Mechanism Of Geniposide In Preventing Atherosclerosis By Improving Macrophage Autophagy

Background:Macrophage autophagy defect is closely related to the progression of atherosclerosis(AS).Triggering receptor expressed on myeloid cell 2(TREM2),an innate immune receptor expressed on the cell membrane of myeloid cells,such as microglia and macrophages,is a key factor in the development of Alzheimer’s disease,the deficiency of which leads to anomalous autophagy in microglia.However,the role of TREM2 in macrophage autophagy in atherosclerosis is still unclear.AS is characterized with phlegm,blood stasis,heat-toxin and deficiency from a perspective of traditional Chinese medicine.Among them,heat-toxin plays an important role in plaque progression and instability.Gardenia jasminoides Ellis is known as a heat-clearing and detoxifying herb.And its main component,geniposide(GP),can inhibit AS and enhance autophagy in cells of several different types;however,the underlying mechanism has not been fully understood.Methods:After 1 week of acclimatization,forty 8-weeks-old ApoE-/-mice and ten age-matched wild-type(WT)C57BL/6 mice were fed with high-fat diet(HFD)and normal diet for 24 weeks,respectively.After 12 weeks of HFD feeding,the ApoE-/mice were randomized into the GP(oral gavage,50 mg/kg/day,n=10),rapamycin(RA;intraperitoneal injection,2 mg/kg,twice a week,n=10),simvastatin(SV;oral gavage,5 mg/kg/day,n=10)and saline control(HFD;oral gavage once daily,n=10)groups;in the meantime,WT mice received oral administration of equivoluminal saline once daily.After 24 weeks,blood samples were collected for biochemical tests,the aortic arch were dissected for morphological analysis,and peritoneal macrophages were isolated to evaluate the level of TREM2 and autophagy signaling.RAW264.7 macrophages were incubated with different concentrations of geniposide,screening out appropriate concentration(40 μM)for following experiments.Trem2 was overexpressed by plasmid transfection(Trem2)or silenced by Trem2 siRNA transfection in macrophages,and RA was utilized to inhibit mTOR signaling,exploring the relationship among TREM2,macrophage autophagy and mTOR signaling and the effect of GP.Results:We observed upregulation of TREM2 and autophagy deficiency in the macrophages of atherosclerotic mice.GP inhibited the progression of AS in ApoE-/mice,decreased the levels of blood glucose,blood lipids and inflammatory cytokines,and suppressed the expression of TREM2.GP reinforced macrophage autophagy and suppressed mTOR signaling pathway.Trem2 overexpression in macrophages inhibited autophagy and activated mTOR signaling pathway;while Trem2 sliencing enhanced macrophage autophagy and suppressed mTOR signaling pathway.The effects of overexpressed Trem2 on macrophage autophagy were eliminated when mTOR signaling pathway was inhibited by RA.The effects of GP on macrophage autophagy and mTOR signaling pathway were weakened,but not eliminated,by the overexpression of Trem2.Conclusions:Our study indicates that TREM2 inhibits macrophage autophagy by activating mTOR signaling pathway,while augmenting the autophagy levels in macrophages by inhibiting the TREM2/mTOR axis can potentially impede atherosclerotic progression.GP prevents AS via enhancement of macrophage autophagy,which is partially dependent on the suppression of TREM2/mTOR axis.