Wnt5a Promotes Cytokines Production and Cell Proliferation in Human Hepatic Stellate Cells Independent of Canonical Wnt Pathway

Background: Wnt5a is involved in the activation of human hepatic stellate cells (HSC) and related with the occurrence of liver fibrosis. As the function and mechanism that Wnt5a mediates HSC activation remains unclear, we sought to investigate them. Methods: Wnt5a levels were determined in the HSC cell line LX-2 after lipopolysaccharide (LPS) and TNF-alpha stimulation. HSC cells showing stable and efficient overexpression or featuring knockdown of Wnt5a were constructed by a lentivirus system. Regulation of cytokine and collagen expressions were confirmed by quantitative PCR or ELISA in stable LX-2 cell lines showing Wnt5a overexpression or knockdown. Proliferation was determined by 5-ethynyl-2'-deoxyuridine labeling. Relevant signaling pathways were identified using specific protein antibodies. Results: LPS and TNF-alpha induced Wnt5a expression in LX-2 cells. Compared with control cells, an increase in IL-1 beta, IL-6, COL1, and COL3 secretion in a stable LX-2 cell line showing Wnt5a overexpression was observed. Knockdown of Wnt5a obviously reduced the production of IL-1 beta, IL-6, COL1, and COL3. Wnt5a overexpression promoted LX-2 proliferation, while Wnt5a knockdown dramatically inhibited cell proliferation. Compared with the effects of Wnt5a knockdown cells, Wnt5a-overexpressing cells triggered the phosphorylation of c-Jun N-terminal kinase (JNK) and beta-catenin, which leads to beta-catenin degradation and inactivates the canonical Wnt/beta-catenin pathway. Conclusions: Wnt5a regulates inflammatory cytokine and collagen production and cell proliferation, which is independent of the canonical Wnt signaling pathway. The results reveal a new signaling mechanism in HSC activation and could provide a new strategy for hepatic fibrosis treatment.