黄芪建中汤调控巨噬细胞M1/M2极化改善癌症恶病质小鼠脂肪消耗OA

Objective:To investigate the effects of Huangqi Jianzhongtang(HQJZ)on macrophage polarization and fat consumption in cancer cachexia(CC)mice.Methods:Ultra-performance liquid chromatography-quadrupole/electrostatic field Orbitrap high-resolution mass spectrometry(UPLC-Q-Orbitrap HRMS)was used to control the quality of HQJZ.(1)In vitro experiment:HQJZ-containing serum was prepared,and the optimal concentration was determined by cytotoxicity assay.Mouse monocyte-derived macrophages(RAW264.7)were cultured and randomly divided into six groups,including a blank group,a classically activated macrophages(M1)group,an alternatively activated macrophages(M2)group,a HQJZ+blank group,a HQJZ+M1 group,and a HQJZ+M2 group.The relative expression of macrophage marker genes CD86,inducible nitric oxide synthase(iNOS),CD206,and arginase-1(Arg1)was detected by real-time quantitative polymerase chain reaction(Real-time PCR).(2)In vivo experiment:Thirty-two BALB/c mice were randomly divided into a control group,a model group,a medroxyprogesterone acetate(MPA)group,and a HQJZ group.Except for the control group,the other mice were injected with CT-26 colon cancer cells to establish a CC model.Mice in the MPA and HQJZ groups were given MPA(0.13 g·kg-1·d-1)or HQJZ(13.13 g·kg-1·d-1)by gavage,respectively,while mice in the control and model groups were given an equal volume of saline by gavage,with interventions continued for 10 d.Real-time PCR was used to detect the expression of macrophage markers(iNOS,Arg1,CD86,CD206)and fat browning-related genes uncoupling protein 1(UCP1)and peroxisome proliferator-activated receptor γ(PPARγ)in epididymal adipose tissue.Western blot(WB)was used to detect protein expression levels of UCP1 and PPARγ.Micro-computed tomography(micro-CT)was used to measure residual fat volume,and hematoxylin-eosin(HE)staining was used to assess fat browning and calculate pathological scores.Results:In vitro,the dominant effective concentration of HQJZ-containing serum was 12.5%.Real-time PCR results showed that,compared with the blank group,Arg1 expression decreased in the HQJZ+blank group(P<0.05),CD206 showed a downward trend without statistical significance,while iNOS and CD86 expression were significantly increased(P<0.05).Compared with the M1 group,Arg1 and CD206 expression decreased in the HQJZ+M1 group(P<0.05).Compared with the M2 group,CD206 expression decreased in the HQJZ+M2 group(P<0.05),CD86 expression increased significantly(P<0.01).In vivo,Real-time PCR results showed that,compared with the control group,CD86 and CD206 expression levels were significantly increased in the model group(P<0.01).Compared with the model group,CD206 expression in the MPA group was significantly decreased(P<0.01).In the HQJZ group,CD206 was significantly decreased(P<0.01).WB results showed that,compared with the model group,protein expression of UCP1 and PPARγ was significantly reduced in the HQJZ group(P<0.05,P<0.01).micro-CT results showed that the total white fat volume in the HQJZ group was greater than that in the model group(P<0.05).HE staining results showed that pathological scores in the HQJZ group were lower than those in the model group(P<0.05).Conclusion:HQJZ may inhibit white adipose tissue browning by promoting macrophage M1 polarization and suppressing M2 polarization,thereby delaying fat consumption in CC mice.