The inflammatory bowel disease ulcerative colitis (UC) frequently progresses to colon cancer. To understand the mechanisms by which UC patients develop colon carcinomas, we used a mouse model of the disease whereby administration of azoxymethane (AOM) followed by repeated dextran sulfate sodium (DSS) ingestion causes severe colonic inflammation and the subsequent development of multiple tumors. We found that treating WT mice with AOM and DSS increased TNF-α expression and the number of infiltrating leukocytes expressing its major receptor, p55 (TNF-Rp55), in the lamina propria and submucosal regions of the colon. This was followed by the development of multiple colonic tumors. Mice lacking TNF-Rp55 and treated with AOM and DSS showed reduced mucosal damage, reduced infiltration of macrophages and neutrophils, and attenuated subsequent tumor formation. WT mice transplanted with TNF-Rp55-deficient bone marrow also developed significantly fewer tumors after AOM and DSS treatment than either WT mice or TNF-Rp55-deficient mice transplanted with WT bone marrow. Furthermore, administration of etanercept, a specific antagonist of TNF-α, to WT mice after treatment with AOM and DSS markedly reduced the number and size of tumors and reduced colonic infiltration by neutrophils and macrophages. These observations identify TNF-α as a crucial mediator of the initiation and progression of colitis-associated colon carcinogenesis and suggest that targeting TNF-α may be useful in treating colon cancer in individuals with UC.

Metastasis proceeds through interaction between cancer cells and resident cells such as leukocytes and fibroblasts. An i.v. injection of a mouse renal cell carcinoma, Renca, into wild-type mice resulted in multiple metastasis foci in lungs and was associated with intratumoral accumulation of macrophages, granulocytes, and fibroblasts. A chemokine, CCL3, was detected in infiltrating cells and, to a lesser degree, tumor cells, together with an infiltration of leukocytes expressing CCR5, a specific receptor for CCL3. A deficiency of the CCL3 or CCR5 gene markedly reduced the number of metastasis foci in the lung, and the analysis using bone marrow chimeric mice revealed that both bone marrow-and non-bone marrow-derived cells contributed to metastasis formation. CCL3-and CCR5-deficient mice exhibited a reduction in intratumoral accumulation of macrophages, granulocytes, and fibroblasts. Moreover, intratumoral neovascularization, an indispensable process for metastasis, was attenuated in these gene-deficient mice. Intrapulmonary expression of matrix metalloproteinase (MMP)-9 and hepatocyte growth factor (HGF) was enhanced in wild-type mice, and the increases were markedly diminished in CCL3-and CCR5-deficient mice. Furthermore, MMP-9 protein was detected in macrophages and granulocytes, the cells that also express CCR5 and in vitro stimulation by CCL3-induced macrophages to express MMP-9. Intratumoral fibroblasts expressed CCR5 and HGF protein. In vitro CCL3 stimulated fibroblasts to express HGF. Collectively, the CCL3-CCR5 axis appears to regulate intratumoral trafficking of leukocytes and fibroblasts, as well as MMP-9 and HGF expression, and as a consequence to accelerate neovascularization and subsequent metastasis formation.

Pro-inflammatory cytokines and chemokines are involved in promoting tumorigenesis by facilitating tumor proliferation and metastasis. The serum levels of interleukin (IL)-6, IL-1β β β β, and tumor necrosis factor-alpha (TNF-α α α α) are significantly elevated in patients with renal cell carcinoma (RCC). However, the mechanisms of how these cytokines participate in the progression of RCC remains unknown. In the present study, we investigated the effects of tumorderived cytokines on invasion and the epithelial-mesenchymal transition (EMT) of RCC cells. We found that expression of IL-1β β β β, IL-6, TNF-α α α α, hypoxia-inducible factor-alpha (HIF-1α α α α), and matrix metalloproteinase-2 (MMP2) were significantly elevated in high malignancy A498 (1) Four types of RCC have been delineated clinically and the genes responsible for them have been characterized. Overall, 75% of RCC are the clear cell type (ccRCC), while the other 25% are made up of the papillary, chromophobe, and oncocytic types. (2) Defects in the von Hippel-Lindau (VHL) gene appear to be responsible for 60% of sporadic ccRCC. A quarter of patients present with locally invasive or metastatic RCC. A third of the patients after resection of the tumor will have a recurrence. Moreover, systemic theraputic treatments of advanced RCC are largely ineffective and do not improve patient survival.(1,2) Therefore, defining the factors involved in disease progression and metastasis will provide molecular targets for the development of effective therapies.A complex network of pro-inflammatory cytokines, chemokines, and their receptors influences the development of primary tumors and metastasis. (3,4) CXCR4 is most commonly found in malignant cells from different cancer types.(4) In addition to CXCR4, CCR3 has been found to be up-regulated in 28% RCC tissues and correlates with a higher grade of malignancy.(5) Furthermore, the levels of IL-6, IL-1β, and TNF-α in serum are significantly higher and correlated with tumor size in RCC.(6) IL-6 blood level is significantly higher in patients with lymph node invasion and distant metastases, while TNF-α level is significantly higher as the stage of the RCC increases.(6,7) Therefore, IL-6 may be one of the factors associated with poor prognosis of patients with RCC. In addition, TNF-α may be useful as marker for the early diagnosis of RCC.(6) However, little is known about how these cytokines affect RCC migration and progression.The epithelial-mesenchymal transition (EMT) is a process in which polarized epithelial cells are converted into motile mesenchymal cells. Alterations in adhesion, morphology, cellular architecture, and migration capacity are the major events that occur during this process.(8) Common molecular markers for the EMT include increased expression of vimentin, nuclear localization of β-catenin, and increased production of transcriptional factors that inhibit E-cadherin production. Phenotypic markers for EMT comprise an increased capacity for migration and three-dimensional invasion as well as resistance t...

Macrophage infiltration is observed during the course of corneal neovascularization. We examined corneal neovascularization process after alkali injury in mice deficient in fractalkine receptor/CX3CR1, which is expressed by macrophages. The mRNA expression of CX3CR1 and its ligand CX3CL1/fractalkine was augmented together in the corneas after alkali injury. Compared with WT mice, CX3CR1‐deficient (KO) mice exhibited enhanced corneal neovascularization 2 weeks after injury, as evidenced by enlarged CD31‐positive areas. Concomitantly, the infiltration of F4/80‐positive macrophages but not Gr‐1‐positive neutrophils was significantly attenuated in KO mice comparing with WT mice. The intraocular mRNA expression of angiogenic factor, vascular endothelial factor, was enhanced to similar extents in WT and KO after the injury. In contrast, the mRNA expression of anti‐angiogenic factors, a disintegrin and metalloprotease with thrombospondin (ADAMTS)‐1 and thrombospondin (TSP)‐1 was enhanced in wild‐type mice but the enhancement was attenuated in KO mice. A double immunofluorescence analysis demonstrated that F4/80‐positive cells also expressed CX3CR1 and ADAMTS‐1, TSP‐1 and ADAMTS‐1 were detected in CX3CR1‐positive cells. CX3CL1 enhanced the ADAMTS‐1 and TSP‐1 expression by peritoneal macrophages. Moreover, topical application of CX3CL1 inhibited corneal neovascularization at 2 weeks, together with enhanced intraocular ADAMTS‐1 and TSP‐1 expression. Thus, these observations indicate that intraocular infiltrating CX3CR1‐positive macrophages can dampen alkali‐induced corneal neovascularization by producing anti‐angiogenic factors, such as ADAMTS‐1 and TSPs.

Chronic low dose of tumor necrosis factor-α (TNF-α) stimulation promotes tumorigenesis by facilitating tumor proliferation and metastasis. The plasma levels of TNF-α are increased in patients with renal cell carcinoma (RCC). Furthermore, high-grade clear cell RCC cell lines secrete more TNF-α than low-grade ones, and allow low-grade cell lines' gain of invasive ability. However, the molecular mechanism of TNF-α in mediating progression of RCC cells remains unclear. In the present study, TNF-α induced epithelial-mesenchymal transition (EMT) of RCC cells by repressing E-cadherin, promoting invasiveness and activating matrix metalloproteinase (MMP) 9 activity. RCC cells underwent promoted growth in vivo following stimulation with TNF-α. In addition, TNF-α induced phosphorylation of extracellular signal-regulated kinase, nuclear factor kappa B (NF-κB) and Akt in a time-dependent manner, and increased nuclear translocation and promoter activity of NF-κB. To investigate the role of NF-κB activation in TNF-α-induced EMT of RCC, we employed chemical inhibitors (NF-κB activation inhibitor and Bay 11-7082) and transfected dominant-negative (pCMV-IκBαM) and overexpressive (pFLAG-p65) vectors of NF-κB. While overexpression of NF-κB p65 alone could induce E-cadherin loss in RCC, EMT phenotypes and MMP9 expressions induced by TNF-α were not reversed by the inhibitors of NF-κB activation. These results suggest that the TNF-α signaling pathway is involved in the tumorigenesis of RCC. However, NF-κB activation is not crucial for invasion and EMT enhanced by TNF-α in RCC cells.