Lanzhou Institute of Biological Products Co. Ltd.

A new chitinase gene, cloned from the biocontrol Chaetomium globosum W7, was designated Cgchi18. Recombinant protein Cgchi18 with 535 amino acids was expressed in Escherichia coli, and purified by means of a column-free purification method relying on split intein, achieving a 12.39-fold purification and a 15.61% recovery yield. The maximum activity of this approximately 60-kDa protein was observed at 45 °C and pH 5.0. Cgchi18 was activated by Mg²⁺ and Ba²⁺, but inhibited by Mn²⁺, Co²⁺, Cu²⁺, Zn²⁺, Ag⁺ and Hg²⁺. Cgchi18 showed high substrate specificity, only hydrolyzing β-1,4-glycoside bond in chitin and its derivatives, to liberate disaccharides or trisaccharides. For the degradation of colloidal chitin under optimal conditions, Vmax and Km of Cgchi18 were calculated as 8.05 μmol/min/mg and 3.18 mg/mL, respectively. Additionally, it exhibited antifungal activity and could have a degrading effect on the spread of hyphae of pathogenic fungi. In conclusion, the chitinase Cgchi18 identified from C. globosum has potential for industrial and agricultural applications.

This study aims to explore the potential mechanism by which Botulinum toxin type A (BoNT/ A) inhibits microglial inflammatory activation through P2X7 receptors (P2X7R).

BoNT/A is a promising analgesic drug, and previous studies have established that it alleviates Neuropathic Pain (NP) by inhibiting microglial inflammatory activation. This study examined the biomarkers and potential mechanisms by which BoNT/A relieves neuropathic pain by mediating microglial P2X7R and analyzing transcriptome sequencing data from mouse BV-2 microglial cells.

The P2X7R agonist Bz-ATP was used to induce microglial inflammatory activation, whilst RNAseq technology was used to explore the biomarkers and potential mechanisms through which BoNT/A suppresses microglial inflammation.

RNA sequencing was performed on three BV-2 cell samples treated with a P2X7R specific activator (Bz-ATP) and three BV-2 cell samples pre-treated with BoNT/A. Only data that successfully passed quality control measures were included in subsequent analysis. Initially, Differentially Expressed Genes (DEGs) were identified from BoNT/A and control samples, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. Biomarkers were then identified by constructing a Protein- Protein Interaction (PPI) network and utilizing the CytoHubba plug-in in Cytoscape software. Lastly, enrichment analysis and regulatory network analysis were performed to elucidate the potential mechanism of BoNT/A in the treatment of NP.

93 DEGs related to the “cell component size regulation” GO term and enriched in the “axon guidance” KEGG pathway were identified. Subsequently, 6 biomarkers were identified, namely PTPRF, CHDH, CKM, Ky, Sema3b, and Sema3f, which were enriched in pathways related to biosynthesis and metabolism, disease progression, signal transduction, and organelle function, including the “ribosome” and “Wnt signaling pathway.” Finally, a competing endogenous RNA (ceRNAs) network was constructed from 6 mRNAs, 66 miRNAs, and 31 lncRNAs, forming a complex relationship network.

Six genes (PTPRF, Sema3b, Sema3f, CHDH, CKM, and Ky) were identified as biomarkers of microglial inflammatory activation following BoNT/A treatment. This finding may provide a valuable reference for the relief and treatment of neuropathic pain.