Figure 1.
Figure 1. GMFG regulates iron metabolism protein expression in murine macrophages. (A-B) RAW264.7 macrophages were treated with FAC (A; 0-150 µM) or DFO (B; 0-150 µM) in 10% FBS/DMEM for 24 hours. Immunoblot analysis of GMFG in cellular lysates (upper). α-tubulin was used as a loading control. Relative quantification of GMFG protein expression levels from densitometric scans after normalizing to the control α-tubulin (lower graphs). (C) Quantitative polymerase chain reaction (qPCR) analysis of mean relative GMFG messenger RNA (mRNA) expression in FAC- or DFO-treated RAW264.7 macrophages. The data were normalized to 18S mRNA expression. (D) Immunoblot analysis of iron metabolism proteins in cellular lysates of RAW264.7 macrophages or BMDMs transfected with control siRNA (Ctrl) or GMFG siRNA for 48 hours. α-tubulin was used as a loading control. Graphs (right) show relative quantification of immunoblot (left). Protein levels from densitometric scans are normalized to the control α-tubulin and presented as fold change relative to control siRNA-transfected cells. (E) qPCR analysis of mean relative mRNA expression levels of iron metabolism proteins in RAW264.7 macrophages transfected with control siRNA or GMFG siRNA for 48 hours. The data were normalized to 18S mRNA expression. (F) Immunoblot analysis of iron metabolism proteins in cellular lysates of RAW264.7 macrophages transfected with control siRNA or GMFG siRNA for 48 hours, then treated with vehicle, FAC (50-100 µM), or DFO (100 µM) for 24 hours. α-tubulin was used as a loading control. Graphs (right) show relative quantification of immunoblot (left). Protein levels from densitometric scans are normalized to the control α-tubulin and presented as fold change relative to control siRNA-transfected cells before treatments. (G) Immunoblot analysis of iron metabolism proteins in cellular lysates of RAW264.7 macrophages transfected with control siRNA or GMFG siRNA for 48 hours, followed by cotransfection of GFP vector or GMFG-GFP plasmid for another 24 hours. α-tubulin was used as a loading control. Data represent the mean ± standard deviation of at least 3 independent experiments. *P < .05 compared with control untreated cells or control siRNA-transfected cells. a.u., arbitrary units.

GMFG regulates iron metabolism protein expression in murine macrophages. (A-B) RAW264.7 macrophages were treated with FAC (A; 0-150 µM) or DFO (B; 0-150 µM) in 10% FBS/DMEM for 24 hours. Immunoblot analysis of GMFG in cellular lysates (upper). α-tubulin was used as a loading control. Relative quantification of GMFG protein expression levels from densitometric scans after normalizing to the control α-tubulin (lower graphs). (C) Quantitative polymerase chain reaction (qPCR) analysis of mean relative GMFG messenger RNA (mRNA) expression in FAC- or DFO-treated RAW264.7 macrophages. The data were normalized to 18S mRNA expression. (D) Immunoblot analysis of iron metabolism proteins in cellular lysates of RAW264.7 macrophages or BMDMs transfected with control siRNA (Ctrl) or GMFG siRNA for 48 hours. α-tubulin was used as a loading control. Graphs (right) show relative quantification of immunoblot (left). Protein levels from densitometric scans are normalized to the control α-tubulin and presented as fold change relative to control siRNA-transfected cells. (E) qPCR analysis of mean relative mRNA expression levels of iron metabolism proteins in RAW264.7 macrophages transfected with control siRNA or GMFG siRNA for 48 hours. The data were normalized to 18S mRNA expression. (F) Immunoblot analysis of iron metabolism proteins in cellular lysates of RAW264.7 macrophages transfected with control siRNA or GMFG siRNA for 48 hours, then treated with vehicle, FAC (50-100 µM), or DFO (100 µM) for 24 hours. α-tubulin was used as a loading control. Graphs (right) show relative quantification of immunoblot (left). Protein levels from densitometric scans are normalized to the control α-tubulin and presented as fold change relative to control siRNA-transfected cells before treatments. (G) Immunoblot analysis of iron metabolism proteins in cellular lysates of RAW264.7 macrophages transfected with control siRNA or GMFG siRNA for 48 hours, followed by cotransfection of GFP vector or GMFG-GFP plasmid for another 24 hours. α-tubulin was used as a loading control. Data represent the mean ± standard deviation of at least 3 independent experiments. *P < .05 compared with control untreated cells or control siRNA-transfected cells. a.u., arbitrary units.

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