Low-dose Adropin Stimulates Inflammasome Activation Of Macrophage Via Mitochondrial ROS To Suppress Colorectal Cancer Progression

Background:Adropin is encoded by an energy homeostasis related(ENHO)gene and widely exists in liver,pancreas,heart,kidney,brain,and vascular tissues.Abnormal expression of Adropin is associated with metabolic,inflammatory,immune,and central nervous system disorders.Gprotein-coupled receptor19(GPR19)is widely expressed in tumor tissues as a receptor for Adropin.Colorectal cancer(CRC)has the third highest incidence rate in the world.At present,the treatment for CRC includes surgery,chemotherapy,radiotherapy,targeted and immunotherapy.However,the 5-year survival rate of patients is only 56.9%.The recurrence and metastasis of CRC are still difficult in clinical treatment.Tumor associated macrophages(TAM),which account for 30%to 70%of tumor stromal cells,utilize the energy provided by fatty acid oxidation to participate in the progression of cancer.It is unclear whether Adropin and its receptor GPR19 are involved in the development of colorectal cancer(CRC).Objective:1.To investigate whether there is a decrease in the expression of Adropin in colorectal cancer samples;To study the expression level and change trend of Adropin in primary and adjacent tissues of colorectal cancer at different stages;To study whether the expression level of Adropin in colorectal cancer of different stages is related to the level of macrophage infiltration;2.To study the effect of ectopic expression of Adropin on tumor;3.To study the changes of inflammatory corpuscles stimulated by macrophages treated with low-dose and high-dose Adropin in vitro and the polarization trend of macrophages in the absence of Adropin in ENHO knockout mice;4.To study the mechanism of Adropin stimulating the activation of inflammatory corpuscles.Methods:Firstly,through bioinformatics analysis and immunohistochemical analysis of colorectal cancer tissues from 68 patients,the expression of Adropin and its receptor GPR19 in adjacent and cancerous tissues and their correlation with immune infiltration were analyzed.At the same time,the colocalization of Adropin and tumor related macrophages in the cancer stroma was investigated by immunofluorescence;Subsequently,the research team analyzed whether Adropin affects tumor growth by regulating macrophage activity through in vivo tumor bearing experiments;Next,the effect of recombinant Adropin protein on macrophage activity was detected through in vitro experiments and the macrophage activity in ENHO-/-mice was analyzed;Finally,the expression of immune related indicators in macrophages treated with different doses of recombinant Adropin protein,WT mice,and ENHO-/-mice was analyzed to explore the effect of Adropin on inflammatory bodies and its preliminary mechanism.Results:1.In patients with colorectal cancer,the expression of Adropin in cancer nests decreased,and there was a negative correlation between the expression of Adropin in cancer nests and the infiltration of macrophages;The expression of Adropin in the cancer stroma is increased,and there is a positive correlation between the expression of Adropin in the cancer stroma and the infiltration of macrophages.2.After overexpression of Adropin in colon cancer(MC38)cells,tumor growth in vivo is inhibited,accompanied by an increase in M1 macrophages,and a decrease in M2 cells;3.Low dose of Adropin(<100 ng/mL)acting on macrophages in vitro can directly increase the mitochondrial reactive oxygen species required for the activation of inflammatory bodies.In addition,there are fewer M1 macrophages in ENHO-/-mice,and the macrophages of ENHO-/-mice are not easily induced into M1 subgroups in vitro;4.Glucose transporter 1(GLUT 1)and hexokinase 2(HK2),related indicators of glucose metabolism,were upregulated by low dose of Adropin(<100 ng/mL).Carnitine Palmitoyltransferase lα(CPT1α)as an indicator of lipid metabolism is only upregulated at high doses of Adropin(>100ng/mL).When Adropin stimulated macrophages are cleared of intracellular ROS,NLRP3,IL-1β and the expression of n-GSDMD was significantly reduced.Conclusion:The change of Adropin level in tumor cells or macrophages has different relationship with the progression of colorectal cancer.Low dose of Adropin stimulated the antitumor activity of macrophages,while high dose of Adropin promoted the tumorigenic activity of macrophages.Increasing or decreasing the level of Adropin can inhibit tumor progression in different CRC stages.