I, Col III, laminin (LN) and hyaluronic acid (HA) were measured using immunoassays. The early apoptosis rates of HSCs were determined via flow cytometry. Reverse transcription-quantitative polymerase chain reaction and western blotting were performed to evaluate the mRNA and protein expression profiles, respectively. The results indicated that Smad7-EGFP-BMSCs stably expressing Smad7 were successfully constructed. Upon co-culturing with rat Smad7-EGFP-BMSCs, the early apoptotic rate of HSCs was significantly increased (P<0.05). Levels of Smad7 in the culture medium were also significantly increased (P<0.05), whereas the levels of TGF-β1, Col I, Col III, LN and HA were significantly decreased (P<0.05). Furthermore, the mRNA and protein levels of Smad7 and matrix metalloproteinase 1 were significantly increased (P<0.05), whereas those of TGF-β1, α-SMA, Smad2, smad3, TGF-β receptor I, Col I, tissue inhibitors of metalloproteinase-1 and Col III were significantly decreased. The results of the present study suggest that rat BMSCs overexpressing Smad7 may inhibit the fibrosis of HSCs by regulating the TGF-β1/Smad signaling pathway. This provides a novel insight into future treatments for liver fibrosis.

RETRACTED: microRNA-155 Modulates Hepatic Stellate Cell Proliferation, Apoptosis, and Cell Cycle Progression in Rats With Alcoholic Hepatitis via the MAPK Signaling Pathway Through Targeting SOCS1

“…Hepatic stellate cells from rats with AH were transfected with miR-155 mimic, miR-155 inhibitor, siRNA targeting SOCS1, and their corresponding miR-155 negative control (NC) plasmids using Lipofectamine 2000. All these plasmids were purchased from Shanghai Beyotime Biotechnology Co. Ltd. (Shanghai, China) (Wu et al, 2017).…”

“…Afterward, the secondary goat anti-rabbit horseradish enzyme marker (1:5000) was added for incubation for 2 hours, and the expression of the target proteins was detected using an enhanced chemiluminescence reagent electrochemiluminescence (ECL) developing solution. GAPDH served as the internal reference, and the ratio of the gray value of the target band to that of the internal reference band was the relative expression of the protein (Wu et al, 2017).…”

“…Then, 50-mL volumes of propidium iodide (PI) were added until the final concentration reached about 65 mg/mL, and the cells were placed on an ice bath and stained in darkness for 30 minutes. The cells were filtered through a 300-mesh (pore size 40-50 microns) nylon mesh, and the blue fluorescence cell cycle progression was recorded by a flow cytometer at an excitation wavelength of 352 nm (Becton, Dickinson and Company, NJ, USA) (Wu et al, 2017).…”

Aristolochic Acid-Induced Nephrotoxicity: Molecular Mechanisms and Potential Protective Approaches

“…Mothers against decapentaplegic homolog 7 (SMAD7) has been reported to attenuate the development of fibrosis by inhibiting TGF-β signaling [99]. In their mouse model of AAN, Dai and co-workers showed that the loss of SMAD7 was associated with activation of TGF-β/Smad3 and NF-κB signaling pathways, resulting in aggravation of progressive fibrosis and inflammation in mice treated with AA, and that restoration of SMAD7 locally in the kidneys of Smad7 knockout mice inactivated TGF-β/Smad3-mediated renal fibrosis and NF-κB-driven renal inflammation, thereby preventing the progression of chronic AAN [100].…”

Mesenchymal stem cell-based Smad7 gene therapy for experimental liver cirrhosis

“…Overexpression of Smad7 inhibits collagen expression and cell proliferation of HSCs [14]. Our previous study has also demonstrated that rat MSCs overexpressing the Smad7 gene were able to inhibit the fibrogenesis of HSCs [17]. To further validate the therapeutic potential of the Smad7 gene, MSCs overexpressing Smad7 will be injected into rats with liver cirrhosis induced by carbon tetrachloride (CCl 4 ).…”