Bioactive compounds from Huashi Baidu decoction possess both antiviral and anti-inflammatory effects against COVID-19.

The coronavirus disease 2019 (COVID-19) pandemic is an ongoing global health concern, and effective antiviral reagents are urgently needed. Traditional Chinese medicine theory-driven natural drug research and development (TCMT-NDRD) is a feasible method to address this issue as the traditional Chinese medicine formulae have been shown effective in the treatment of COVID-19. Huashi Baidu decoction (Q-14) is a clinically approved formula for COVID-19 therapy with antiviral and anti-inflammatory effects. Here, an integrative pharmacological strategy was applied to identify the antiviral and anti-inflammatory bioactive compounds from Q-14. Overall, a total of 343 chemical compounds were initially characterized, and 60 prototype compounds in Q-14 were subsequently traced in plasma using ultrahigh-performance liquid chromatography with quadrupole time-of-flight mass spectrometry. Among the 60 compounds, six compounds (magnolol, glycyrrhisoflavone, licoisoflavone A, emodin, echinatin, and quercetin) were identified showing a dose-dependent inhibition effect on the SARS-CoV-2 infection, including two inhibitors (echinatin and quercetin) of the main protease (Mpro), as well as two inhibitors (glycyrrhisoflavone and licoisoflavone A) of the RNA-dependent RNA polymerase (RdRp). Meanwhile, three anti-inflammatory components, including licochalcone B, echinatin, and glycyrrhisoflavone, were identified in a SARS-CoV-2-infected inflammatory cell model. In addition, glycyrrhisoflavone and licoisoflavone A also displayed strong inhibitory activities against cAMP-specific 3',5'-cyclic phosphodiesterase 4 (PDE4). Crystal structures of PDE4 in complex with glycyrrhisoflavone or licoisoflavone A were determined at resolutions of 1.54 Å and 1.65 Å, respectively, and both compounds bind in the active site of PDE4 with similar interactions. These findings will greatly stimulate the study of TCMT-NDRD against COVID-19.

SARS-CoV-2 Z-RNA activates the ZBP1-RIPK3 pathway to promote virus-induced inflammatory responses.

SARS-CoV-2 infection can trigger strong inflammatory responses and cause severe lung damage in COVID-19 patients with critical illness. However, the molecular mechanisms by which the infection induces excessive inflammatory responses are not fully understood. Here, we report that SARS-CoV-2 infection results in the formation of viral Z-RNA in the cytoplasm of infected cells and thereby activates the ZBP1-RIPK3 pathway. Pharmacological inhibition of RIPK3 by GSK872 or genetic deletion of MLKL reduced SARS-CoV-2-induced IL-1ß release. ZBP1 or RIPK3 deficiency leads to reduced production of both inflammatory cytokines and chemokines during SARS-CoV-2 infection both in vitro and in vivo. Furthermore, deletion of ZBP1 or RIPK3 alleviated SARS-CoV-2 infection-induced immune cell infiltration and lung damage in infected mouse models. These results suggest that the ZBP1-RIPK3 pathway plays a critical role in SARS-CoV-2-induced inflammatory responses and lung damage. Our study provides novel insights into how SARS-CoV-2 infection triggers inflammatory responses and lung pathology, and implicates the therapeutic potential of targeting ZBP1-RIPK3 axis in treating COVID-19.

Pre-B-cell colony enhancing factor regulates the alveolar epithelial sodium-water transport system through the ERK and AKT pathways.

The sodium-water transport system is crucial for alveolar fluid clearance. The pulmonary edema caused by extracorporeal circulation is mainly due to increased alveolar capillary permeability and reduced fluid clearance. We previously demonstrated that pre-B-cell colony enhancing factor (PBEF) increases alveolar capillary permeability and inhibits the sodium-water transport system. However, the specific mechanism by which PBEF inhibits the sodium-water transport system is unclear. In this study, we used HPAEpiC (alveolar type II epithelial cells) to construct an anoxia-reoxygenation model and simulate the extracorporeal circulation microenvironment. The impact of PBEF on the expression of genes and proteins implicated in sodium transport and its effect on the activation status of the ERK, P38, and AKT signaling pathways were explored in HPAEpiC by real-time fluorescent PCR and western blotting. Specific inhibitors were employed to verify the role of the three signaling pathways in the regulation of the sodium-water transport system. PBEF was substantially non-toxic to alveolar epithelial cells, inhibited the expression of ENaC, NKA, and AQP1, and affected the ERK, P38, and AKT signaling pathways. ERK pathway inhibitors attenuated PBEF-induced downregulation of EnaC, NKA, and AQP1, and increased NKA activity. P38 pathway inhibitors only attenuated PBEF-induced suppression of NKA expression. AKT pathway inhibitors potentiated the inhibitory effects of PBEF, reducing EnaC, AQP1, and NKA expression, as well as NKA activity. In conclusion, PBEF inhibited the sodium-water transport system by activation of ERK and suppression of AKT signaling.

pre-B cell colony enhancing factor negatively regulates Na+ and fluid transport in lung epithelial cells.

This study was undertaken to investigate the effect of pre-B cell colony enhancing factor (PBEF) on Na+ and fluid transport in lung epithelial cells. METHODS: Type 1 and 2 cells were isolated from lung epithelium. After hypoxia reoxygenation treatment, the primary cell cultures were transfected with a plasmid over-expressing PBEF. Sodium-potassium ATPase (NKA), epithelial sodium channel (ENaC), type I cell marker rT140, surfactant protein (SP) and PBEF protein were analyzed at mRNA and protein levels using PCR and Western blot analysis. Immunofluorescence assays showed type 1 and 2 cells were successfully isolated. After the transfection with PBEF over-expression vector, PBEF and RTI40 levels were increased, while ENaC and SP as well as NKA, were decreased in both cells. It is clear that PBEF negatively regulates the expression of ENaC and NKA in the Na+ and fluid transport in lung epithelial cells.