The Mechanism of Action of the Active Ingredients of Coptidis rhizoma against Porcine Epidemic Diarrhea Was Investigated Using Network Pharmacology and Molecular Docking Technology.

The objective of this study was to elucidate the mechanism of action of the active components of Coptidis rhizoma against porcine epidemic diarrhea and to provide a theoretical foundation for further development of novel anti-PED therapeutic agents based on Coptidis rhizoma. The potential targets of Coptidis rhizoma against PEDV were identified through a comprehensive literature review and analysis using the TCMSP pharmacological database, SwissDrugDesign database, GeneCards database, and UniProt database. Subsequently, the STRING database and Cytoscape 3.7.1 software were employed to construct a protein-protein interaction (PPI) network and screen key targets. Gene Ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were conducted on the identified targets. Molecular docking studies were performed using AutoDock 1.5.7 software to analyze the binding energy and modes of interaction between the active components of Coptidis rhizoma and the target proteins. The PyMOL 2.5.0a0 software was employed to visualize the docking results. Through comprehensive analysis, 74 specific targets of active components of Coptidis rhizoma against PEDV were identified. The core gene targets were screened, and an interaction network diagram was subsequently generated. Ultimately, 14 core targets were identified, with STAT3, ESR1, CASP3, and SRC exhibiting the most significant interactions. GO enrichment analysis revealed a total of 215 molecular items, including 48 biological function items, 139 biological process items, and 28 cellular component items. KEGG enrichment analysis identified 140 signaling pathways. Molecular docking analysis demonstrated that epiberberine and palmatine exhibited high binding affinity with STAT3 protein, worenine showed high binding affinity with ESR1 protein, obacunone exhibited high binding affinity with CASP3 protein, and epiberberine, obacunone, berberine, and berberruine exhibited high binding affinity with SRC protein. A network pharmacology and molecular docking technology approach was employed to screen six important active components of Coptidis rhizoma and four important potential targets against PEDV infection. The findings indicated that the active components of Coptidis rhizoma could serve as promising pharmaceutical agents for the prevention and control of PEDV, with significant potential for clinical application.

A strategy for improving silk yield and organ size in silk-producing insects.

Insect silks possess excellent biodegradability, biocompatibility and mechanical properties, and have numerous applications in biomedicine and tissue engineering. However, the application of silk fiber is hindered by its limited supply, especially from non-domesticated insects. In the present study, the silk yield and organ size of Bombyx mori were significantly improved through genetic manipulation of the target of rapamycin complex 1 (TORC1) pathway components. Silk protein synthesis and silk gland size were decreased following rapamycin treatment, inhibiting the TORC1 signaling pathway both in vivo and ex vivo. The overexpression of posterior silk gland-specific Rheb and BmSLC7A5 improved silk protein synthesis and silk gland size by activating the TORC1 signaling pathway. Silk yield in BmSLC7A5-overexpression silkworms was significantly increased by approximately 25%. We have demonstrated that the TORC1 signaling pathway is involved in the transcription and translation of silk genes and transcriptional activators via phosphorylation of p70 S6 kinase 1 and 4E-binding protein 1. Our findings present a strategy for increasing silk yield and organ size in silk-producing insects.

Serpin-1a and serpin-6 regulate the Toll pathway immune homeostasis by synergistically inhibiting the Spätzle-processing enzyme CLIP2 in silkworm, Bombyx mori.

The Toll receptor signaling pathway is an important innate immune response of insects to pathogen infection; its extracellular signal transduction involves serine protease cascade activation. However, excessive or constitutive activation of the Toll pathway can be detrimental. Hence, the balance between activation and inhibition of the extracellular protease cascade must be tightly regulated to achieve favorable outcomes. Previous studies have shown that serpins-serine protease inhibitors-negatively regulate insect innate immunity by inhibiting extracellular protease cascade signaling. Although the roles of serpins in insect innate immunity are well described, the physiological mechanisms underlying their synergistic effects remain poorly understand. Here, we characterize the molecular mechanism by which serpin-1a and serpin-6 synergistically maintain immune homeostasis of the silkworm Toll pathway under physiological and pathological conditions. Through in vitro biochemical assays and in vivo bioassays, we demonstrate that clip-domain serine protease 2 (CLIP2), as the Toll cascade-activating terminal protease, is responsible for processing proSpätzle1 to induce the expression of antimicrobial peptides. Further biochemical and genetic analyses indicate that constitutively expressed serpin-1a and inducible serpin-6 synergistically target CLIP2 to maintain homeostasis of the silkworm Toll pathway under physiological and pathological conditions. Taken together, this study provides new insights into the precise regulation of Toll cascade activation signals in insect innate immune responses and highlights the importance and complexity of insect immune homeostasis regulation.

Dynamic Changes and Characterization of the Metal Ions in the Silk Glands and Silk Fibers of Silkworm.

Metal ions are involved in the conformational transition of silk fibroin and influence the structure and mechanical properties of silk fibers. However, the dynamic characteristics of metal ions during the formation of silk fibers remain unclear. In this study, we found that the silk glands of silkworms contain various metal elements, with varying levels of the metal elements in different zones of the glands and higher levels in the anterior silk glands. Additionally, the content of various metallic elements in the silk glands varied greatly before and after spinning, similar to their content in different cocoon layers, thus, indicating that the anterior silk glands maintain a certain metal ion environment for the transport and conformational transformation of the silk proteins. Most of the metallic elements located in fibroin were confirmed using degumming experiments. For the first time, a scanning electron microscope energy spectrometry system was used to characterize the metal elements in the cross-section of silk and cocoons. These findings have deepened our understanding of the relationship between the overall metal ion environment and silk fiber formation and help us further conceptualize the utilization of metal ions as targets to improve the mechanical properties of the silk fibers.