Generation of cytotoxic aptamers specifically targeting fibroblast-like synoviocytes by CSCT-SELEX for treatment of rheumatoid arthritis.

OBJECTIVES: Rheumatoid arthritis (RA) is an autoimmune disease characterised by aggressive fibroblast-like synoviocytes (FLSs). Very few RA patients-derived FLSs (RA-FLSs)-specific surface signatures have been identified, and there is currently no approved targeted therapy for RA-FLSs. This study aimed to screen therapeutic aptamers with cell-targeting and cytotoxic properties against RA-FLSs and to uncover the molecular targets and mechanism of action of the screened aptamers. METHODS: We developed a cell-specific and cytotoxic systematic evolution of ligands by exponential enrichment (CSCT-SELEX) method to screen the therapeutic aptamers without prior knowledge of the surface signatures of RA-FLSs. The molecular targets and mechanisms of action of the screened aptamers were determined by pull-down assays and RNA sequencing. The therapeutic efficacy of the screened aptamers was examined in arthritic mouse models. RESULTS: We obtained an aptamer SAPT8 that selectively recognised and killed RA-FLSs. The molecular target of SAPT8 was nucleolin (NCL), a shuttling protein overexpressed on the surface and involved in the tumor-like transformation of RA-FLSs. Mechanistically, SAPT8 interacted with the surface NCL and was internalised to achieve lysosomal degradation of NCL, leading to the upregulation of proapoptotic p53 and downregulation of antiapoptotic B-cell lymphoma 2 (Bcl-2) in RA-FLSs. When administrated systemically to arthritic mice, SAPT8 accumulated in the inflamed FLSs of joints. SAPT8 monotherapy or its combination with tumour necrosis factor (TNF)-targeted biologics was shown to relieve arthritis in mouse models. CONCLUSIONS: CSCT-SELEX could be a promising strategy for developing cell-targeting and cytotoxic aptamers. SAPT8 aptamer selectively ablates RA-FLSs via modulating NCL-p53/Bcl-2 signalling, representing a potential alternative or complementary therapy for RA.

Synthesis of metal framework-modified carbon dots with super large stokes shift using Hami melon as a green precursor for detecting thiophanate-methyl residue in leafy vegetables.

Consuming leafy vegetables with excessive thiophanate-methyl (TM) residue poses serious risks to human health. To realize rapidly and sensitively detecting TM in leafy vegetables, we developed a fluorescent probe based on zeolitic imidazolate framework-8-modified carbon dots using Hami melon as the green precursor (HM-CDs@ZIF-8). Meanwhile, the mechanism of HM-CDs@ZIF-8 for detecting TM was investigated and explained. The results of the performance tests showed that the prepared HM-CDs@ZIF-8 exhibited high sensitivity, excellent selectivity, robust anti-interference capability, reliable reproducibility and repeatability, and long-term stability. After optimization experiments, the fluorescence intensity of HM-CDs@ZIF-8 showed a strong linear correlation with the concentration of TM (0.00171-3.4239 mol/L) with a detection limit of 2.025 µmol/L. The HM-CDs@ZIF-8 was successfully applied to determine TM in spiked leafy vegetables with satisfactory recoveries of 96-105%. The relative standard deviations were in the range of 0.26-2.55%. The sensor has a promising application for detecting TM in leafy vegetables.

Host-Gut Microbiota Metabolic Interactions and Their Role in Precision Diagnosis and Treatment of Gastrointestinal Cancers.

The critical role of the gut microbiome in gastrointestinal cancers is becoming increasingly clear. Imbalances in the gut microbial community, referred to as dysbiosis, are linked to increased risks for various forms of gastrointestinal cancers. Pathogens like Fusobacterium and Helicobacter pylori relate to the onset of esophageal and gastric cancers, respectively, while microbes such as Porphyromonas gingivalis and Clostridium species have been associated with a higher risk of pancreatic cancer. In colorectal cancer, bacteria such as Fusobacterium nucleatum are known to stimulate the growth of tumor cells and trigger cancer-promoting pathways. On the other hand, beneficial microbes like Bifidobacteria offer a protective effect, potentially inhibiting the development of gastrointestinal cancers. The potential for therapeutic interventions that manipulate the gut microbiome is substantial, including strategies to engineer anti-tumor metabolites and employ microbiota-based treatments. Despite the progress in understanding the influence of the microbiome on gastrointestinal cancers, significant challenges remain in identifying and understanding the precise contributions of specific microbial species and their metabolic products. This knowledge is essential for leveraging the role of the gut microbiome in the development of precise diagnostics and targeted therapies for gastrointestinal cancers.

Repurposing AS1411 for constructing ANM-PROTACs.

Proteolysis-targeting chimeras (PROTACs) are heterobifunctional molecules consisting of two ligands joined by a linker, enabling them to simultaneously bind with an E3 ligase and a protein of interest (POI) and trigger proteasomal degradation of the POI. Limitations of PROTAC include lack of potent E3 ligands, poor cell selectivity, and low permeability. AS1411 is an antitumor aptamer specifically recognizing a membrane-nucleus shuttling nucleolin (NCL). Here, we repurpose AS1411 as a ligand for an E3 ligase mouse double minute 2 homolog (MDM2) via anchoring the NCL-MDM2 complex. Then, we construct an AS1411-NCL-MDM2-based PROTAC (ANM-PROTAC) by conjugating AS1411 with large-molecular-weight ligands for "undruggable" oncogenic STAT3, c-Myc, p53-R175H, and AR-V7. We show that the ANM-PROTAC efficiently penetrates tumor cells, recruits MDM2 and degrades the POIs. The ANM-PROTAC achieves tumor-selective distribution and exhibits excellent antitumor activity with no systemic toxicity. This is a PROTAC with built-in tumor-targeting and cell-penetrating capacities.

Hepatitis B virus-mediated sodium influx contributes to hepatic inflammation via synergism with intrahepatic danger signals.

The NACHT, LRR, and PYD domains-containing protein 3 (NLRP3) inflammasome has been involved in the pathogenesis of various chronic liver diseases. However, its role in hepatitis B virus (HBV)-associated hepatitis remains unknown. Here we demonstrate the synergistic effect of HBV with potential intrahepatic danger signals on NLRP3 inflammasome activation. HBV exposure at the appropriate temporal points enhances potassium efflux-dependent NLRP3 inflammasome activation in macrophages and also increases NLRP3 inflammasome-mediated inflammation in HBV-transgenic mouse model. HBV-mediated synergism with intrahepatic signals represented by ATP molecules on NLRP3 activation was observed via relevance analysis, confocal microscopy, and co-immunoprecipitation, and its effector cytokines exhibit positive associations with hepatic inflammation in patients with severe hepatitis B. Furthermore, the synergism of HBV on NLRP3 inflammasome activation owes to increased sodium influx into macrophages. Our data demonstrate that HBV contributes to hepatic inflammation via sodium influx-dependent synergistic activation of NLRP3 inflammasome, which provides a deeper understanding of immune pathogenesis in HBV-associated hepatitis.

The elucidation of the anti-inflammatory mechanism of EMO in rheumatoid arthritis through an integrative approach combining bioinformatics and experimental verification.

Introduction: Emodin (EMO), a natural derivative of the anthraquinone family mainly extracted from rhubarb (Rheum palmatum), has previously been demonstrated to possess superior anti-inflammatory properties from a single target or pathway. In order to explore the underlying mechanism of action of EMO against rheumatoid arthritis (RA), a network pharmacology approach was employed. Methods: A gene expression profile from GSE55457 available from the Gene Expression Omnibus (GEO) database was used to identify the targets of EMO action. Further, single cell RNA sequencing data from GEO database of RA patients (GSE159117) were downloaded and analysed. To further investigate the anti-RA effect of EMO on MH7A cells, the expression of IL-6 and IL-1ß were monitored. Finally, RNA-seq analyses were conducted on synovial fibroblasts from EMO-treated. Result: We screened the key targets of EMO against RA using network pharmacology methods, including HMGB1, STAT1, EGR1, NR3C1, EGFR, MAPK14, CASP3, CASP1, IL4, IL13, IKBKB and FN1, and their reliability was verified using ROC curve. Single-cell RNA sequencing data analysis showed that these core target proteins mainly played a role by modulating monocytes. The anti-RA effect of EMO was further verified with MH7A cells, which showed that EMO could block cell differentiation and reduce the expression of IL-6 and IL-1ß. WB experiments confirmed that EMO could affect the expression of COX2, HMBG1 and the phosphorylation of p38. Finally, sequencing of synovial fibroblasts from rats treated with EMO showed consistent results with those predicted and verified, further proving the anti-inflammatory effect of EMO. Conclusion: Our research shows that EMO inhibits inflammatory response of rheumatoid arthritis (RA) by targeting HMGB1, STAT1, EGR1, NR3C1, EGFR, MAPK14, CASP3, CASP1, IL4, IL13, IKBKB, FN1 and Monocytes/macrophages.

Deciphering the action mechanism of paeoniflorin in suppressing pancreatic cancer: A network pharmacology study and experimental validation.

Background: Paeoniflorin (PF) is the main active component of Chinese herbaceous peony that has been shown to have an anti-tumor effect. However, there are few studies on the prevention and treatment of pancreatic cancer with PF. Methods: We gathered Microarray data pertaining to paeoniflorin intervention in pancreatic cancer by utilizing the GEO database (GSE97124). Then, the DEGs were filtered by the 33R program. RNA-seq data of pancreatic cancer and normal tissue samples were taken from the TCGA and GTEx databases, respectively, and the WGCNA technique was utilized to examine the pancreatic cancer-specific genes. Paeoniflorin target genes for the treatment of pancreatic cancer were determined based on the overlap between DEGs and WGCNA. GO and KEGG enrichment analyses were then performed on paeoniflorin target genes to discover which biological processes were impacted. Using the 3 hierarchical methods included in the Cytohubba plugin, we re-screened the hub genes in the target genes to find the genes most relevant to paeoniflorin treatment. The overall survival effects of hub genes were confirmed using the TCGA database. Finally, the paeoniflorin targets identified by the network pharmacology analysis were validated using PANC-1 and Capan-2 cells. Results: We identified 148 main potential PF targets, and gene enrichment analysis suggested that the aforementioned targets play a crucial role in the regulation of MAPK, PI3K-AKT, and other pathways. The further screening of the prospective targets resulted in the identification of 39 hub genes. Using the TCGA database, it was determined that around 33.33% of the hub gene's high expression was linked with a bad prognosis. Finally, we demonstrated that PF inhibits IL-6 and IL-10 expression and p38 phosphorylation in pancreatic cancer cells, thereby reducing inflammation. Conclusion: PF may regulate inflammatory factors mainly through the p38 MAPK signal pathway. These findings provide theoretical and experimental evidence suggesting the PF as a promising natural source of anti-tumor compounds for pancreatic cancer.

Metabolomics combined with network pharmacology to explore the mechanisms of modified Guishen pill to ameliorate polycystic ovary syndrome.

BACKGROUND: The modified Guishen pill (MGP) has a prominent therapeutic effect on polycystic ovary syndrome (PCOS). However, its mechanism is still unclear. This study aimed to uncover the mechanism of MGP for PCOS treatment through a comprehensive strategy integrating metabolomics and network pharmacology. METHODS: A letrozole-induced PCOS model was used to evaluate ovarian function in rats. Plasma metabolomics was used to authenticate differential metabolites and enriched related pathways using the MetaboAnalyst platform. Network pharmacology was utilized to explore the endogenous targets of MGP treatment for PCOS. Finally, the potential targets and related biological functions were verified experimentally. RESULTS: MGP improved PCOS symptoms by regulating abnormal levels of sex hormones and alleviating ovarian pathological changes in rats; fifty-four potential differential metabolites involved in MGP treatment for PCOS, and the hub genes derived from network pharmacology were consistent with the metabolomic analysis results to varying degrees. The comprehensive analysis identified that a key novel target for endothelial nitric oxide synthase (eNOS/NOS3), five key metabolites (ornithine, citrulline, l-glutamic acid, acetylornithine, and hydroxyproline), and one pathway (arginine and proline metabolism) were related to the therapy of PCOS with MGP. Subsequently, we verified the localization and expression of eNOS in the ovaries, and it significantly improved insulin resistance, apoptosis, and oxidative stress in letrozole-induced PCOS rats. CONCLUSION: Our work reveals the complex mechanism of MGP therapy for PCOS. This study is a successful paradigm for elucidating the pharmacological mechanism of the traditional Chinese medicine compound.

Electro-Acupuncture at Zusanli Acupoint (ST36) Suppresses Inflammation in Allergic Contact Dermatitis Via Triggering Local IL-10 Production and Inhibiting p38 MAPK Activation.

Acupuncture has shown beneficial effect in the treatment of multiple dermatologic conditions including dermatitis, pruritus, urticaria, and hyperhidrosis; however, the detailed mechanisms are still kept unclear. This study aimed to investigate if electro-acupuncture (EA) treatment prevents 2,4-dinitrofluorobenzene (DNFB)-induced allergic contact dermatitis (ACD) in rats and explore its underlying mechanisms. ACD was induced by sensitizing and challenging with DNFB topically. Rats were treated daily following bilateral subcutaneous stimulation of EA at Zusanli acupoint (ST36) for 1 week. Ear swelling and serum IgE levels were measured. The ear biopsies were obtained for histology. Inflammatory cytokines on the dermatological ear and local acupoint tissue were assayed. Spleen lymphocytes and the homogenized supernatant of local acupuncture area were used to co-culture for flow cytology and immune analysis, respectively. EA treatment at ST36 notably inhibited ear swelling and inflammatory cell infiltration on DNFB-induced ACD. EA also decreased serum IgE concentrations and alleviated the production of inflammatory cytokines in dermatological ear. Additionally, EA treatment attenuated the percentage of CD4+IFN-γ+ and CD4+IL-4+ T cells associated with ACD. Interestingly, secretion of interleukin (IL)-10 in the local acupoint tissue following EA stimulation was increased and showed suppressive function when co-cultured with the spleen lymphocytes from DNFB group. Lastly, EA treatment demonstrably suppressed p38 MAPK activation in DNFB-treated rats. Our findings suggest that EA treatment at ST36 may ameliorate inflammation associated with DNFB-induced ACD via triggering local IL-10 production and inhibiting p38 MAPK activation, which provide an alternative and promising therapy for ACD.