Prostaglandin I2 synthase regulates macrophage polarization and promotes lymph node metastasis in colorectal cancer
Objective To explore the biological role of lipid metabolism genes in colorectal cancer(CRC)lymph node metastasis and its mechanisms.Methods First,8 cases of lymph node metastasis and non-metastatic CRC tissues were selected for high throughput sequencing,and public databases were com-bined to screen for lipid metabolism genes related to lymph node metastasis and prognosis,namely prosta-glandin I2 synthase(PTGIS).Subsequently,the relationship between PTGIS expression and lymph node metastasis was verified in 70 CRC tissues.Next,PTGIS was silenced in CRC cell lines using small interfer-ing RNA(siRNA),the cells were divided into transfected(siPTGIS)and control groups,and silencing ef-ficiency was verified by real-time fluorescent quantitative polymerase chain reaction(RT-qPCR)and Western blotting.Cell counting kit-8(CCK-8)assay,monoclonal assay and 5-ethynyl-2-deoxyuridine(EdU)staining were used to analyze the effect of silencing PTGIS on the proliferative ability of CRC.Tran-swell assay was used to analyze the effect of PTGIS silencing on the migratory and invasive ability of CRC cells.Nile green staining and Triglyceride quantification kit were used to detect the lipid level of the cells.Flow cytometry was used to analyze the effect of silencing PTGIS on the macrophage phenotypic changes of CRC cells co-cultured with THP-1 after 24 h of co-culture with THP.Finally,immunofluorescence(IF)was used to verify the relationship between PTGIS and the expression abundance of M1-type macrophages.T-test was used for comparisons between groups.Results Immunohistochemistry(IHC)showed higher ex-pression levels of PTGIS in CRC tissues with lymph node metastasis than in CRC tissues without lymph node metastasis(25.74±1.36 vs.5.10±0.57,t=13.96,P＜0.01).The proliferation,migration and invasion ability of CRC cells after PTGIS silencing was significantly lower than that of the control group(HCT8:CCK-8:0.64±0.01 vs.1.06±0.01,t=22.31,P＜0.01;EdU:35.20±0.92 vs.64.70± 1.62,t=15.75,P＜0.01;monoclonal assay:761.00±46.36 vs.1 672.00±166.70,t=5.26,P＜0.01;Transwell migration:53.00±1.73 vs.92.67±1.20,t=18.82,P＜0.01;Transwell invasion:27.67±1.20 vs.65.00±2.08,t=15.53,P＜0.01).The number of lipid droplets and triglyceride con-tent were significantly lower in the transfected group than in the control group(lipid droplets:0.39±0.01 vs.1.24±0.07,t=10.41,P＜0.01;triglycerides:74.50±1.50 vs.142.50±1.50,t=32.06,P＜0.01).The proportion of M1-type macrophages after co-culture of CRC cells with THP-1 was higher in the transfected group than in the control group(1.86±0.06 vs.1.00±0.00,t=13.97,P＜0.01).M1-type macrophage expression was higher in PTGIS low-expressing CRC tissues than in PTGIS high-expressing CRC tissues(1.17±0.01 vs.0.66±0.02,t=23.12,P＜0.01).Conclusion Silencing PTGIS inhib-its CRC cell metastasis and promotes macrophage M1 polarization.
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