ABSTRACT
Notch signaling has been shown to be important in osteoblast differentiation. Therapeutic radiation has been shown to alter the skeletal system, yet little information is available on the changes in Notch signaling in irradiated osteoblasts. The purpose of this study was to analyze the effect of radiation therapy with 2 and 4 Gy on Notch signaling in osteoblasts. In order to assess the radiation damage on osteoblast differentiation, total RNA and protein were collected three days after exposure to radiation. The effects of radiation on Notch signaling at the early and terminal stages of osteoblastic MC3T3-E1 cell differentiation was analyzed by qRT-PCR and western blot analysis. Our study applied a previously established method to induce MC3T3-E1 cell differentiation into osteoblasts and osteoblast precursors. Our results showed that the expression of Notch receptors (Notch1-4), ligands (Jagged1, Jagged2 and Delta1), target of Notch signaling (Hes1) and markers (ALP, M-CSF, RANKL and OPG) were altered following 2 and 4 Gy of irradiation. The present research did not indicate a strong relationship between Notch1 regulation and suppression of osteoblast differentiation. We found Hes1 may play a role in the radiation effect on osteoblast differentiation. Our results indicate that radiated osteoblast precursors and osteoblasts promoted osteoclast differentiation and proliferation.
Subject(s)
Osteoblasts/radiation effects , Receptors, Notch/radiation effects , Signal Transduction/radiation effects , Alkaline Phosphatase/biosynthesis , Alkaline Phosphatase/radiation effects , Animals , Basic Helix-Loop-Helix Transcription Factors/biosynthesis , Basic Helix-Loop-Helix Transcription Factors/radiation effects , Calcium-Binding Proteins/biosynthesis , Calcium-Binding Proteins/radiation effects , Cell Differentiation/radiation effects , Cell Line , Gamma Rays/therapeutic use , Homeodomain Proteins/biosynthesis , Homeodomain Proteins/radiation effects , Intercellular Signaling Peptides and Proteins/biosynthesis , Intercellular Signaling Peptides and Proteins/radiation effects , Intracellular Signaling Peptides and Proteins/biosynthesis , Intracellular Signaling Peptides and Proteins/radiation effects , Jagged-1 Protein , Jagged-2 Protein , Macrophage Colony-Stimulating Factor/biosynthesis , Macrophage Colony-Stimulating Factor/radiation effects , Membrane Proteins/biosynthesis , Membrane Proteins/radiation effects , Mice , Osteoblasts/cytology , Osteoblasts/metabolism , Osteoprotegerin/biosynthesis , Osteoprotegerin/radiation effects , RANK Ligand/biosynthesis , RANK Ligand/radiation effects , Receptors, Notch/metabolism , Serrate-Jagged Proteins , Transcription Factor HES-1ABSTRACT
Ever since the skin was recognized as a site of immunologic activity, a number of laboratories have studied the production of cytokines by skin cells and the effects of chemicals, allergens, contact irritants and UVB radiation on their production. Most research to date has been carried out using either purified populations of primary cells, or established cell lines. Cytokines, however, do not function in isolation but they appear in human tissues within the context of other cytokines that can, in turn, strongly influence the final biological outcome. Therefore, in vivo studies using whole skin are more physiologically relevant since all cell types are present and interactions among them are allowed to proceed. We report here the results of a detailed timecourse study using whole mouse skin, consisting of both dermis and epidermis, irradiated with either a low or high dose of UVB and analyzed using a Multi-probe RNase protection assay system. The results show that in whole skin the kinetics of cytokine induction are different than what was previously observed in tissue culture. In addition to already known skin-associated cytokines, we report here the presence and UVB induction of cytokines not previously reported.
Subject(s)
Cytokines/radiation effects , Skin/radiation effects , Ultraviolet Rays/classification , Animals , Autoradiography , Cytokines/genetics , Cytokines/immunology , Dermis/immunology , Dermis/radiation effects , Epidermis/immunology , Epidermis/radiation effects , Female , Growth Inhibitors/radiation effects , Humans , Interleukin-1/radiation effects , Interleukin-10/radiation effects , Interleukin-11/radiation effects , Interleukin-2/radiation effects , Interleukin-6/radiation effects , Leukemia Inhibitory Factor , Lymphokines/radiation effects , Lymphotoxin-alpha/radiation effects , Macrophage Colony-Stimulating Factor/radiation effects , Macrophage Migration-Inhibitory Factors/radiation effects , Mice , Mice, Inbred BALB C , RNA Probes , Radiation Dosage , Ribonucleases , Skin/immunology , Time Factors , Transforming Growth Factor beta/radiation effectsABSTRACT
Experiments were conducted to explore the possible effect of low-dose irradiation on cytokine production in mice. SJL/J and C57BL/6J mice were exposed to 3 Gy and 4 Gy respectively. At various time intervals thereafter, lung-conditioned media, bone-marrow-conditioned media and sera were collected. Interleukin-6 and macrophage-colony-stimulating-factor activities were tested. Interleukin-6 in lung-conditioned media from both strains was found to be significantly induced by irradiation. Macrophage-colony-stimulating-factor activity was greatly enhanced in sera of irradiated SJL/J mice as well as in bone-marrow-conditioned media of both strains. The kinetics of the radiation-induced interleukin 6 and macrophage-colony-stimulating-factor activity differed in the 2 strains. SJL/J and C57BL/6J mice have been previously shown to be susceptible to low-dose-radiation-induced leukemia. The coleukemogenic effects of both cytokines in 3 different models of experimental leukemogenesis is demonstrated.
