Fig 1: STOM promotes oxidative stress and inflammation in LPS-induced MLE-12 cells. Detection of CD36 (A) protein and (B) mRNA levels in MLE-12 cells transfected with Ov-NC and Ov-CD36. Detection of (C) LDH, (D) MDA, (E) SOD and (F) ROS levels in LPS-induced MLE-12 cells transfected with Ov-NC and Ov-CD36. Detection of TNF-α and IL-6 (G) protein and (H) mRNA levels in MLE-12 cells. The results are representative of at least three independent experiments. ***P<0.001 vs. Control group; ###P<0.001 vs. LPS + si-NC group; $P<0.05, $$P<0.01, $$$P<0.001 vs. LPS + si-STOM + Ov-NC group. LPS, lipopolysaccharide; Ov-, overexpression; NC, negative control; LDH, lactate dehydrogenase; MDA, malondialdehyde; SOD, superoxide dismutase; ROS, reactive oxygen species; si-, small interfering RNA; STOM, stomatin.
Fig 2: STOM expression in LPS-induced MLE-12 cells. (A) Gene expression of STOM was analyzed using the Gene Expression Omnibus database. ***P<0.001. Detection of STOM (B) protein and (C) mRNA expression levels in MLE-12 cells. (D) Detection of STOM protein and (E) mRNA expression levels in MLE-12 cells transfected with siRNA. (F) A Cell Counting Kit-8 assay was performed to determine cell viability. The results are representative of at least three independent experiments. **P<0.01, ***P<0.001 vs. Control group; #P<0.05 vs. LPS + si-NC group. LPS, lipopolysaccharide; siRNA/si-, small interfering RNA; NC, negative control; STOM, stomatin.
Fig 3: STOM-knockdown reduces oxidative stress and inflammation in LPS-induced MLE-12 cells. MLE-12 cells were treated with LPS, followed by transfection with si-NC and si-STOM. Detection of (A) LDH, (B) MDA, (C) SOD and (D) ROS levels in MLE-12 cells. Detection of TNF-α and IL-6 (E) protein expression levels and (F) content levels in MLE-12 cells. The results are representative of at least three independent experiments. ***P<0.001 vs. Control group; ##P<0.01, ###P<0.001 vs. LPS + si-NC group. LPS, lipopolysaccharide; si-, small interfering RNA; NC, negative control; LDH, lactate dehydrogenase; MDA, malondialdehyde; SOD, superoxide dismutase; ROS, reactive oxygen species; STOM, stomatin.
Fig 4: STOM positively regulates CD36. (A) Association between STOM and CD36 was predicted using the Search Tool for the Retrieval of Interacting Genes/Proteins database. (B) Detection of CD36 protein levels in MLE-12 cells transfected with si-STOM. **P<0.01 vs. Control group. (C) An RNA immunoprecipitation assay was conducted to determine the interaction between STOM and CD36. The results are representative of at least three independent experiments. ***P<0.001 vs. Anti-IgG group; ##P<0.01 vs. Anti-CD36 group. si-, small interfering RNA; STOM, stomatin; NC, negative control.
Fig 5: Enrichment of sphingolipids in lipid raft microdomains. (A) RAW264.7 and Stom−/− macrophages were infected with CFW-labeled A. fumigatus wild-type or pksP conidia (CFW labeled; blue) and stained for GM1 ganglioside (CTB-Alexa Fluor 647; red); white arrows indicate phagosomes. (B) Percentage of GM1-positive phagosomes in RAW264.7 wild-type and Stom−/− macrophages after infection with A. fumigatus wild-type or pksP conidia.
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