Hirsutella sinensis mycelium regulates autophagy of alveolar macrophages via TLR4/NF-κB signaling pathway

Abstract

Background : Hirsutella sinensis mycelium (HSM) has potent anti-pulmonary fibrotic activities and has been proposed as an effective treatment for idiopathic pulmonary fibrosis. Macrophages are the main innate immune cells in the lung tissue, playing key roles in pulmonary fibrosis repair and homeostasis. Excessive macrophage autophagy plays a vital role in pulmonary fibrosis. The protective effect of HSM on macrophages of bleomycin (BLM)-induced pulmonary fibrotic mice remain unclear. Methods : In this study, we collected lung tissue and bronchoalveolar lavage fluid (BALF) samples from pulmonary fibrotic mice. Meanwhile, alveolar macrophages were isolated and murine macrophage RAW264.7 cell line was cultured for further study of HSM autophagy. Results: First, we found that HSM decreased the number of autophagosomes, as well as the levels of LC3B and ATG5, and increased the protein level of P62 during the development of pulmonary fibrosis. Meanwhile, HSM reduced alveolar macrophages infiltration into the BALF and inhibited their accumulation in the fibrotic lung tissue. Flow cytometry analysis showed that HSM administration inhibited the autophagy marker LC3B expression in CD11b lo CD11c hi alveolar macrophages in BLM-induced lung fibrosis without affecting CD11b hi CD11c lo interstitial macrophages. Transmission electron microscopy and JC-1 staining for mitochondrial membrane potential of alveolar macrophages also verified that the HSM significantly decreased autophagy in the alveolar macrophages of BLM-treated mice. In vitro , autophagosomes-lysosome fusion inhibitor chloroquine (CQ) was pre-incubated with RAW264.7 cells, and HSM reduced CQ-induced autophagosomes accumulation. TLR4 signaling inhibitor CLI095 reversed the above effects, suggesting HSM could reduce the cumulation of autophagosomes dependent on TLR4. Furthermore, lipopolysaccharide (LPS)-stimulated TLR4-related autophagy was significantly inhibited by HSM treatment. In addition, the protein expressions of TLR4 and phospho-NF-κB p65 were markedly inhibited in cells treated with HSM. Conclusions : These results indicated that HSM could inhibit the autophagy of alveolar macrophages through TLR4/NF-κB signaling pathway to achieve anti-fibrotic effect.

Bushen Wenyang Huayu Decoction Targets TLR4/NF-κB Mediated Autophagy to Treat Endometriosis Effectively

“…Te TLR4/NF-κB pathway is a signal transduction system that has been linked to infammatory immunological mechanisms and the regulation of infammation as well as the regulation of autophagy [38]. Some researchers have discovered that the TLR4/NF-κB signaling pathway can promote autophagy in the pathological phase of idiopathic pulmonary fbrosis, demonstrating that TLR4/NF-κB is required for autophagy [23].…”

“…It is important to fnd novel strategies to cure EMs by modulating autophagy. TLR4/NF-κB constitutes the classical infammatory signaling pathway, can play crucial roles in mediating immune and infammatory responses, and participate in the pathological process of EMs [21,22]; furthermore, autophagy was thought to be regulated via TLR4/NF-κB signaling pathway [23,24]. Recently, TLR4/NF-κB has been implicated in EMs due to the critical regulatory roles of infammation, indicating that targeting TLR4/NF-κB might be a strategy for EMs therapy.…”

Scorpion and centipede alleviates severe asthma through M2 macrophage-derived exosomal miR-30b-5p

“…To identify the effect of SC on macrophages, primary macrophages were isolated from the BALF of mice [ 21 ]. The isolation of macrophages was performed according to the methods in previous studies.…”

The Role of Macrophage Autophagy in Asthma: A Novel Therapeutic Strategy

“…These results indicate that inflammatory stimulus promote macrophage autophagy through the TLR4 signaling pathway. Thus, targeting TLR4 through Hirsutella sinensis mycelium, a natural agent, reduces the cumulation of autophagosomes, and achieves anti-inflammation effects [ 50 ].…”