Specificity protein 1/ZIP8/zinc axis reduced macrophage extracellular traps to alleviate severe acute pancreatitis in mice

Figure 5 Reduced ZIP8 expression in macrophages reduces zinc levels and is linked to enhanced macrophage extracellular trap formation; zinc supplementation can lower reactive oxygen species levels and suppress macrophage extracellular trap formation. A: A transcriptomic analysis was conducted to identify differentially expressed genes in the macrophages of the macrophage extracellular trap (MET)-induced group and negative control groups; B: The expression level of ZIP8 messenger RNA (mRNA) in bone marrow-derived macrophages (BMDMs) was measured using quantitative real-time polymerase chain reaction (q-PCR) (n = 3); C: Representative immunofluorescence images showing ZIP8 expression in BMDMs; D: After ZIP8 was overexpressed, ZIP8 mRNA expression in BMDMs was measured using q-PCR (n = 3); E: Zinc levels in BMDMs were assessed using a fluorescent zinc probe (n = 4); F: Representative fluorescence images of citrullinated histone H3 (CitH3) and F4/80 in macrophages, together with quantitative analysis of CitH3 fluorescence intensity (n = 4); G: Changes in the mRNA expression levels of peptidylarginine deiminase 4, nicotinamide adenine dinucleotide phosphate hydrogen oxidase 2, and toll-like receptor 2 in macrophages within the MET induction system, with or without ZIP8 overexpression, were measured by q-PCR (n = 4); H: Intracellular reactive oxygen species levels were measured using a 2’,7’-dichlorodihydrofluorescein diacetate fluorescent probe assay (n = 4). ^aP < 0.05. ^bP < 0.01. ^cP < 0.001. NS: Not significant; PMA: Phorbol 12-myristate 13-acetate; Nc: Negative control; CitH3: Citrullinated histone H3; BMDMs: Bone marrow-derived macrophages; DAPI: 4’,6-diamidino-2-phenylindole; mRNA: Messenger RNA; Oe: Overexpression; MFI: Mean fluorescence intensity; PAD4: Peptidylarginine deiminase 4; TLR2: Toll-like receptor 2; NOX2: Nicotinamide adenine dinucleotide phosphate hydrogen oxidase 2; ROS: Reactive oxygen species; DCFH-DA: 2’,7’-dichlorodihydrofluorescein diacetate.