Study On The Role Of Lgr4 Regulated Macrophage In Acute Liver Injury

Liver is the main distribution site of macrophages,it can be roughly divided into two groups: Kupffer cells(KCs)which can self renew and macrophages derived from peritoneal bone marrow.The KCs located in the liver sinusoids can remove microorganisms from the portal vein to maintain liver homeostasis.When the liver is damaged,macrophages derived from bone marrow and abdominal cavity are recruited in the liver.Studies have shown that liver macrophages can participate in the initial stage of liver disease and can affect the outcome of liver disease.Liver macrophages,as antigen presenting cells,aggravate or inhibit inflammation by producing different cytokines,chemokines and mediators.In addition,liver macrophages can also promote liver tissue repair by releasing transforming growth factor(TGF-β)and matrix metalloproteinases(MMP).As the most abundant group of immune cells in liver tissues,liver macrophages mainly sense extracellular ligands through cell membrane protein receptors,and activate downstream signaling pathways to regulate various physiological functions.G protein-coupled receptors(GPCRs)are the largest protein receptor family on the cell membrane.There are more than 800 types of GPCRs that have been discovered,most of which are for tumor diseases.Few studies are directed towards acute liver injury.Therefore,in order to explore the role of GPCR-related receptors in acute liver injury,we used CCL4 to induce an acute liver injury model,extracted liver macrophages,and screened out the(Leucine-rich repeat-containing G protein coupled receptor)family member Lgr4 in the acute liver injury model,the surface of liver macrophages was significantly upregulated.Based on this phenomenon,we constructed Lgr4 macrophage conditional knockout mice.By inducing a knockout mouse model,we found that after the macrophage Lgr4 receptor was knocked out,the acute liver injury state of mice was aggravated,and we verified this conclusion in pathology,serology and cytology.It has been reported in the literature that tumor necrosis factor(TNF-α)and interferon(IFN-γ)are the central link of acute liver injury.Subsequently,we used mouse bone marrow induced macrophages in vitro cell function experiments and found that under the stimulation of LPS,macrophages Lgr4 knockout promoted the secretion of macrophages TNF-α.At the same time,we detected that the level of IFN-γ in the serum of Lgr4 macrophage knockout acute liver injury model mice also increased.We have already known that IFN-γ is mainly derived from lymphocyte secretion.Therefore,we guessed whether the macrophage Lgr4 receptor knockout will affect the secretion of related chemokines.Subsequently,we confirmed this hypothesis by immunofluorescence staining technology,and by in vitro cell RT-q PCR experiments,it was detected that the macrophage Lgr4 receptor knockout promoted the secretion of CXCL9,CXCL10 and CXCL11,of which CXCL10 was the most obvious.In order to further prove that the macrophage Lgr4 receptor knockout promotes the expression of TNF-α and CXCL10,we extracted mouse bone marrow macrophages,stimulated with LPS and Lgr4 ligand Rspondin 1 and Rspondin 3 in vitro,and detected the expression of TNF-α and CXCL10.We found that Lgr4 ligand can effectively reverse the function of LPS to stimulate bone marrow macrophages,indicating that the downstream signal of Lgr4 receptor regulates inflammation inhibitory signals,and the knockout aggravates acute liver injury in mice.In summary,our results reveal that the macrophage Lgr4 receptor participates in the regulation of the secretion of TNF-α and IFN-γ in acute liver injury,thereby affecting the outcome of acute liver injury.Therefore,Lgr4 receptor on macrophage membrane is expected to become a target for drug therapy of acute liver injury.