Melatonin Suppresses Macrophage M1 Polarization and ROS-Mediated Pyroptosis via Activating ApoE/LDLR Pathway in Influenza A-Induced Acute Lung Injury

Figure 8

re-ApoE3 remedied the loss of regulatory ability of melatonin on macrophage polarization and oxidative injury. (a) Western blot images of the expression of ApoE, LDLR, iNOS, and Arg1 to β-actin in wild-type (WT) and ApoE-/- BMDMs infected by influenza A (H3N2) (, 12 h) with/without melatonin pretreatment or combined pretreatment of melatonin and recombinant ApoE3 (10 μg/ml, 3 h before H3N2 infection). (b) Representative immunofluorescence images of F4/80 (red), iNOS (green), and Arg1 (green) expression in WT and ApoE-/- BMDMs (original magnification ×400). Quantitative RT-PCR measurement of the relative mRNA levels of TNF-α (c), MCP1 (d), CD86 (e), Arg1 (f), Fizz1 (g), CD206 (h), and IL-1β (j) in WT and ApoE-/- BMDMs. (i) Intracellular ROS levels were quantificationally detected by DCFH-DA using a fluorescence plate reader in WT and ApoE-/- BMDMs. (k) Western blot images of the expression of NLRP3, Caspase1, and GSDMD-N to β-actin in WT and ApoE-/- BMDMs. (l) The lactic dehydrogenase (LDH) released into the medium was assessed based on OD₄₉₀ values of LDH release in WT and ApoE-/- BMDMs. Data expressed as (). , , and compared with influenza A- (H3N2-) infected WT or ApoE-/- BMDMs.

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