Theaflavine inhibits hepatic stellate cell activation by modulating the PKA/ LKB1/AMPK/GSK38 cascade and subsequently enhancing Nrf2 signaling

Activation of hepatic stellate cells (HSCs) constitutes a crucial etiological factor leading to liver fibrosis. Theaflavine (TF) is a characteristic bioactive compound in fermented tea. Here, we found that TF attenuated the activation of LX-2 HSCs induced by transforming growth factor-81 (TGF-81). TF potentiated nuclear factor erythroid 2-related Factor 2 (Nrf2) signaling. Knockdown of Nrf2 abrogated TF-mediated resistance to TGF-81. Liver kinase B1 (LKB1), AMP-activated kinase (AMPK), and glycogen synthase kinase-38 (GSK38) are upstream regulators of Nrf2. TF modulated the LKB1/AMPK/GSK38 axis. Inhibition of AMPK or knockdown of LKB1 crippled TF-mediated potentiation of Nrf2. Protein kinase A (PKA) catalyzes LKB1 phosphorylation. In LX-2 cells, TF increased the LKB1/PKA interaction without affecting their contents. Inhibition of PKA abolished TF-mediated potentiation of LKB1/Nrf2 and abrogated the inhibitory effects of TF on their activation. TF also enhanced direct binding between purified catalytic subunit & alpha; of PKA (PKA-C & alpha;) and LKB1 proteins in vitro. Molecular docking indicated that TF showed binding activity with both LKB1 and PKA-C & alpha; proteins. In mouse primary HSCs, TF elevated LKB1/PKA-C & alpha; binding, boosted LKB1 phosphorylation, potentiated Nrf2 and suppressed their spontaneous activation. PKA inhibition or LKB1 knockdown eliminated TF-mediated induction of Nrf2 and suppression of HSC activation. Furthermore, TF considerably alleviated CCl4-induced mouse liver fibrosis. In mouse livers, TF increased the LKB1/PKA-C & alpha; interaction, upregulated LKB1 phosphorylation and modulated its downstream AMPK/GSK38/Nrf2 cascade. Our findings collectively indicated that TF suppresses HSC activation. Mechanistically, TF elevated the LKB1/PKA interaction in HSCs, which increased LKB1 phosphorylation and subsequently modulated the downstream AMPK/GSK38/Nrf2 axis.

Automated summary

Theaflavine (TF) attenuated the activation of hepatic stellate cells (HSCs) induced by transforming growth factor-β (TGF-β) through potentiating the nuclear factor erythroid 2-related Factor 2 (Nrf2) signaling pathway. TF modulated the LKB1/AMPK/GSK38 axis to enhance the activity of the Nrf2 pathway. Moreover, TF bound to LKB1 and PKA-Cα proteins, leading to increased phosphorylation of LKB1 and modulation of the AMPK/GSK38/Nrf2 cascade.