IgG expression in human colorectal cancer and its relationship to cancer cell behaviors

Abstract

Increasing evidence indicates that various cancer cell types are capable of producing IgG. The exact function of cancer-derived IgG has, however, not been elucidated. Here we demonstrated the expression of IgG genes with V(D)J recombination in 80 cases of colorectal cancers, 4 colon cancer cell lines and a tumor bearing immune deficient mouse model. IgG expression was associated with tumor differentiation, pTNM stage, lymph node involvement and inflammatory infiltration and positively correlated with the expressions of Cyclin D1, NF-κB and PCNA. Furthermore, we investigated the effect of cancer-derived IgG on the malignant behaviors of colorectal cancer cells and showed that blockage of IgG resulted in increased apoptosis and negatively affected the potential for anchor-independent colony formation and cancer cell invasion. These findings suggest that IgG synthesized by colorectal cancer cells is involved in the development and growth of colorectal cancer and blockage of IgG may be a potential therapy in treating this cancer.

Figure 1. Expression of IgG and mRNA in CRC tissues.

A: Co-expression of CEA, IgG protein and IGHG1 mRNA in CRC cells. B–D: Expression of IgG (red signals) in normal colon tissue (B), well differentiated (C) and poorly differentiated (D) CRC. E: Expression of IgG in cells of cancer nests (arrows) infiltrating into deep muscle layer of the colon wall. The expression level of IgG is higher in infiltrating cancer cells (E) than in the cancer cells of the “primary” tumor shown in D. F: CRC nests (CRC) and lymph node (LN) before and after LCM. G: Agarose gel electrophoresis of RT-PCR amplification of microdissected CRC cells and lymph nodes. H: Co-localization of IgG protein expressed in the cytoplasm (red signal) and Cyclin D1 (left panel), NF-κB (middle panel) and PCNA with nuclear localization (right panel) (purple-blue signals). ML: Muscle layer. Bars = 50 µm (A–E, G), 20 µm (F).

Figure 2. Expression of IgG protein and mRNA in human CRC cell lines.

A: Igγ and Igκ are detected in all four CRC lines by Double staining immunofluorescence (Igγ, TRITC; Igκ, FITC; Merge, yellow.). Bars = 50 µm. B: Percentage of IgG positive cells in Raji, HT29, SW480, and LOVO. C: Agarose gel electrophoresis of RT-PCR amplification products. R, DNase treated RNA as template; C, cDNA as template. D: IgG detection by Western blot in all four cell lines. E: IgG protein and mRNA are detected with IHC and ISH in tumors harvested on 21 days of LOVO cells in SCID mice. IgG is detectable in the sera at day 7, 14, and 21.

Figure 3. Blockage of cancer-derived IgG by anti-human IgG antibody inhibits biological behaviors of CRC cells.

A: MTS with a series of concentrations of anti-human IgG (10–1000 nM) antibody. B: Treatment with rabbit anti-mouse IgG increases the apoptosis rates in LOVO cells. C: Apoptotic percentage of LOVO cells is higher after incubation with IgG antibody than with IgM antibody or mouse serum. D: Blockage of cancer-derived IgG results in morphological changes. E: IgG antibody decreases malignant anchor-independent growth of CRC cells. F: Blockade of cancer-derived IgG suppresses the invasion potential of CRC cells. Bars = 50 µm.