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Laminin γ2 Mediates Epithelial to mesenchymal Transition in Hepatocytes AML12 Cell Line |
CHEN Hui*(),ZHANG Ji-shun |
Digestive Department, Beijing Chaoyang Hospital, Capital Medical University,Beijing 100043,China |
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Abstract Objective: To investigate the effect of TGF-β1 on laminin γ2 (LAMC2) expression in mouse normal hepatocyte lines AML12, and the role of LAMC2 on the differentiation of AML12 cells. Methods: Lentiviral vector of LAMC2 (Lent-LAMC2) was transfected with AML12 cells, Lenti-NC was used as the control group, the transfection efficiency was detected by Western blot, and the role of LAMC2 on the AML12 differentiation was detected by qPCR analysis. The AML12 cells were stimulated with TGF-β1 (5 ng/mL) from 6 h to 24 h, and the mRNA expression of TGF-β1 and LAMC2 was detected by real-time PCR (qPCR). AML12 cells were stimulated with TGF-β1 or TGF-β1 + TGF-β inhibitor (SB431542) for 24 h, and the mRNA of LAMC2 was detected by qPCR analysis. Finally, small interfering RNA (siRNA) was used to inhibit the expression of LAMC2 in AML12 cells, and the differentiation of AML12 cells was detected by Western blot followed by stimulation of TGF-β1 at dose of 5 ng/mL. Results: Compared with the Lenti-NC vector group, the protein level of LAMC2 was significantly increased in the LAMC2 overexpressed cells, the expressions of α-SMA and Col1a1 mRNA were up-regulated (all P<0.05), and ALB expressions were significantly down-regulated (P< 0.05). With the prolongation of the AML12 cell time of TGF-β1, the mRNA expression of LAMC2 gradually increased, which was positively correlated with the stimulation time of TGF-β1 (P<0.05). The transcription level of LAMC2 in TGF-β1+SB431542 group was significantly lower than that in TGF-β1 group (P<0.05). Importantly, compared with the siNC group, the siLAMC2 group can effectively block the effect of TGF-β1 on the upregulation of expression in AML12 cells α-SMA and Col1a1. Conclusion: Laminin γ2 overexpression induces mesenchymal-like cell changes in AML12 cells. TGF-β1 promotes the expression of LAMC2 in AML12 cells, and LAMC2 specific siRNAs block the fibrogenic effect of TGF-β1 on AML12 cells.
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Received: 28 May 2023
Published: 01 December 2023
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Corresponding Authors:
*Hui CHEN
E-mail: chenhui_0516@126.com
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