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OBJECTIVE@#To explore the effect of leucine-rich α-2-glycoprotein (LRG1) derived from hepatocytes on activation of hepatic M1 Kupffer cells.@*METHODS@#A metabolic dysfunction-associated fatty liver disease (MAFLD) model was established in BALB/c mice by high-fat diet (HFD) feeding for 16 weeks. Oleic acid was used to induce steatosis in primary cultures of mouse hepatocytes. The mRNA and protein expressions of LRG1 in mouse liver tissues and hepatocytes were detected by real-time PCR and Western blotting. Primary hepatic macrophages were stimulated with the conditioned medium (CM) from steatotic hepatocyte along with LRG1 or transforming growth factor-β1 (TGF-β1), or both for 24 h, and the expression levels of inducible nitric oxide synthase (iNOS) was detected with Western botting, and the mRNA expressions of iNOS, chemokine ligand 1 (CXCL-1) and interleukin-1β (IL-1β) were measured by RT-PCR. The MAFLD mice were injected with LRG1 (n=6), TGF-β1 (n=6), or both (n=6) through the caudal vein, and the live tissues were collected for HE staining and immumohistochemical detection of F4/80 expression; the mRNA expressions of iNOS, CXCL-1 and IL-1β in liver tissues were detected using RT-PCR.@*RESULTS@#The mRNA and protein expression levels of LRG1 were significantly downregulated in the liver tissues of MAFLD mice and steatotic hepatocytes (P < 0.05). Treatment of the hepatic macrophages with CM from steatosis hepatocytes significantly enhanced the mRNA expression levels of iNOS, CXCL-1 and IL-1β, and these changes were significantly inhibited by the combined treatment with TGF-β1 and LRG1 (P < 0.05). In MAFLD mice, injections with either LRG1 or TGF-β1 alone reduced hepatic lipid deposition and intrahepatic macrophage infiltration, and these effects were significantly enhanced by their combined treatment, which also more strongly inhibited the mRNA expression levels of iNOS, CXCL-1 and IL-1β (P < 0.05).@*CONCLUSION@#LRG1 inhibits hepatic macrophage infiltration by enhancing TGF-β1 signaling to alleviate fatty liver inflammation in MAFLD mice.
Subject(s)
Animals , Mice , Transforming Growth Factor beta1 , Macrophage Activation , Signal Transduction , Non-alcoholic Fatty Liver Disease , Culture Media, Conditioned , GlycoproteinsABSTRACT
Objective To illustrate that tumor necrosis factor-α (TNF-αt) / nuclear factor of activated T cells pathway is the main function way that mesenchymal stem cells (MSC) inhibit proliferation of pulmonary vascular smooth muscle cells during treating pulmonary hypertension (PH).Methods MSC from human umbilical cord was used to treat PH rat models induced by monocrotaline.Rats were divided into the control group,the PH model group and the MSC group.The general conditions of the rats were observed.Haemodynamics was detected.Pathological sections and immunohistochemistry method were used to detect the lung structure and tissue changes.Changing conditions of TNF-αt/NFAT were detected.Results Compared with rats in the PH model group,the general conditions of the MSC group tended to be normal evidently:the right ventricular systolic pressure (RVSP) dropped [(30.37 ±3.13) mmHg vs.(47.90 ± 3.45) mmHg,1 mmHg =0.133 kPa],the aortic pressure (MAoP) increased [(115.03 ± 16.01) mmHg vs.(92.78 ± 16.28 mmHg)],the thickening condition of arterial intima-media was evidently relieved [(17.22 ±1.21)％ vs.(31.68 ±2.26)％],the plasma TNF-α level decreased obviously [(842 ±76) ng/L vs.(245 ±24)ng/L],and the lung tissue TNF-o level decreased (0.172 ±0.024 vs.0.248 ± 0.051),and all the differences were statistically significant (all P ＜ 0.05).The activation of pulmonary artery NFATc2 in the MSC treatment group was apparently inhibited.Conclusions MSC therapy may perform the treating effect in PH by inhibiting the over-proliferation of inflammation related pulmonary vascular smooth muscle cells via TNF-oα/NFAT pathway.
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Objective Lamins are the major components of nuclear lamina underneath the inner nuclear membrane (INM).Lamins express in most cells and are involved in the whole process of growth, also play a major role in cell stability and embryonic development.Mutant in human LMNA gene may lead to a series of disorders, which are similar to progeria or other aging-associate syndrome.In this study, we report a new lmna knockdown animal model generated in our laboratory in order to provide a useful tool for studying laminopathies.Methods Two plasmids tagged to zebrafish lmna gene were designed based on morpholino oligonucleotides technology.Co-microinjected the plasmids into zebrafish embryos to knockdown lmna gene.Imagining and western blot detection were used to identify the mutants.Results Two different proteins, Lamin A/C, were expressed in the zebrafish embryos.Two plasmids lmna-MO and lmna-EGFP-pCS 2 + were generated and co-microinjected into embryos.The results of imagining and western blot showed that the expression of lmna gene was downregulated in the zebrafish embryos.Conclusions Lamin A/C are expressed in zebrafish.lmna gene can be knocked down by the injection of lmna-MO and lmna-EGFP-pCS 2 +.This new animal model may be a powerful tool for study on laminopathies.
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The treatments of shoulder joint subluxation following apoplexy with acupuncture, tuina, acupoint injection, and comprehensive methods in recent 10 years were reviewed. By comparing and analyzing the clinical effects of different methods, acupuncture combing with rehabilitation therapy is considered as a satisfactory one in treating subluxation following apoplexy.