Identifying the Potential of miRNAs in Houttuynia cordata-Derived Exosome-Like Nanoparticles Against Respiratory RNA Viruses.

Introduction: Pathogenic respiratory RNA viruses, including influenza A virus (IAV), respiratory syncytial virus (RSV), and SARS-CoV-2, are major causes of causes of acute respiratory infection globally. Plant-derived exosome-like nanoparticles containing miRNAs have shown substantial cross-kingdom regulatory effects on both viral and human transcripts. Houttuynia cordata (H. cordata), a traditional Chinese medicine frequently used to treat respiratory diseases. However, the role of H. cordata-derived exosome-like nanoparticles (HELNs) and the miRNA they encapsulated are unclear. Methods: HELNs were isolated from fresh underground roots (uHELNs) and above ground stems and leaves (aHELNs) using differential centrifugation. The HELNs were identified using transmission electron microscopy, nanoparticle tracking analysis, and zeta potential. Small RNA sequencing and RT-PCR were employed to determine the miRNA expression in uHELNs and aHELNs. All genomes were sourced from the NCBI database. Target prediction of viral genomes was performed using RNAhybrid, while human target prediction was conducted using both RNAhybrid and Miranda. Functional enrichment analysis was applied to the predicted human targets to explore the hub targets and their roles in antiviral effects. The accessibility of miRNA target sites was determined through the MFOLD web server, and customized dual-luciferase reporter assays were administered to validate the computational findings. Results: A total of 12 highly enriched miRNAs were identified in both uHELNs and aHELNs. Upon prediction and verification, miR858a and miR858b were shown to target the NP gene in H1N1, while miR166a-3p targeted the ORF1ab in SARS-CoV-2. However, no valid miRNA targets were found for RSV. Regarding human transcripts, miR168a-3p, miR168b-3p, and miR8175 were found to inhibit MAPK3 expression, and novel_mir2 could suppress both AKT1 and MAPK3 expression. Discussion: This study sheds light on the collaborative antiviral mechanism of miRNAs in HELNs across two species and explores the potential antiviral scopes of both H. cordata miRNAs and HELNs.

Flavonoid-Rich Extract of Oldenlandia diffusa (Willd.) Roxb. Inhibits Gastric Cancer by Activation of Caspase-Dependent Mitochondrial Apoptosis.

OBJECTIVE: To evaluate the apoptosis and cycle arrest effects of Oldenlandia diffusa flavonoids on human gastric cancer cells, determine the action mechanisms in association with the mitochondrial dependent signal transduction pathway that controls production of reactive oxygen species (ROS), and evaluate the pharmacodynamics of a mouse xenotransplantation model to provide a reference for the use of flavonoids in prevention and treatment of gastric cancer. METHODS: Flavonoids were extracted by an enzymatic-ultrasonic assisted method and purified with D-101 resin. Bioactive components were characterized by high-performance liquid chromatography. Cell lines MKN-45, AGS, and GES-1 were treated with different concentrations of flavonoids (64, 96, 128, 160 µg/mL). The effect of flavonoids on cell viability was evaluated by MTT method, and cell nuclear morphology was observed by Hoechst staining. The apoptosis rate and cell cycle phases were measured by flow cytometry, the production of ROS was detected by laser confocal microscope, the mitochondrial membrane potential (MMP) were observed by fluorescence microscope, and the expression of apoptotic proteins related to activation of mitochondrial pathway were measured by immunoblotting. MKN-45 cells were transplanted into BALB/c nude mice to establish a xenograft tumor model. Hematoxylin and eosin staining was used to reveal the subcutaneous tumor tissue. The tumor volume and tumor weight were measured, the expression levels of proliferation markers proliferating cell nuclear antigen (PCNA) and Ki-67 were detected by immunohistochemistry, and the expression levels of CA72-4 were measured by enzyme linked immunosorbent assay. RESULTS: Oldenlandia diffusa flavonoids inhibited proliferation of MKN-45 and AGS human gastric cancer cells, arrested the cell cycle in G1/S phase, induced accumulation of ROS in the process of apoptosis, and altered MMP. In addition, flavonoids increased Apaf-1, Cleaved-Caspase-3, and Bax, and decreased Cyclin A, Cdk2, Bcl-2, Pro-Caspase-9, and Mitochondrial Cytochrome C (P<0.05). The MKN-45 cell mouse xenotransplantation model further clarified the growth inhibitory effect of flavonoids towards tumors. The expression levels of PCNA and Ki-67 decreased in each flavonoid dose group, the expression level of CA72-4 decreased (P<0.05). CONCLUSION: Flavonoids derived from Oldenlandia diffusa can inhibit proliferation and induce apoptosis of human gastric cancer cells by activating the mitochondrial controlled signal transduction pathway.