FOXP3 expression and clinical characteristics of hepatocellular carcinoma

Figure 1 Expression of forkhead box protein 3 in hepatoma cell lines. A: Expression of forkhead box protein 3 (FOXP3) mRNA is different in hepatoma cell lines by reverse-transcription polymerase chain reaction. β-actin was used to verify the integrity of the template cDNA preparations. 1: SMMC-7721, 2: Hepa-G2, 3: Melanocytes, 4: Foxp3-plasmid; B: Expression of FOXP3 protein was different in hepatoma cell lines by Western blotting. Melanocytes served as a negative control. Foxp3 transiently transferred 293 cells were established as a positive control. 1: SMMC-7721, 2: Hepa-G2, 3: Melanocytes, 4: Foxp3/293 cells.

Figure 2 Flow cytometry of forkhead box protein 3 expression in various hepatoma cell lines. Melanocytes served as a negative control. The grey underlaid plot represents staining with the isotype, and the white underlaid plot represents staining with anti-human forkhead box protein 3 antibody. FITC: Fluorescein isothiocyanate.

Figure 3 Double-label immunofluorescence analysis of forkhead box protein 3 expression in hepatoma cell line. A: SMMC-7721 cells express forkhead box protein 3 (FOXP3) in both nuclei and perinuclear cytoplasm (imaged with green fluorescent) (A1: Melanocytes FOXP3-FITC, A2: SMMC-7721 FOXP3-FITC); B: Imaged with diamidino-phenyl-indole (DAPI) to identify the nuclei of SMMC-7721 and melanocytes (B1: Melanocytes DAPI, B2: SMMC-7721 DAPI); C: Image superimposed on a differential interference contrast background confirmed colocalization (C1: Melanocytes merge, C2: SMMC-7721 merge).

Figure 4 Hepatocellular carcinoma immunohistochemical score (× 100). Forkhead box protein 3 (FOXP3) expression in hepatocellular carcinoma (HCC) by immunohistochemical (IHC). Images represent HCC tissues with IHC scores of 0 (A), 1 (B), 2 (C) and 3 (D). In IHC score 0, many FOXP3-positive lymphocytes are tumor-infiltrating Tregs.

Figure 5 Hepatocellular carcinoma immunohistochemical staining (× 200). Immunohistochemical (IHC) staining of paraffin embedded hepatocellular carcinoma (HCC) tissues revealed different levels of forkhead box protein 3 (FOXP3) expression in HCC cells (PCH101 antibody, eBioscience). A: Well-differentiated HCC (nuclear FOXP3 staining); B: Moderately-differentiated HCC (cytoplasmic FOXP3 staining); C: Poorly-differentiated HCC (cytoplasmic FOXP3 staining); D: Normal liver tissue (nuclear FOXP3 staining in Tregs).