Colorectal Cancer Cells-Fibroblasts Interaction Regulats Connexin43 And IGFBP-7 Expression In Colorectal Carcinoma

Background Colorectal cancer is one of the most common gastrointestinal malignancies that severely threaten the health of human. It is of great significance to study the underlying mechanisms of colorectal cancer for its prevention and treatment. Tumor microenvironment can significantly influence tumorigenesis. Tumour microenvironment includs extracellular matrix, blood vasculature, inflammatory cells and fibroblasts. In cancer-associated stroma, but not normal stroma, fibroblasts can be activated to myofibroblasts, also be called "cancer-associated"fibroblasts. α-SMA is a marker for"cancer-associated" fibroblasts. Several experimental systems have further shown that such "cancer-associated"fibroblasts can influence the tumorigenic properties of the epithelial compartment.Direct intercellular communication is mainly mediated by gap junctions, which is named as gap junctional intercellular communication(GJIC). GJIC has been implicated in the ability of a cell to regulate growth control via adaptive responses: differentiation, proliferation and apoptosis. Gap junctions are always abnormal or defective in tumor cells, leading to tumor cell uncontrolled growth. Gap junctions are made up of protein subunits termed connexins. Connexin43 protein (Cx43) is one of the most universally connexins in many tissues.Insulin-like growth factor binding protein 7 (IGFBP7) was screened from the cDNA subtraction library of colonic adenocarcinoma-normal mucosa by suppression subtractive hybridization (SSH) in our laboratory in 1999. IGFBP7 has been proposed as a potential tumor suppressor gene. However, it is of interest that the expression of IGFBP7 is quite different in some cancers. IGFBP7 expression was upregulated incolorectal cancers and small blood vessels in the tumor tissue, but lost in most coloncancer cell lines. This paradoxical phenomenon implied us: whether tumor-stromainteraction plays a role in regulating IGFBP7 expression?Objective To test the interaction between colorectal cancer cells and fibroblasts,further more study the variation in the two sorts cells, including fibroblasts activationand IGFBP7 expression in the co-culture system.Methods To simulate tumor microenvironment, we create a special model fortesting cell-cell interaction, two-layer cell spheroid. Colorectal cell line SW620 wasmade for the spheroid core and fibroblasts HELF for the outer. We tested Cx43expression in cell spheroid by IHC. We examined α-SMA and IGFBP7 expression inthe co-culture system by IF and RT-PCR.Results In the spheroid, the cells were morphologically well and the boundarybetween two sorts of cells was clear. We found Cx43 expression in the boundary betweenSW620 cells and HELF cells. In the SW620-HELF and SW620-HSF co-culturesystem, we found α-SMA remarkably highly expressed in the fibroblasts co-cultured withSW620. However, we haven't seen IGFBP7 expression in SW620-HELF co-culturesystem.Conclusions:1. Two-layer cell spheroid is a good model for cell-cell interaction.2. Colorectal tumor cells-fibroblasts interaction exists in colorectal cancer, and the interaction can induce Cx43 expression, restoring GJIC.3. Colorectal cancer cells-fibroblasts interaction is able to induce fibroblasts activation.4. SW620-HELF interaction can't induce IGFBP7 expression in the co-culture system.