The efficacy of Xiangmei Pills on rats with ulcerative colitis(UC) was investigated by characterizing the spectrum of the active chemical components of Xiangmei Pills. Rapid identification and classification of the main chemical components were performed,and the therapeutic effects of Xiangmei Pills on the proteins and intestinal flora of UC rats were analyzed to explore the mechanism of its action in treating UC. Fifty SD rats were acclimatized to feeding for 3 d and randomly divided into blank group, model group,mesalazine group(0. 4 g·kg~(-1)), low-dose group of Xiangmei Pills(1. 89 g·kg~(-1)), and high-dose group of Xiangmei Pills(5. 67 g·kg~(-1)), with 10 rats in each group. 5% dextrose sodium sulfate(DSS) was given by gavage to induce the male SD rat model with UC,and the corresponding medicinal solution was given by gavage after 10 days, respectively. The therapeutic effect of Xiangmei Pills on rats with UC was evaluated according to body mass, disease activity index(DAI), and hematoxylin-eosin(HE) staining, and the histopathological changes in the colon were observed. Ultra-high performance liquid chromatography-quadrupole/electrostatic field orbitrap high-resolution mass spectrometry(UHPLC-Q-Orbitrap HRMS) technique was used to rapidly and accurately identify the main chemical constituents of Xiangmei Pills. Immunohistochemistry was used to detect the expression of aryl hydrocarbon receptor(AhR),interferon-γ(IFN-γ), mucin-2(MUC-2), and cytochrome P450 1A1(CYP1A1) in colon tissue. Interleukin-22(IL-22) expression in colon tissue was detected by immunofluorescence. The 16S r DNA high-throughput sequencing technique was used to study the modulatory effects of Xiangmei Pills on the intestinal flora structure of rats with UC. Pharmacodynamic results showed that compared with that of the blank group, the colon tissue of the model group was congested, and ulcers were visible in the mucosa; compared with that in the model group, the histopathology of the colon of the rats with UC in the groups of Xiangmei Pills were improved, with scattered ulcers and reduced inflammatory cell infiltration. Chemical analysis showed that a total of 45 components were identified by mass spectrometry information, including 15 phenolic acids, 8 coumarins, 15 organic acids, 3 amino acids, 2 flavonoids, and 2 other components. Compared with those in the blank group, the levels of Ah R, CYP1A1, MUC-2, and IL-22 proteins in the colon tissue of rats in the model group were significantly decreased, and the level of IFN-γ protein was significantly increased; the intestinal flora of rats in the model group was disorganized, with a decrease in the abundance of the flora; the relative abundance of Bacteroidetes,unclassified genera of Ascomycetes, Prevotella of the Prevotella family, and Prevotella decreased significantly, and that of Firmicutes decreased, but the difference was not statistically significant. The relative abundance of Bacteroidetes, Bifidobacterium, and Lactobacillus increased significantly. Compared with those of the model group, the levels of Ah R, CYP1A1, MUC-2, and IL-22proteins in the colonic tissue of the groups of Xiangmei Pills were significantly higher, and the levels of IFN-γ proteins were significantly lower. The recovery of the intestinal flora was accelerated, and the diversity of the intestinal flora was significantly increased. The relative abundance of Bacteroidetes was significantly increased, and that of unclassified genera of Ascomycetes,Lactobacillus, Prevotella of the Prevotella family, and Prevotella was significantly increased. The relative abundance of Bacteroidetes and Bifidobacterium was significantly decreased. This study demonstrated that Xiangmei Pills can effectively treat UC, mainly through the phenolic acid and organic acid components to stimulate the intestinal barrier, regulate protein expression and the relative abundance and diversity of intestinal flora, and play a role in the treatment of UC.
Animals ; Colitis, Ulcerative/metabolism* ; Drugs, Chinese Herbal/chemistry* ; Rats, Sprague-Dawley ; Male ; Rats ; Gastrointestinal Microbiome/genetics* ; Chromatography, High Pressure Liquid ; Humans ; Mass Spectrometry ; RNA, Ribosomal, 16S/genetics* ; Bacteria/drug effects*

Messenger RNA (mRNA) therapeutics involve delivering in vitro transcribed mRNA into specific cells to produce target proteins for the treatment or prevention of diseases. However, the development of mRNA therapeutics relies largely on mRNA delivery systems. Lipid nanoparticles (LNPs) represent the most widely used mRNA carriers in clinical applications. Composed of ionizable lipids, zwitterionic phospholipids, cholesterol, and polyethylene glycol-lipids, LNPs can address critical challenges in mRNA drug development, such as poor in vivo stability and the difficulty in crossing biological barriers. Ultimately, LNPs enable safe, efficient, and targeted mRNA delivery to the liver, lung, spleen, and other organs. This review outlines the roles of the four lipid components in LNPs for mRNA delivery. It then introduces targeted mRNA delivery to various organs/tissues such as the liver, lung, spleen, pancreas, bone marrow, and placenta, using strategies such as antibody modification, lipid structure alteration, and specialized administration routes. Additionally, this review discusses the applications and challenges of LNP-based mRNA therapeutics in disease treatment, aiming to provide insights for the clinical translation of mRNA therapies and for further innovations in LNP delivery systems.
Humans ; RNA, Messenger/administration & dosage* ; Nanoparticles/chemistry* ; Lipids/chemistry* ; Drug Delivery Systems ; Animals ; Liposomes

BACKGROUND: Although many studies have reported the therapeutic effects of hot spring bathing on various diseases, its influence on healthy individuals is not well understood. Myoban Onsen, a sulfur-rich hot spring in Beppu City, Japan, is traditionally believed to improve skin conditions, relieve fatigue, and promote relaxation. However, scientific verification of these effects, particularly their impact on gut microbiota and related metabolic outcomes in healthy individuals, remains scarce. This study aimed to evaluate the effects of Myoban hot spring bathing on gut microbiota composition and SCFA concentrations in healthy individuals. METHODS: In this study, 16 healthy adult males (n = 16) participated in Myoban hot spring bathing four times over two weeks. Fecal samples were collected before and after the intervention, and 16S rRNA sequencing and gas chromatography-mass spectrometry (GC-MS) were performed to analyze gut microbiota composition and organic acid concentrations. The effects of hot spring bathing were evaluated using the Wilcoxon matched-pair signed-rank test to compare pre- and post-intervention. RESULTS: After Myoban hot spring bathing, there was a significant increase in beneficial gut bacteria, Bifidobacterium, Blautia, and Anaerostipes, compared to pre-bathing (p = 0.0012, p = 0.0103, and p = 0.0017, respectively). Conversely, significant decreases were observed in Parabacteroides, Alistipes, and Oscillibacter (p = 0.0125, p = 0.0215, and p = 0.0125, respectively). Significant increases in SCFAs, including acetic acid, propionic acid, and butyric acid, were observed after Myoban hot spring bathing (p = 0.0067, p = 0.0125, and p = 0.0302, respectively). These findings suggest that Myoban hot spring bathing may benefit healthy adult males. CONCLUSIONS: This study suggests that Myoban hot spring bathing may improve gut health in healthy males. The observed increases in beneficial bacteria and SCFAs indicate a potential contribution to improved health status through modulation of the gut environment. TRIAL REGISTRATION Registration number: UMIN000055229, retrospectively registered.
Humans ; Gastrointestinal Microbiome ; Male ; Hot Springs ; Pilot Projects ; Fatty Acids, Volatile/analysis* ; Adult ; Japan ; Feces/chemistry* ; Bacteria/genetics* ; Young Adult ; Baths ; RNA, Ribosomal, 16S/analysis* ; Middle Aged

Nipah virus (NiV) and related viruses form a distinct henipavirus genus within the Paramyxoviridae family. NiV continues to spillover into the humans causing deadly outbreaks with increasing human-bat interaction. NiV encodes the large protein (L) and phosphoprotein (P) to form the viral RNA polymerase machinery. Their sequences show limited homologies to those of non-henipavirus paramyxoviruses. We report two cryo-electron microscopy (cryo-EM) structures of the Nipah virus (NiV) polymerase L-P complex, expressed and purified in either its full-length or truncated form. The structures resolve the RNA-dependent RNA polymerase (RdRp) and polyribonucleotidyl transferase (PRNTase) domains of the L protein, as well as a tetrameric P protein bundle bound to the L-RdRp domain. L-protein C-terminal regions are unresolved, indicating flexibility. Two PRNTase domain zinc-binding sites, conserved in most Mononegavirales, are confirmed essential for NiV polymerase activity. The structures further reveal anchoring of the P protein bundle and P protein X domain (XD) linkers on L, via an interaction pattern distinct among Paramyxoviridae. These interactions facilitate binding of a P protein XD linker in the nucleotide entry channel and distinct positioning of other XD linkers. We show that the disruption of the L-P interactions reduces NiV polymerase activity. The reported structures should facilitate rational antiviral-drug discovery and provide a guide for the functional study of NiV polymerase.
Nipah Virus/chemistry* ; Cryoelectron Microscopy ; Viral Proteins/genetics* ; RNA-Dependent RNA Polymerase/genetics* ; Phosphoproteins/genetics* ; Humans ; Models, Molecular ; Protein Binding

Loss of protein homeostasis is a hallmark of cellular senescence, and ribosome pausing plays a crucial role in the collapse of proteostasis. However, our understanding of ribosome pausing in senescent cells remains limited. In this study, we utilized ribosome profiling and G-quadruplex RNA immunoprecipitation sequencing techniques to explore the impact of RNA G-quadruplex (rG4) on the translation efficiency in senescent cells. Our results revealed a reduction in the translation efficiency of rG4-rich genes in senescent cells and demonstrated that rG4 structures within coding sequence can impede translation both in vivo and in vitro. Moreover, we observed a significant increase in the abundance of rG4 structures in senescent cells, and the stabilization of the rG4 structures further exacerbated cellular senescence. Mechanistically, the RNA helicase DHX9 functions as a key regulator of rG4 abundance, and its reduced expression in senescent cells contributing to increased ribosome pausing. Additionally, we also observed an increased abundance of rG4, an imbalance in protein homeostasis, and reduced DHX9 expression in aged mice. In summary, our findings reveal a novel biological role for rG4 and DHX9 in the regulation of translation and proteostasis, which may have implications for delaying cellular senescence and the aging process.
G-Quadruplexes ; Cellular Senescence ; Ribosomes/genetics* ; Humans ; Animals ; Mice ; DEAD-box RNA Helicases/genetics* ; Protein Biosynthesis ; RNA/chemistry* ; Neoplasm Proteins

OBJECTIVES: The mechanism of the odontogenic differentiation of apical papillary cells (APCs) stimulated by bioactive glass 45S5 is still unclear. This study aims to investigate the effect of autophagy on the odontogenic differentiation of APCs stimulated by bioactive glass 45S5. METHODS: APCs were isolated and cultured in vitro, and the cell origin was identified by flow cytometry. The culture medium was prepared with 1 mg/mL 45S5, and its pH and ion concentration were determined. The experiments were divided into control, 45S5, and 3-methyladenine (3-MA) 45S5 groups. In the 45S5 group, APCs were induced to culture with 1 mg/mL 45S5. In the 3-MA 45S5 group, the autophagy inhibitor 3-MA was added to 1 mg/mL 45S5. Protein immunoblotting assay (Western blot) was used to detect the expression of autophagy-associated proteins of microtubule-associated protein 1 light-chain 3β (LC3B) and P62 after 24 h of induction culture in each group. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the expression of bone sialoprotein (BSP), Runt-related transcription factor 2 (Runx2), dentin sialophosphoprotein (DSPP), and dentin matrix protein-1 (DMP-1) after 7 d of induction culture. Cellular alkaline phosphatase (ALP) staining analyzed cellular ALP activity at 7 d of induction, and alizarin red staining evaluated the formation of mineralized nodules at 21 d of induction. RESULTS: The pH of the 45S5 extract culture medium was 8.65±0.01, which was not significantly different from that of the control group (P>0.05). The silicon ion concentration of the 45S5 induction culture medium was (1.56±0.07) mmol/L, which was higher than that of the control group (0.08±0.01) mmol/L (P<0.05). The calcium ion concentration of the 45S5 induction culture was (1.57±0.15) mmol/L, which was not significantly different from that of the control group (P>0.05). Western blot results showed that LC3B-Ⅱ/Ⅰ ratio increased and P62 expression decreased in the 45S5 group compared with those in the control group (P<0.05). By contrast, the ratio decreased and the expression increased in the 3-MA 45S5 group compared with those in the 45S5 group (P<0.05). RT-qPCR results showed that the expression of BSP, Runx2, DMP-1, and DSPP enhanced in the 45S5 group compared with that in the control group (P<0.05), but the expression decreased in the 3-MA 45S5 group compared with that in the 45S5 group (P<0.05). Semi-quantitative analysis of ALP staining and alizarin red staining showed that the ALP activity was enhanced, and the formation mineralized nodule increased in the 45S5 group compared with those in the control group. The ALP activity weakened, and the formation mineralized nodules were reduced in the 3-MA 45S5 group compared with that those in the 45S5 group. CONCLUSIONS Cell autophagy participates in the odontogenic differentiation of APCs induced by 1 mg/mL 45S5 in vitro.
Autophagy/drug effects* ; Cell Differentiation/drug effects* ; Odontogenesis/drug effects* ; Dental Papilla/cytology* ; Humans ; Microtubule-Associated Proteins/metabolism* ; Glass/chemistry* ; Cells, Cultured ; Core Binding Factor Alpha 1 Subunit/metabolism* ; Extracellular Matrix Proteins/metabolism* ; Ceramics/pharmacology* ; Adenine/pharmacology* ; Sialoglycoproteins/metabolism* ; Phosphoproteins/metabolism* ; Integrin-Binding Sialoprotein/metabolism* ; Alkaline Phosphatase/metabolism* ; RNA-Binding Proteins

With the continuous development of messenger RNA (mRNA) technology, mRNA-based drugs have shown broad application prospects in recent years. Since mRNA is easy to be degraded and difficult to enter cells directly, the mRNA delivery vectors have always been one of the focuses in the development of mRNA-based drugs. Although lipid nanoparticles (LNPs) have been widely used for the delivery of mRNA, they tend to accumulate in the liver, and repeated administration can easily induce inflammatory response which leads to tissue damage. Compared with LNPs, virus-like particles (VLPs) have the advantages of high biocompatibility and safety, being expected to offer new solutions for mRNA delivery. Based on the practical application requirements, this review summarized the research progress in VLPs according to the mRNA delivery steps: particle assembly, delivery into cells, and intracellular release. We hope to provide a basis and design ideas for the development of new VLPs as delivery vectors, promote the application of VLPs in mRNA delivery, and provide new possibilities for the research and application of mRNA-based therapeutics.
RNA, Messenger/administration & dosage* ; Humans ; Nanoparticles/chemistry* ; Genetic Vectors ; Lipids/chemistry* ; Drug Delivery Systems/methods* ; Virion ; Animals ; Gene Transfer Techniques ; Liposomes

The development of safe and effective carriers is crucial for improving the in vivo stability of siRNA drugs and facilitating their clinical translation. Chitosan (CS), a natural cationic polymer, shows great potential in nucleic acid drug delivery. To optimize the physicochemical properties of CS/siRNA nanoparticles (NPs) and increase their siRNA delivery efficacy, in this study, hyaluronic acid (HA) was added into CS to form stable complex NPs through electrostatic interactions. The HA component is able to target the CD44 receptors on the surface of tumor cells, facilitating efficient siRNA delivery. First, we systematically investigated the effects of the molecular weights and mass ratio of CS and HA on the physicochemical properties of CS/HA NPs. The results showed that at HA: CS mass ratios of approximately 5:5 and 6:4, the complex NPs exhibited small particle sizes, narrow size distribution, and high storage stability. Under similar conditions, the size of CS/HA NPs increased with the increase in the molecular weights of CS and HA. Based on these findings, suitable conditions were selected to prepare CS/HA NPs for siRNA delivery. Cell experiments demonstrated that the introduction of HA effectively reduced the cytotoxicity of the CS delivery system and enhanced the NP uptake. The CS/HA/siRNA NPs achieved 50% to 60% silencing of the luciferase gene in HeLa-Luc cells. CS/HA NPs formed smaller nanoparticles with siRNA than pure CS and mediated specific interactions with tumor cells via HA, leading to efficient siRNA delivery. These findings provide valuable insights into the construction of natural polymer composite nanoparticles for application in siRNA delivery.
Hyaluronic Acid/chemistry* ; Chitosan/chemistry* ; RNA, Small Interfering/administration & dosage* ; Nanoparticles/chemistry* ; Humans ; Particle Size ; HeLa Cells ; Hyaluronan Receptors

This study aims to investigate the potential of oncolytic nanoparticles encapsulating Coxsackievirus A21 (CVA21) full-genome mRNA (CVA21@ONP) to resurrect CVA21 and induce apoptosis in host cells, as well as the antitumor immune effects of CVA21@ONP in immunocompetent tumor-bearing BALB/c mice. We used lipid nanoparticles (LNPs) to encapsulate CVA21 full-genome mRNA, thus preparing CVA21@ONP. The killing efficacy of CVA21@ONP was determined by the plaque assay and cell counting kit-8 (CCK-8), and the apoptosis in HT29 and CT26-iRFP cells was evaluated by flow cytometry. Mice were administrated with CVA21@ONP at high and low doses intratumorally, and the growth of tumors expressing infra-red fluorescent protein (iRFP) was monitored. Additionally, the types and changes of immune cells in the spleen were analyzed by flow cytometry. The results demonstrated that CVA21@ONP successfully resurrected CVA21 in both HT29 and U87MG cells. The plaque assay revealed robust killing effects of CVA21@ONP against both human and murine cell lines, and flow cytometry results showed increased early and late apoptotic cells. Notably, intratumoral detection revealed significantly down-regulated expression of iRFP in both high- and low-dose CVA21@ONP groups. Flow cytometry results further indicated that CVA21@ONP treatment effectively reduced the levels of immunosuppressive cells, including myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs), in the spleen, while enhancing T cell-dependent antitumor immune responses. These findings suggest that CVA21@ONP can replicate and survive extensively both in vitro and in vivo, activating the immune system of mice administrated with CVA21@ONP to target cells at the tumor site, thereby remodeling the tumor immune microenvironment and accelerating the suppression or even complete regression of tumors. The oncolytic performance of CVA21@ONP has been verified through intratumoral injection administration in this study, aimed at further exploring its therapeutic potential and promoting the development of the field of tumor treatment.
Animals ; Nanoparticles/chemistry* ; Mice ; Mice, Inbred BALB C ; Humans ; Apoptosis ; Oncolytic Viruses/genetics* ; Oncolytic Virotherapy/methods* ; Cell Line, Tumor ; RNA, Messenger/genetics* ; HT29 Cells

The production of healthy agricultural products has increased the demand for innovative and sustainable plant protection technologies, and the rapid advancement of nanotechnology has brought revolutionary breakthroughs to traditional agriculture. Nanocarrier-based drug delivery systems can not only significantly improve the utilization efficiency of pesticides, achieving enhanced efficacy and reduced application, but also decrease the pesticide residues and environmental pollution. Additionally, they have made breakthrough progress in the stability and persistence of RNA pesticides. This review summarized the research progress on nanopesticides and RNA pesticides, focusing on the mechanisms of nanocarriers in improving pesticide bioactivity and RNA interference (RNAi) efficiency. It also systematically summarized the types of nanomaterials and their applications in pest and disease management and provided an in-depth outlook for the future development of nanopesticides and RNA pesticides, which provided technical support for the high-quality development of agriculture in the future.
Pesticides/chemistry* ; Nanotechnology ; Nanostructures ; RNA ; Agriculture/methods* ; RNA Interference ; Drug Delivery Systems