Transcriptomic and proteomic investigation of the ameliorative effect of total polyphenolic glycoside extract on hepatic fibrosis in Lamiophlomis rotata Kudo via the AGE/RAGE pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: During the regression of liver fibrosis, a decrease in hepatic stellate cells (HSCs) can occur through apoptosis or inactivation of activated HSCs (aHSCs). A new approach for antifibrotic therapy involves transforming hepatic myofibroblasts into a quiescent-like state. Lamiophlomis rotata (Benth.) Kudo (L. rotata), an orally available Tibetan herb, has traditionally been used to treat skin disease, jaundice, and rheumatism. In our previous study, we found that the total polyphenolic glycoside extract of L. rotata (TPLR) promotes apoptosis in aHSCs for the treatment of hepatic fibrosis. However, whether TPLR induces aHSCs to become inactivated HSCs (iHSCs) is unclear, and the underlying mechanism remains largely unknown. PURPOSE: This study aimed to examine the impact of TPLR on the phenotypes of hepatic stellate cells (HSCs) during the regression of liver fibrosis and explore the potential mechanism of action. METHODS: The effect of TPLR on the phenotypes of hepatic stellate cells (HSCs) was assessed using immunofluorescence (IF) staining, reverse transcription-polymerase chain reaction (RT-PCR), and Western blotting. Transcriptomic and proteomic methods were employed to identify the main signaling pathways involved. Based on the omics results, the likely mechanism of TPLR on the phenotypes of aHSCs was confirmed through overexpression and knockdown experiments in TGF-ß1-activated LX-2 cells. Using a CCl4-induced liver fibrosis mouse model, we evaluated the anti-hepatic fibrosis effect of TPLR and explored its potential mechanism based on omics findings. RESULTS: TPLR was found to induce the differentiation of aHSCs into iHSCs by significantly decreasing the protein expression of α-SMA and Desmin. Transcriptomic and proteomic analyses revealed that the AGE/RAGE signaling pathway plays a crucial role in the morphological transformation of HSCs following TPLR treatment. In vitro experiments using RAGE overexpression and knockdown demonstrated that the mechanism by which TPLR affects the phenotype of HSCs is closely associated with the RAGE/RAS/MAPK/NF-κB axis. In a model of liver fibrosis, TPLR obviously inhibited the generation of AGEs and alleviated liver tissue damage and fibrosis by downregulating RAGE and its downstream targets. CONCLUSION: The AGE/RAGE axis plays a pivotal role in the transformation of activated hepatic stellate cells (aHSCs) into inactivated hepatic stellate cells (iHSCs) following TPLR treatment, indicating the potential of TPLR as a therapeutic agent for the management of liver fibrosis.

The total iridoid glycoside extract of Lamiophlomis rotata Kudo induces M2 macrophage polarization to accelerate wound healing by RAS/ p38 MAPK/NF-κB pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Lamiophlomis rotata (Benth.) Kudo (L. rotata), a Tibetan medicinal plant, is used to treat "yellow-water diseases", such as skin disease, jaundice and rheumatism. Our previous study showed that the iridoid glycoside extract of L. rotata (IGLR) is the major constituent of skin wound healing. However, the role of IGLR in the biological process of trauma repair and the probable mechanism of the action remain largely unknown. AIM OF THE STUDY: To investigate the role of IGLR in the biological process of trauma repair and the probable mechanism of the action. MATERIALS AND METHODS: The role of IGLR in wound healing was investigated by overall skin wound in mice with Hematoxylin and Eosin (H&E) and Masson trichrome staining. The anti-inflammatory, angiogenesis-promoting and fibril formation effects of IGLR were visualized in wound skin tissue by immunofluorescence staining, and the proinflammatory factors and growth factors were assayed by real-time polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). Macrophages, dermal fibroblasts, and endothelial cells were cultured to measure the direct/indirect interaction effects of IGLR on the proliferation and migration of cells, and flow cytometry was employed to assess the role of IGLR on macrophage phenotype. Network pharmacology combined with Western blot experiments were conducted to explore possible mechanisms of the actions. RESULTS: IGLR increased the expression of CD206 (M2 markers) through the RAS/p38 MAPK/NF-κB signaling pathway during wound injury in vivo and in vitro. IGLR suppressed the inflammatory cytokines iNOS, IL-1ß and TNF-α in the early stage of wound healing. During the proliferation step of wound repair, IGLR promoted angiogenesis and fibril formation by increasing the expression of VEGF, CD31, TGF-ß and α-SMA in wound tissue, and similar results were verified by RT-PCR and ELISA. In a paracrine mechanism, the extract promoted the proliferation of dermal fibroblasts, and endothelial cells were founded by the conditioned medium (CM). CONCLUSION: IGLR induced M2 macrophage polarization in the early stage of wound healing; in turn, IGLR played a key role in the transition from inflammation to cell proliferation during the biological process of wound healing.