ABSTRACT
Tumor necrosis factor-like weak inducer of apoptosis (TWEAK), a member of the tumor necrosis factor (TNF) family of cytokines, acts on responsive cells via binding to a cell surface receptor called Fn14. TWEAK binding to an Fn14 receptor or constitutive Fn14 overexpression has been shown to activate nuclear factor κB signaling which is important in tumorigenesis and cancer therapy resistance. In the present study, we demonstrate that TWEAK and Fn14 are expressed in neuroblastoma cell lines and primary tumors, and both are observed at increased levels in high-stage tumors. The treatment of neuroblastoma cell lines with recombinant TWEAK in vitro causes increased survival, and this effect is partially due to the activation of NF-κB signaling. Moreover, TWEAK induces the release of matrix metalloprotease-9 (MMP-9) in neuroblastoma cells, suggesting that TWEAK may play a role in the invasive phase of neuroblastoma tumorigenesis. TWEAK-induced cell survival was significantly reduced by silencing the TWEAK and Fn14 gene functions by siRNA. Thus, the expression of TWEAK and Fn14 in neuroblastoma suggests that TWEAK functions as an important regulator of primary neuroblastoma growth, invasion and survival and that the therapeutic intervention of the TWEAK/Fn14 pathway may be an important clinical strategy in neuroblastoma therapy.
Subject(s)
Cell Transformation, Neoplastic/metabolism , Neuroblastoma/metabolism , Receptors, Tumor Necrosis Factor/metabolism , Tumor Necrosis Factors/metabolism , Cell Line, Tumor , Cell Nucleus/metabolism , Cell Survival , Child , Child, Preschool , Cytokine TWEAK , Enzyme Precursors/metabolism , Female , Gene Expression , Humans , Infant , Infant, Newborn , Male , Matrix Metalloproteinase 2/metabolism , Matrix Metalloproteinase 9 , NF-kappa B/metabolism , Neuroblastoma/pathology , Protein Transport , TWEAK ReceptorABSTRACT
BACKGROUND: Osteoprotegerin (OPG), a soluble member of the tumor necrosis factor family, is produced by various cell types and tissues and plays a key role in the physiological regulation of osteoclast differentiation and activity. Also, OPG is a soluble decoy receptor for tumor necrosis factor-related apoptosis-inducing factor (TRAIL). In the present study we investigated whether the human colon cancer cell lines HT-29 and SW-480 produce and secrete OPG in vitro. MATERIALS AND METHODS: Expression of OPG mRNA was examined by RT-PCR. OPG protein was analysed by ELISA assay and immunostaining methods. The effect of OPG secretion on TRAIL-mediated apoptosis was also investigated. RESULTS: By RT-PCR, it was demonstrated that mRNA transcripts for OPG were produced by both cell lines. By ELISA analysis, OPG was detected in the culture medium; and treatment of cells with proinflammatory cytokines TNF-alpha and IL-1beta increased OPG secretion significantly. Tumor xenografts in nude mice also were shown to express OPG by immunohistochemistry. When RANKL, which selectively binds OPG, was added to cell cultures along with recombinant TRAIL, apoptosis was shown to increase significantly. CONCLUSION: These data indicate that OPG may be involved in tumorigenesis and the progression of colon cancer.
Subject(s)
Colonic Neoplasms/metabolism , Glycoproteins/biosynthesis , Receptors, Cytoplasmic and Nuclear/biosynthesis , Receptors, Tumor Necrosis Factor/biosynthesis , Animals , Apoptosis/drug effects , Apoptosis Regulatory Proteins/pharmacology , Carrier Proteins/pharmacology , Cell Line, Tumor , Colonic Neoplasms/genetics , Glycoproteins/genetics , Glycoproteins/metabolism , HT29 Cells , Humans , Interleukin-1/pharmacology , Membrane Glycoproteins/pharmacology , Mice , Mice, Inbred BALB C , Mice, Nude , Osteoprotegerin , RANK Ligand , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Receptor Activator of Nuclear Factor-kappa B , Receptors, Cytoplasmic and Nuclear/genetics , Receptors, Tumor Necrosis Factor/genetics , Reverse Transcriptase Polymerase Chain Reaction , TNF-Related Apoptosis-Inducing Ligand , Tumor Necrosis Factor-alpha/pharmacologyABSTRACT
Two innovative 20-S-camptothecin (CPT) formulations, previously found suitable to achieve therapeutically relevant CPT concentrations, were assessed for their in vitro cytotoxic potential as compared to an aqueous CPT solution, using the MTT assay. The formulations, cationic CPT-containing liposomes (CPT-Lip), hydroxypropyl-beta-cyclodextrin (HP-beta-CD) complexed CPT (CPT-CD) and a saturated aqueous CPT solution (CPT-Sol), were diluted in culture medium to appropriate CPT concentrations (4.7-300 ng/ml), and incubated with HT-29 and SW-480 human colon carcinoma cell lines. IC50 values were calculated after 48 and 72 h incubation for the HT-29 and SW-480 cell lines, respectively, and were found to be of the same magnitude for all formulations, with only a slight difference (CPT-SolSubject(s)
Camptothecin/chemistry
, Camptothecin/toxicity
, Colonic Neoplasms/pathology
, Chemistry, Pharmaceutical
, Colonic Neoplasms/drug therapy
, Colonic Neoplasms/metabolism
, Dose-Response Relationship, Drug
, Drug Screening Assays, Antitumor/methods
, HT29 Cells
, Humans
