Study On The Mechanism Of ALOX15 Regulating Macrophage Ferroptosis Mediating Chronic Sinusitis With Nasal Polyps Type 2 Inflammatio

BACKGROUND:Ferroptosis,which is an iron-dependent programmed cell death characterized by lipid peroxidation,has been identified in recent years and plays an important role in tumor and inflammation diseases.However,the action mechanism of ferroptosis in chronic rhinosinusitis has not been reported.METHODS:Immunohistochemistry and Western blot were used to analyze the expression and distribution of ferroptosis regulatory molecules in various groups of clinical tissues.Immunofluorescence was used to verify the co-localization of molecules such as 15lipoxygenase(ALOX15)in macrophages.Transmission electron microscopy was performed to observe the morphology of macrophage mitochondria.Macrophage models derived from human peripheral blood mononuclear cells(PBMC)and THP-1 cell line were constructed.We induced macrophage ferroptosis by RSL3 and validated the regulatory role of ALOX15 on ferroptosis and inflammatory response in M2 type macrophages by using ALOX15 inhibitor PD 146176 and others.The signaling pathways regulated by ALOX15 on macrophages were explored by RNA-seq.The sensitivity of each subtype of macrophages to ferroptosis was analyzed by CCK-8 and LDH release.The regulation of macrophage lipid peroxidation by ALOX15 was analyzed using BODIPYTM 581/591 C11 and the effect of ferroptosis on macrophage CCL18 secretion levels was verified by ELISA.Results:1.eCRSwNP had high levels of ferroptosis driving factors such as ALOX15 and AcylCoA synthetase long-chain family member 4(ACSL4)and low levels of ferroptosis inhibitory molecules such as glutathione peroxidase 4(GPX4).2.eCRSwNP macrophages showed high expression of ALOX15,while transmission electron microscopy showed that eCRSwNP macrophages displayed morphological alterations about ferroptosis in mitochondria.3.RNA-seq results showed that ALOX15 affected the chemokine secretion function of macrophages,as well as the expression of ferroptosis-related molecules solute carrier family 7 member 11(SLC7A11)and prostaglandin endoperoxide synthase 2(PTGS2).4.M2 macrophages were more sensitive to ferroptosis induced by RSL3 than M0 and M1 macrophages,and inhibitors of ALOX15 were able to rescue the occurrence of ferroptosis in the cells.5.ALOX15 mediates ferroptosis in M2 macrophages by affecting lipid peroxidation levels,and ferroptosis can affect the secretion of C-C motif chemokine ligand 18(CCL18)in macrophages.Conclusion:There are molecular and morphological alterations related to ferroptosis in macrophages in eCRSwNP,M2-type macrophages are more sensitive to ferroptosis.ALOX15 can be involved in the ferroptosis process of M2 type macrophages through mediating lipid peroxidation,and ferroptosis could affect the ability of M2 macrophages to secrete CCL18.