Effect of human umbilical cord mesenchymal stem cells in treatment of mice with liver fibrosis and its mechanism / 临床肝胆病杂志

ObjectiveTo investigate the effect of human umbilical cord mesenchymal stem cells （hUCMSCs） in the treatment of mice with liver fibrosis and its mechanism. MethodsA total of 18 specific pathogen-free C57BL/6 mice， aged 6 weeks， were selected and divided into control group （n=6）， carbon tetrachloride （CCl4） model group （CCl4 group， n=6）， and hUCMSCs treatment group （MSC group， n=6） using a random number table. The mice in the CCl4 group and the MSC group were given intraperitoneal injection of CCl4 solution to establish a mouse model of liver fibrosis， while those in the control group were injected with the same dose of corn oil， and the mice in the MSC group were injected with hUCMSCs via the caudal vein during the injection of CCl4. At the end of week 8， mouse serum was collected， and the mice were sacrificed to collect and fix the liver. Enzyme-linked immunosorbent assay was used to measure the levels of inflammatory factors； an automatic biochemical detector was used to measure liver function parameters； HE staining， Masson staining， Sirius Red staining， and α-SMA immunofluorescence assay were used to evaluate liver fibrosis. Hepatic stellate cells （HSCs） stimulated by TGF-β were co-cultured with hUCMSCs in the medium with or without chitinase-3 like-protein-1 （CHI3L1）， and Western blot was used to measure the expression levels of proteins. A one-way analysis of variance was used for comparison of continuous data between multiple groups， and the Dunnett’s t-test was used for further comparison between two groups. ResultsMasson staining and Sirius Red staining showed that the CCl4 group had a significantly higher degree of fibrosis than the control group （both P<0.05）， and the MSC group had significant alleviation of fibrosis compared with the CCl4 group （both P<0.05）. Compared with the control group， the CCl4 group had significant increases in the levels of interleukin-1β， interleukin-6 （IL-6）， aspartate aminotransferase （AST）， alanine aminotransferase （ALT）， and alkaline phosphatase （ALP） （all P<0.05）， and compared with the CCl4 group， the MSC group had significant reductions in the levels of IL-6， AST， ALT， and ALP （all P<0.05）. The CCl4 group had significantly higher expression levels of CHI3L1 and α-SMA than the control group and the MSC group （all P<0.05）. The cell culture experiment showed that the MSC+HSC group had a significantly higher expression level of Bax than the HSC group and the MSC+CHI3L1 group （both P<0.05）， suggesting that CHI3L1 reversed the pro-apoptotic effect of MSC on activated HSCs. ConclusionThis study shows that hUCMSCs can improve liver fibrosis in mice， possibly by inhibiting CHI3L1 to promote the apoptosis of HSCs.

Isolation and Cs+ resistance mechanism of Escherichia coli strain ZX-1.

This research aims to elucidate the physiological mechanisms behind the accidental acquisition of high-concentration cesium ions (Cs+) tolerance of Escherichia coli and apply this understanding to develop bioremediation technologies. Bacterial Cs+ resistance has attracted attention, but its physiological mechanism remains largely unknown and poorly understood. In a prior study, we identified the Cs+/H+ antiporter TS_CshA in Microbacterium sp. TS-1, resistant to high Cs+ concentrations, exhibits a low Cs+ affinity with a Km value of 370 mM at pH 8.5. To enhance bioremediation efficacy, we conducted random mutagenesis of TS_cshA using Error-Prone PCR, aiming for higher-affinity mutants. The mutations were inserted downstream of the PBAD promoter in the pBAD24 vector, creating a mutant library. This was then transformed into E. coli-competent cells. As a result, we obtained a Cs+-resistant strain, ZX-1, capable of thriving in 400 mM CsCl-a concentration too high for ordinary E. coli. Unlike the parent strain Mach1™, which struggled in 300 mM CsCl, ZX-1 showed robust growth even in 700 mM CsCl. After 700 mM CsCl treatment, the 70S ribosome of Mach1™ collapsed, whereas ZX-1 and its derivative ΔZX-1/pBR322ΔAp remained stable. This means that the ribosomes of ZX-1 are more stable to high Cs+. The inverted membrane vesicles from strain ZX-1 showed an apparent Km value of 28.7 mM (pH 8.5) for Cs+/H+ antiport activity, indicating an approximately 12.9-fold increase in Cs+ affinity. Remarkably, the entire plasmid isolated from ZX-1, including the TS_cshA region, was mutation-free. Subsequent whole-genome analysis of ZX-1 identified multiple SNPs on the chromosome that differed from those in the parent strain. No mutations in transporter-related genes were identified in ZX-1. However, three mutations emerged as significant: genes encoding the ribosomal bS6 modification enzyme RimK, the phage lysis regulatory protein LysB, and the flagellar base component protein FlgG. These mutations are hypothesized to affect post-translational modifications, influencing the Km value of TS_CshA and accessory protein expression. This study unveils a novel Cs+ resistance mechanism in ZX-1, enhancing our understanding of Cs+ resistance and paving the way for developing technology to recover radioactive Cs+ from water using TS_CshA-expressing inverted membrane vesicles.